InteractiveFly: GeneBrief

Kinesin-like protein at 61F: Biological Overview | References

Gene name - Kinesin-like protein at 61F

Synonyms -

Cytological map position - 61F4-61F4

Function - Kinesin motor protein

Keywords - mitotic spindle assembly, cytoskeleton, microtubule antiparallel orientation preference, microtubule crosslinking and sliding activities, pole-pole separation

Symbol - Klp61F

FlyBase ID: FBgn0004378

Genetic map position - 3L:1,241,568..1,245,939 [-]

Classification - KISc_BimC_Eg5, Kinesin motor domain

Cellular location - mitotic spindle

NCBI link: EntrezGene
Klp61F orthologs: Biolitmine
Recent literature
Brust-Mascher, I., Civelekoglu-Scholey, G. and Scholey, J. M. (2015). Mechanism for anaphase B: Evaluation of "slide-and-cluster" versus "slide-and-flux-or-elongate" models. Biophys J 108: 2007-2018. PubMed ID: 25902440
Elongation of the mitotic spindle during anaphase B contributes to chromosome segregation in many cells. This study quantitatively tested the ability of two models for spindle length control to describe the dynamics of anaphase B spindle elongation using experimental data from Drosophila embryos. In the slide-and-flux-or-elongate (SAFE) model, kinesin-5 motors persistently slide apart antiparallel interpolar microtubules (ipMTs). During pre-anaphase B, this outward sliding of ipMTs is balanced by depolymerization of their minus ends at the poles, producing poleward flux, while the spindle maintains a constant length. Following cyclin B degradation, ipMT depolymerization ceases so the sliding ipMTs can push the poles apart. The competing slide-and-cluster (SAC) model proposes that MTs nucleated at the equator are slid outward by the cooperative actions of the bipolar kinesin-5 and a minus-end-directed motor, which then pulls the sliding MTs inward and clusters them at the poles. In assessing both models, it is assumed that kinesin-5 preferentially cross-links and slides apart antiparallel MTs while the MT plus ends exhibit dynamic instability. However, in the SAC model, minus-end-directed motors bind the minus ends of MTs as cargo and transport them poleward along adjacent, parallel MT tracks, whereas in the SAFE model, all MT minus ends that reach the pole are depolymerized by kinesin-13. Remarkably, the results show that within a narrow range of MT dynamic instability parameters, both models can reproduce the steady-state length and dynamics of pre-anaphase B spindles and the rate of anaphase B spindle elongation. However, only the SAFE model reproduces the change in MT dynamics observed experimentally at anaphase B onset. Thus, although both models explain many features of anaphase B in this system, the quantitative evaluation of experimental data regarding several different aspects of spindle dynamics suggests that the SAFE model provides a better fit.

Bouge, A. L. and Parmentier, M. L. (2016). Tau excess impairs mitosis and kinesin-5 function, leading to aneuploidy and cell death. Dis Model Mech [Epub ahead of print]. PubMed ID: 26822478
In neurodegenerative diseases like Alzheimer's disease (AD), cell cycle defects and associated aneuploidy have been described. However, the importance of these defects in the physiopathology of AD and the underlying mechanistic processes are largely unknown in particular with respect to the microtubule-binding protein Tau, which is found in excess in the brain and cerebrospinal fluid of patients. Although it has long been known that Tau is phosphorylated during mitosis to generate a lower affinity for microtubules, there has been no indication that an excess of this protein could affect mitosis. The effect of an excess of human Tau (hTau) protein on cell mitosis was studied in vivo. Using the Drosophila developing wing disc epithelium as a model, this study shows that an excess of hTau induces a mitotic arrest, with the presence of monopolar spindles. This mitotic defect leads to aneuploidy and apoptotic cell death. The mechanism of action of hTau was studied and it was found that the MT-binding domain of hTau is responsible for these defects. hTau effects occur via the inhibition of the function of the kinesin Klp61F, the Drosophila homologue of kinesin-5 (also called Eg5 or KIF11). This deleterious effect of hTau is also found in other Drosophila cell types (neuroblasts) and tissues (the developing eye disc) as well as in human Hela cells.By demonstrating that microtubule-bound Tau inhibits the Eg5/KIF11 kinesin and cell mitosis, this work provides a new framework to consider the role of Tau in neurodegenerative diseases.

Ye, A. A. and Maresca, T. J. (2016). Generating a "humanized" Drosophila S2 cell line sensitive to pharmacological inhibition of Kinesin-5. J Vis Exp [Epub ahead of print]. PubMed ID: 26863489

Kinetochores are large protein-based structures that assemble on centromeres during cell division and link chromosomes to spindle microtubules. Proper distribution of the genetic material requires that sister kinetochores on every chromosome become bioriented by attaching to microtubules from opposite spindle poles before progressing into anaphase. However, erroneous, non-bioriented attachment states are common and cellular pathways exist to both detect and correct such attachments during cell division. The process by which improper kinetochore-microtubule interactions are destabilized is referred to as error correction. To study error correction in living cells, incorrect attachments are purposely generated via chemical inhibition of kinesin-5 motor, which leads to monopolar spindle assembly, and the transition from mal-orientation to biorientation is observed following drug washout. The large number of chromosomes in many model tissue culture cell types poses a challenge in observing individual error correction events. Drosophila S2 cells are better subjects for such studies as they possess as few as 4 pairs of chromosomes. However, small molecule kinesin-5 inhibitors are ineffective against Drosophila kinesin-5 (Klp61F). This study describes how to build a Drosophila cell line that effectively replaces Klp61F with human kinesin-5, which renders the cells sensitive to pharmacological inhibition of the motor and suitable for use in the cell-based error correction assay (Ye, 2016).

Radford, S. J., Go, A. M. and McKim, K. S. (2016). Cooperation between kinesin motors promotes spindle symmetry and chromosome organization in oocytes. Genetics [Epub ahead of print]. PubMed ID: 27932541
The oocyte spindle in most animal species is assembled in the absence of the microtubule-organizing centers called centrosomes. Without the organization provided by centrosomes, acentrosomal meiotic spindle organization may rely heavily on the bundling of microtubules by kinesin motor proteins. Indeed, the minus-end directed kinesin-14 NCD and the plus-end directed microtubules by kinesin motor proteins. Indeed, the minus-end directed kinesin-6 Subito are known to be required for oocyte spindle organization in Drosophila melanogaster How multiple microtubule-bundling kinesins interact to produce a functional acentrosomal spindle is not known. In addition, there have been few studies on the meiotic function of one of the most important microtubule-bundlers in mitotic cells, the kinesin-5 KLP61F. This study found that the kinesin-5 KLP61F is required for spindle and centromere symmetry in oocytes. The asymmetry observed in the absence of KLP61F depends on NCD, the kinesin-12 KLP54D, and the microcephaly protein ASP. In contrast, KLP61F and Subito work together in maintaining a bipolar spindle. It is proposed that the prominent central spindle, stabilized by Subito, provides the framework for the coordination of multiple microtubule-bundling activities. The activities of several proteins, including NCD, KLP54D, and ASP, generate asymmetries within the acentrosomal spindle, while KLP61F and Subito balance these forces resulting in the capacity to accurately segregate chromosomes.
Lv, Z., Rosenbaum, J., Aspelmeier, T. and Grosshans, J. (2018). A 'molecular guillotine' reveals an interphase function of Kinesin-5. J Cell Sci. PubMed ID: 29361546
Motor proteins are important for transport and force generation in a variety of cellular processes and morphogenesis. This study designed a general strategy for conditional motor mutants by inserting a protease cleavage site into the 'neck' between the head domain and the stalk of the motor protein, making the protein susceptible to proteolytic cleavage at the neck by the corresponding protease. To demonstrate the feasibility of this approach, the cleavage site of TEV protease was inserted into the neck of the tetrameric motor Kinesin-5. Application of TEV protease led to a specific depletion and functional loss of Kinesin-5 in Drosophila embryos. With this approach, it was revealed that Kinesin-5 stabilized the microtubule network during interphase in syncytial embryos. The "molecular guillotine' can potentially be applied to many motor proteins due to the conserved structures of kinesins and myosins with accessible necks.
Tubman, E., He, Y., Hays, T. S. and Odde, D. J. (2018). Kinesin-5 mediated chromosome congression in insect spindles. Cell Mol Bioeng 11(1): 25-36. PubMed ID: 29552234
The microtubule motor protein kinesin-5 is well known to establish the bipolar spindle by outward sliding of antiparallel interpolar microtubules. In yeast, kinesin-5 also facilitates chromosome alignment "congression" at the spindle equator by preferentially depolymerizing long kinetochore microtubules (kMTs). The motor protein kinesin-8 has also been linked to chromosome congression. Therefore, this study sought to determine whether kinesin-5 or kinesin-8 facilitates chromosome congression in insect spindles. RNAi of the kinesin-5 Klp61F and kinesin-8 Klp67A were performed separately in Drosophila melanogaster S2 cells to test for inhibited chromosome congression. Klp61F RNAi, Klp67A RNAi, and control metaphase mitotic spindles expressing fluorescent tubulin and fluorescent Cid were imaged, and their fluorescence distributions were compared. RNAi of Klp61F with a weak Klp61F knockdown resulted in longer kMTs and less congressed kinetochores compared to control over a range of conditions, consistent with kinesin-5 length-dependent depolymerase activity. RNAi of the kinesin-8 Klp67A revealed that kMTs relative to the spindle lengths were not longer compared to control, but rather that the spindles were longer, indicating that Klp67A acts preferentially as a length-dependent depolymerase on interpolar microtubules without significantly affecting kMT length and chromosome congression. This study demonstrates that in addition to establishing the bipolar spindle, kinesin-5 regulates kMT length to facilitate chromosome congression in insect spindles. It expands on previous yeast studies, and it expands the role of kinesin-5 to include kMT assembly regulation in eukaryotic mitosis.
Costa, M. F. A. and Ohkura, H. (2019). The molecular architecture of the meiotic spindle is remodeled during metaphase arrest in oocytes. J Cell Biol. PubMed ID: 31278080
Before fertilization, oocytes of most species undergo a long, natural arrest in metaphase. Before this, prometaphase I is also prolonged, due to late stable kinetochore-microtubule attachment. How oocytes stably maintain the dynamic spindle for hours during these periods is poorly understood. This study reports that the bipolar spindle changes its molecular architecture during the long prometaphase/metaphase I in Drosophila melanogaster oocytes. By generating transgenic flies expressing GFP-tagged spindle proteins, it was found that 14 of 25 spindle proteins change their distribution in the bipolar spindle. Among them, microtubule cross-linking kinesins, MKlp1/Pavarotti and kinesin-5/Klp61F, accumulate to the spindle equator in late metaphase. The late equator accumulation of MKlp1/Pavarotti is regulated by a mechanism distinct from that in mitosis. While MKlp1/Pavarotti contributes to the control of spindle length, kinesin-5/Klp61F is crucial for maintaining a bipolar spindle during metaphase I arrest. This study provides novel insight into how oocytes maintain a bipolar spindle during metaphase arrest.
Hwang, J. H., Vuong, L. T. and Choi, K. W. (2020). Crumbs, Galla and Xpd are required for kinesin-5 regulation in mitosis and organ growth in Drosophila. J Cell Sci. PubMed ID: 32501288
Xeroderma Pigmentosum D (XPD) is a multi-function protein involved in transcription, DNA repair, and chromosome segregation. In Drosophila, Xpd interacts with Crumbs (Crb) and Galla to regulate mitosis during embryogenesis. It is unknown how these proteins are linked to mitosis. This study shows that Crb, Galla-2 and Xpd regulate nuclear division in syncytial embryo by interacting with Klp61F, the Drosophila mitotic kinesin-5 associated with bipolar spindles. Crb, Galla-2 and Xpd physically interact with Klp61F and co-localize to mitotic spindles. Knockdown of any of these proteins results in similar mitotic defects. These phenotypes are restored by overexpressing Klp61F, suggesting that Klp61F is a major effector. Mitotic defects of galla-2 RNAi are suppressed by Xpd overexpression but not vice versa Depletion of Crb, Galla-2 or Xpd results in a reduction of Klp61F levels. Reducing proteasome function restores Klp61F levels and suppress mitotic defects caused by knockdown of Crb, Galla-2 or Xpd. Further, eye growth is regulated by Xpd and Klp61F. Hence, this study proposes that Crb, Galla-2 and Xpd interact to maintain the level of Klp61F during mitosis and organ growth.
Jang, Y. G., Choi, Y., Jun, K. and Chung, J. (2020). Mislocalization of TORC1 to Lysosomes Caused by KIF11 Inhibition Leads to Aberrant TORC1 Activity. Mol Cells 43(8): 705-717. PubMed ID: 32759469
While the growth factors like insulin initiate a signaling cascade to induce conformational changes in the mechanistic target of rapamycin complex 1 (mTORC1), amino acids cause the complex to localize to the site of activation, the lysosome. The precise mechanism of how mTORC1 moves in and out of the lysosome is yet to be elucidated in detail in Drosophila. This study reports that microtubules and the motor protein KIF11 are required for the proper dissociation of mTORC1 from the lysosome upon amino acid scarcity. When microtubules are disrupted or KIF11 is knocked down, mTORC1 localizes to the lysosome even in the amino acid-starved situation where it should be dispersed in the cytosol, causing an elevated mTORC1 activity. Moreover, in the mechanistic perspective, this study discovered that mTORC1 interacts with KIF11 on the motor domain of KIF11, enabling the complex to move out of the lysosome along microtubules. These results suggest not only a novel way of the regulation regarding amino acid availability for mTORC1, but also a new role of KIF11 and microtubules in mTOR signaling.


The segregation of genetic material during mitosis is coordinated by the mitotic spindle, whose action depends upon the polarity patterns of its microtubules (MTs). Homotetrameric mitotic kinesin-5 motors can crosslink and slide adjacent spindle MTs, but it is unknown whether they or other motors contribute to establishing these MT polarity patterns. This study explored whether the Drosophila embryo kinesin-5 KLP61F, which plausibly crosslinks both parallel and antiparallel MTs (Tao, 2006; Sharp, 1999), displays a preference for parallel or antiparallel MT orientation. In motility assays, KLP61F was observed to crosslink and slide adjacent MTs, as predicted. Remarkably, KLP61F displayed a 3-fold higher preference for crosslinking MTs in the antiparallel orientation. This polarity preference was observed in the presence of ADP or ATP plus nonhydrolyzable ATP analog AMPPNP, but not AMPPNP alone, which induces instantaneous rigor binding. Also, a purified motorless tetramer containing the C-terminal tail domains displayed an antiparallel orientation preference, confirming that motor activity is not required. The results suggest that, during morphogenesis of the Drosophila embryo mitotic spindle, KLP61F's crosslinking and sliding activities could facilitate the gradual accumulation of KLP61F within antiparallel interpolar MTs at the equator, where the motor could generate force to drive poleward flux and pole-pole separation (van den Wildenberg, 2008).

By using fluorescence microscopy-based microtubule (MT)-MT sliding assays, it was first tested whether purified, full-length KLP61F, like its vertebrate ortholog, Eg5, is able to facilitate MT-MT sliding (Kapitein, 2005). To this end, biotinylated Cy-5-labeled MTs were specifically attached to a glass surface. Subsequently, the surface was blocked with the amphiphilic block copolymer Pluronic F108 to prevent nonspecific binding of MTs and KLP61F to the surface. Purified KLP61F and rhodamine-labeled MTs were added together with adenosine triphosphate (ATP). Then time series of images were acquired that showed clear movement of rhodamine-labeled MTs over immobilized Cy5-labeled MTs. Rhodamine MTs did not land or slide on regions of the surface where no MT was immobilized. This excludes the possibility that MTs were driven by KLP61F directly attached to the glass surface. In most of the recorded events, crosslinked, nonaligned MTs, were observed with a crossover point moving relative to both filaments with an average velocity (± standard deviation) of 11.0 ± 3.1 nm/s, which was independent of the crossing angle. Occasionally, the sliding MT rotated into alignment with the immobilized MT, whereupon the two relative velocities of sliding added up to approximately twice the individual velocities, indicating that these MTs all ended up aligned antiparallel. In some of the recorded events, the sliding MT had already been aligned. The average velocity measured for all aligned, sliding MTs was 26.7 ± 4.5 nm/s. These observations suggest that KLP61F can crosslink MTs in either parallel or antiparallel orientation and that it moves with a well-defined velocity along both crosslinked MTs, largely independent of their relative orientation, just like its Xenopus ortholog, Eg5 (Kapitein, 2005). However, the question remains whether either of these kinesin-5 motors preferentially crosslinks MTs into parallel or antiparallel polarity patterns (van den Wildenberg, 2008).

As a prelude to assaying kinesin-5's MT-crosslinking polarity preference, standard bundling assays were used to test the MT-bundling activity of the following constructs: (1) purified full-length KLP61F (a 520 kDa tetrameric holoenzyme), (2) a tetrameric 272 kDa native molecular weight (MW) 'stalk' fragment lacking both the N-terminal motor and the C-terminal tail domains, and (3) a tetrameric 378 kDa MW native 'motorless' (a.k.a. 'headless') fragment. As expected, highly purified motorless KLP61F, like the full-length protein, displayed robust MT-bundling activity, whereas the purified stalk subfragment displayed no detectable bundling activity, supporting the idea that KLP61F homotetramers must contain either N-terminal motor domains or C-terminal tail domains to be capable of bundling MTs (Tao, 2006; van den Wildenberg, 2008).

To determine whether KLP61F has a preference for crosslinking MTs into either parallel or antiparallel bundles, polarity-marked MTs and purified KLP61F (Tao, 2006) were mixed for 1 min in assay buffer containing nucleotides, and they were subsequently introduced into a microscope chamber with an aminosilanized glass surface, which led to a fixation of the relative orientation of MTs upon attachment. After rinsing the sample, the parallel and antiparallel MT bundles attached to the surface were counted to determine their relative abundance. In these assays, unlike the more routine bundling assays, the relative concentration of KLP61F and MTs was optimized to generate bundles consisting of two MTs and not more (van den Wildenberg, 2008).

It was observed that, in saturating concentrations of the nonhydrolyzable ATP analog AMPPNP, equal numbers of parallel and antiparallel MT pairs were formed. It was reasoned that this occurred because AMPPNP facilitates the strong binding of KLP61F motor domains to the MT tracks, immediately locking them in place in a tight binding state. In other words, AMPPNP freezes the on and off kinetics of the motors and will not allow potential differences in binding affinity of either the motor domains or the binding domains in the tails between antiparallel and parallel MTs to establish a preferred polarity pattern. The result further suggests that each individual KLP61F motor has considerable rotational flexibility because the pairs of motor domains at opposite ends of the stalk domain must be capable of rotating by 180° in order to crosslink MTs oriented in either parallel or antiparallel configurations. It should also be noted that, even if the orientational preference of a single crosslink was small compared to thermal energies, several motors could still collectively cause a strong orientational bias over time if the crosslinking is transient (van den Wildenberg, 2008).

In principle, the two sets of binding sites, on the motor domains and on the tails, could each cause an orientational bias, the bias could be equal or opposite, or just one set could cause the bias. The following experiments were designed to differentiate between the various scenarios. The existence of a bias implies a certain degree of mechanical torsional stiffness in the tetramers. Note that a bias caused by only one set of binding sites allows one to roughly localize flexibility in the molecule. To avoid the initial 'orientation quench' caused by AMPPNP on the motor domains, which appears to lock KLP61F-MT complexes in a random initial tight binding configuration, the assay was modified. MTs and KLP61F were first incubated in the presence of ATP for 1 min to allow the system to equilibrate. This time is appropriate because it exceeds the residence time of individual kinesin-5 motors on MTs but is short enough to prevent sliding to the end of travel, whereupon kinesin-5 reaches the ends of 'sorted' MTs. When this is allowed to occur, complicating events (e.g., 'snap-backs' of dangling MTs, et cetera) can introduce artifacts into the assays. After incubation, the crosslinked MTs were attached to the glass surface, and AMPPNP was flushed in to lock the KLP61F motors in an immotile state. Under these conditions, three times more antiparallel MT pairs were observed than parallel ones, indicating that the full-length KLP61F has a preference for generating antiparallel MT pairs in the presence of ATP (van den Wildenberg, 2008).

Earlier studies had shown that the KLP61F homolog Eg5 can diffuse axially along the MT polymer lattice in the presence of adenosine diphosphate (ADP) (Kwok, 2006); this process presumably does not involve specific and strong binding states of the motor domains but probably depends on interactions with the C-terminal tail domains instead (Tao, 2006; Kwok, 2006). To examine the MT-bundling behavior of KLP61F in this 'diffusive mode,' in which the binding via the motor domains is likely switched off, MT-MT crosslinking in the presence of KLP61F and ADP were examined. Three times more antiparallel than parallel MT crosslinking were observed under these conditions. All results thus suggest that the tail binding sites are responsible for the bias. To entirely exclude the possiblity that the motor domains are required, whether KLP61F's C-terminal MT-binding domains alone can cause the orientational preference of these kinesin-5 motors was tested. The orientation was determined of MTs bundled by motorless constructs (which were already shown to bundle MTs in the presence of ADP. Again three times more antiparallel than parallel MT bundles were observed. KLP61F thus has an approximately 3-fold preference for bundling antiparallel MTs over parallel ones. This preference is preserved when the motor domains are totally absent, as was the case for the motorless subfragment, or when they are switched off in a weakly and dynamically bound MT-binding state in the absence of ATP and in the presence of ADP (van den Wildenberg, 2008).

Taken together, these results demonstrate that the homotetrameric kinesin-5 KLP61F, like its homolog, Eg5, can crosslink and slide MTs. These findings further suggest that kinesin-5 motors display a preference for crosslinking MTs into antiparallel bundles. It may be reasonable to assume that the bipolar structure observed for Drosophila KLP61F (Kashina, 1996) and the MT-MT sliding activity demonstrated for Xenopus Eg5 (Kapitein, 2005) are shared by all members of the kinesin-5 family (Scholey, 2009). However, kinesin-5 motors appear to be deployed to play different roles in spindles from different systems, and these roles could be correlated with system-specific differences in the molecular architecture and mechanism of action of kinesin-5 motors. KLP61F is the first member of the kinesin-5 family explicitly shown to display both a bipolar ultrastructure (Kashina, 1996) and MT-MT sliding activity, both of which underlie the proposed kinesin-5-dependent 'sliding filament' mechanism (van den Wildenberg, 2008).

The molecular mechanism by which KLP61F preferentially crosslinks MTs into antiparallel orientations is not known. This is a fascinating problem that merits further detailed analysis. The observation that tetramers of both ADP-bound full-length KLP61F and motorless KLP61F subfragments preferentially crosslink MTs into antiparallel orientations shows that the mechanochemical activity of the motor domains is not essential for the antiparallel polarity preference. In this context, it is interesting to note that MT crosslinking is also brought about by the nonmotor MT-binding protein Ase1p, which displays a similar antiparallel orientation preference. Note that the antiparallel MT orientation preference of motorless KLP61F suggests that the C-terminal tail domains may control the polarity preference of full-length KLP61F, but the possibility cannot be excluded that active KLP61F motor domains (in contrast to those trapped in the presence of AMPPNP) could contribute as well. It is also noted that the tail domains contain the cyclin-dependent kinase (cdk)-dependent phosphorylatable bimC box, which may target kinesin-5 to spindle MTs (Sharp, 1999; Sawin, 1995), so it is tempting to speculate that the phosphorylation state of the bimC box influences the polarity preference of kinesin-5. For addressing the above issues, detailed structure-function studies of the MT-crosslinking polarity preference of headless and tailless, phosphorylated and nonphosphorylated KLP61F constructs are planned (van den Wildenberg, 2008).

On the basis of the results of the relative sliding experiments, together with the absence of any MT-crosslinking orientation preference in the presence of AMPPNP, it is apparent that full-length KLP61F is flexible enough to crosslink MTs in any orientation. However, to explain the orientation preference that is observed in the presence of ADP and ATP, it is imagined that some part of the tetramer must have sufficient torsional rigidity to form and maintain the antiparallel MT orientation. This apparent contradiction is resolved if one assumes that the stalk between the opposing tail domains is relatively rigid, that the C-terminal tail domains specifically interact with a MT, resulting in an antiparallel orientation preference, and that the flexibility of the motor domains resides in the neck and/or neck linker. An improved understanding of the torsional rigidity of different domains of the KLP61F homotetramer would therefore be illuminating (van den Wildenberg, 2008).

What are the implications of kinesin-5's antiparallel polarity preference for the mechanism of mitosis? At present, there is considerable interest in the mechanisms for establishing MT polarity patterns within mitotic spindles and in other MT-based structures, such as axons and dendrites. In astral mitotic spindles, such as those in the early Drosophila embryo, spindle MTs are organized into two overlapping radial arrays, with their minus ends located at the centrosomes and their plus ends facing the equator of the spindle. Consequently, MTs around and near the centrosomes are oriented parallel, whereas MTs overlapping with their plus ends at the equator are likely to encounter antiparallel neighbors. These antiparallel pairs are crucial for generating forces between the spindle poles. In some spindles, such as Drosophila embryo mitotic spindles, motor-dependent crosslinking and relative sliding of antiparallel MTs at the spindle equator is thought to underlie poleward flux within interpolar MT (ipMT) bundles and pole-pole separation during anaphase spindle elongation. It is plausible that antiparallel ipMT-MT crosslinking and sliding by kinesin-5, acting in concert with nonmotor MT-associated proteins and with nucleated-MT assembly around centrosomes and chromosomes, could play significant roles in establishing the MT polarity patterns found in spindles (van den Wildenberg, 2008).

The specific MT orientation preference of KLP61F motors is so far unique among mitotic sliding motors. The fact that purified kinesin-5 motors all appear to be slow, plus-end-directed bipolar homotetramers capable of crosslinking adjacent MTs is consistent with the idea that kinesin-5 homotetramers serve as dynamic MT-MT crosslinks that both bundle parallel MTs and drive antiparallel MT sliding and that this is their main contribution to mitotic spindle morphogenesis and function. The results suggest that in the Drosophila embryo, KLP61F could initially bind and crosslink MTs of either polarity throughout the spindle, thereby 'zipping' together parallel MTs to form MT bundles. This might be aided by an additional 'stickiness' caused by the tail domains. Then via on and off kinetics or after moving toward crosslinked MT plus ends, the antiparallel preference mediated by the tails would cause KLP61F to accumulate in the overlap region of antiparallel ipMTs at the spindle equator to efficiently slide them apart, thereby contributing to poleward flux and spindle elongation (van den Wildenberg, 2008).

Trim9 and Klp61F promote polymerization of new dendritic microtubules along parallel microtubules

Axons and dendrites are distinguished by microtubule polarity. In Drosophila, dendrites are dominated by minus-end-out microtubules while axons contain plus-end-out microtubules. Local nucleation in dendrites generates microtubules in both orientations. To understand why dendritic nucleation does not disrupt polarity, this study used live imaging to analyze the fate of microtubules generated at branch points. It was found that they had different rates of success exiting the branch based on orientation: correctly oriented minus-end-out microtubules succeeded in leaving about twice as often as incorrectly oriented microtubules. Increased success relied on other microtubules in a parallel orientation. From a candidate screen, Trim9 and kinesin-5 (Klp61F) were identified as machinery that promoted growth of new microtubules. In S2 cells, EB1 recruited Trim9 to microtubules. Klp61F promoted microtubule growth in vitro and in vivo, and could recruit Trim9 in S2 cells. In summary, the data argue that Trim9 and kinesin-5 act together at microtubule plus ends to help polymerizing microtubules parallel to pre-existing ones resist catastrophe (Feng, 2021).

The almost uniform minus-end-out microtubule polarity in Drosophila dendrites has proven a useful system for the identification of mechanisms that control microtubule organization and polarity. One surprise is that multiple mechanisms operate in parallel even in this very confined space with simple microtubule layout. Two basic types of polarity control mechanisms have been identified: those that can establish microtubule polarity independently of existing microtubules, and those that act as positive feedback loops to reinforce the predominant polarity (Feng, 2021).

Two mechanisms have been identified that can autonomously contribute to dendritic minus-end-out microtubule polarity: local nucleation and minus-end growth. Very early in the development of dendritic arborization neurons in the Drosophila embryo, new dendrites are populated by plus-end-out microtubules that grow in from the cell body (Feng, 2019). The next step is for slower-growing minus ends to enter dendrites from the cell body adding minus-end-out microtubules. Dendrites then remain with mixed polarity for the rest of embryogenesis, and eventually this resolves to minus-end-out in larvae. Minus-end-out microtubules can also be generated locally in dendrites by nucleation. Like microtubule growth from the cell body, nucleation can contribute plus-end-out and minus-end-out microtubules. In mature dendritic arborization neurons, nucleation at branch points is biased towards generating minus-end-out microtubules. Several mechanisms to bias nucleation have been shown to operate in neurons, but they have not been shown to act in mature dendritic arborization neurons. In developing C. elegans sensory dendrites, nucleation sites cluster close to the tip, resulting in a short region of plus-end-out microtubules beyond the cluster and the proximal dendrite dominated by minus-end-out microtubules (Liang, 2020). However, in ddaE dendrites, nucleation sites are found not just at the dendrite tip but throughout the arbor at branch points. In ddaE dendrites, nucleation has been proposed to be biased by recruitment to only one side of Golgi outposts. However, nucleation sites have recently been shown to be recruited to endosomes rather than Golgi in ddaE dendrites (Weiner, 2020), making the earlier findings difficult to interpret. In summary, both growth from microtubule ends and nucleation of new microtubules create minus-end-out, as well as plus-end-out, microtubules in dendrites (Feng, 2021).

In addition to the de novo mechanisms that add minus-end-out microtubules to dendrites described above, feedback mechanisms reinforce the dominant polarity. One of these controls the direction of microtubule growth at branch points. As microtubules grow from distal regions of the dendrite into branch points, they encounter a choice to grow towards the cell body or away from the cell body. Kinesin-2, together with Eb1, Apc and Apc2, allows the growing plus end to track existing microtubules, reinforcing polarity. If this mechanism is eliminated, polarity in dendrites remains mixed. The final percentage of minus-end-out microtubules seems to depend on the angle of branches such that in ddaE dendrites, in which branch angles are close to 90° and microtubules can easily turn either direction at branch points, ~50% of microtubules are minus-end-out. In dendrites with more acute branch angles that help direct growing plus ends towards the cell body, 70% of microtubules remain minus-end-out, even when steering is eliminated. The mechanism described in this study, that selectively promotes growth of new microtubules out of branch points along parallel microtubules, seems to act as an additional feedback loop that helps align newly nucleated microtubules with pre-existing ones. Impairing this mechanism seems to have little impact on overall polarity in dendrites, unlike eliminating steering. Quality control of newly nucleated microtubules may have a small effect on overall polarity because nucleation itself is somewhat biased and nucleation may contribute at relatively low levels to the overall microtubule population at steady state in mature neurons. Quality control of new microtubules may exist as a backup mechanism to help maintain microtubule organization under stressful conditions like axon injury, which can upregulate nucleation (Feng, 2021).

The data suggest that Trim9 and Klp61F are both needed for new microtubules to grow in parallel bundles with pre-existing microtubules. The fact that reduction of either results in a phenotype suggests that they act together, rather than in parallel, to control plus-end behavior. A vertebrate Trim9 family member, Trim46, organizes parallel microtubules at the axon initial segment and has autonomous parallel bundling activity in vitro. A role for the single Drosophila Trim9 family member in parallel orientation of microtubules suggests that this may be an ancestral function of this family, rather than a new function that evolved with the expansion of the family in vertebrates. If this is the case, then the other vertebrate family members in the C-1 subfamily containing a microtubule-binding COS box may all engage parallel microtubules, perhaps in different cell types or subcellular regions. In vitro, Trim46 has a strong preference for interacting with parallel bundles of microtubules over individual microtubules, and so tends to accumulate some distance behind dynamic plus ends. When a depolymerizing plus end encounters a bundled region decorated with Trim46, catastrophe is strongly inhibited. Although the ability of Trim46 to rescue catastrophes would be expected to promote growth like Drosophila Trim9, the site of action seems to be somewhat different. Trim46 acts at bundled regions of microtubules behind the dynamic plus end, and Drosophila Trim9 prevents catastrophes from happening, likely at the plus end, with Eb1 and Klp61F (Feng, 2021).

The in vivo role of Klp61F in promoting continued growth of plus ends out of the branch point is consistent with previous in vitro studies showing that Eg5 dimers can accumulate at the plus end and promote polymerization. Klp61F tetramers form rods ~95 nm in length that crosslink microtubules with spacing of at least 60 nm. The spacing of microtubules bundled with Trim46 is ~37 nm. Based on these general size considerations, as well as in vitro activities of Klp61F and Trim46, the following model is proposed. Klp61F could be constantly traveling along stable bundles of microtubules. If a new microtubule approaches within 60 nm, then it could be captured. As Klp61F interacts with the growing plus end it could promote polymerization. At the same time, Trim9 could be recruited to the growing plus end by Eb1, perhaps through [S/T]-x-[I/L]-P motifs, which interact with Eb1. Both Drosophila Trim9 proteins contain two potential motifs at amino acids: the first is at 42-SALP-46 in both, and the second is 479-TILP-483 in the RB isoform and 468-TILP-472 in the RA isoform. After Klp61F grabs the growing microtubule with Eb1 and Trim9 at its tip, Trim9 could reinforce the parallel interaction. Although this model provides an initial framework for the function of Klp61F and Trim9 in dendrites, it is quite speculative and raises many additional questions. For example, if Klp61F is in constant flux towards the plus end of microtubules, how is it transported into dendrites? And do these proteins really act sequentially? If so, perhaps family members collaborate in a similar way in other cellular scenarios; for example, at the axon initial segment (Feng, 2021).

Kinesin-5 is essential for growth-cone turning

Inhibition of chick kinesin-5, a mitotic motor protein also expressed in neurons, causes axons to grow faster as a result of alterations in the forces on microtubules (MTs) in the axonal shaft. This study investigated whether kinesin-5 plays a role in growth-cone guidance. Growth-cone turning requires that MTs in the central (C-) domain enter the peripheral (P-) domain in the direction of the turn. Inhibition of kinesin-5 in cultured neurons prevents MTs from polarizing within growth cones and causes them to grow past cues that would normally cause them to turn. It was found that kinesin-5 is enriched in the transition (T-) zone of the growth cone and that kinesin-5 is preferentially phosphorylated on the side opposite the invasion of MTs. Moreover, when a growth cone encounters a turning cue, phospho-kinesin-5 polarizes even before the growth cone turns. Additional studies indicate that kinesin-5 works in part by antagonizing cytoplasmic dynein and that these motor-driven forces function together with the dynamic properties of the MTs to determine whether MTs can enter the P-domain. It is proposed that kinesin-5 permits MTs to selectively invade one side of the growth cone by opposing their entry into the other side (Nadar, 2009).

Novel nuclear defects in KLP61F-deficient mutants in Drosophila are partially suppressed by loss of Ncd function

KLP61F in Drosophila and other BimC kinesins are essential for spindle bipolarity across species; loss of BimC function generates high frequencies of monopolar spindles. Concomitant loss of Kar3 kinesin function increases the frequency of bipolar spindles although the underlying mechanism is not known. Recent studies raise the question of whether BimC kinesins interact with a non-microtubule spindle matrix rather than spindle microtubules. This study presents cytological evidence that loss of KLP61F function generates novel defects during M-phase in the organization and integrity of the nuclear lamina, an integral component of the nuclear matrix. Larval neuroblasts and spermatocytes of klp61F mutants showed deep involutions in the nuclear lamina extending toward the centrally located centrosomes. Repositioning of centrosomes to form monopolar spindles probably does not cause invaginations as similar invaginations formed in spermatocytes lacking centrosomes entirely. Immunofluorescence microscopy indicated that Non-claret disjunctional (Ncd) is a component of the nuclear matrix in somatic cells and spermatocytes. Loss of Ncd function increases the frequency of bipolar spindles in klp61F mutants. Nuclear defects were incompletely suppressed; micronuclei formed near telophase at the poles of bipolar spindle in klp61F ncd spermatocytes. These results are consistent with a model in which KLP61F prevents Ncd-mediated collapse of a nonmicrotubule matrix derived from the interphase nucleus (Wilson, 2004).

This study present cytological evidence that loss of KLP61F function generates spindle defects as well as novel defects in organization of the nuclear matrix during M-phase in somatic cells and spermatocytes. These results also show that Ncd is nuclear during interphase and spindle-associated in M-phase in the soma and male germ line. Loss of Ncd function increases the frequency of biastral spindles in klp61F mutants, but fails or incompletely restores nuclear matrix defects. These findings raise new questions about the molecular basis of genetic interactions between KLP61F and Ncd (Wilson, 2004).

Somatic cells in klp61F and klp61F ncd mutants with monopolar spindles show deep invaginations in the nuclear lamina that extended toward centrally located centrosomes. Similar involutions were found in klp61F mutant spermatocytes judged to be near prometaphase, irrespective of the presence or absence of centrosomes. These observations suggest that the driving force in forming invaginations in the nuclear lamina is associated with nuclear and/or cytoplasmic material rather than with centrosomes or centrosome organized microtubules. A contribution of nuclear forces to repositioning of centrosomes has precedence in yeast; spindle-pole bodies in preassembled spindles move through the nuclear envelope to side-by-side positions when temperature-dependent BimC function is inactivated at non-permissive temperatures. Because spindle pole bodies assume face-to-face positions when microtubules are depolymerized, side-by-side positions suggests that nuclear forces contribute to spindle defects in BimC-deficient yeast as well (Wilson, 2004).

Nuclear defects in somatic cells differed from those in spermatocytes, raising the question of whether KLP61F function in somatic cells and spermatocytes is mediated by a common mechanism or two different mechanisms. Arguments can be made for and against a common mechanism. The strongest argument for a common mechanism is the striking similarity of spindle defects in somatic cells and spermatocytes. Another argument is the ability of ncd mutants to suppress the klp61F mutant phenotype in both cell types. At first glance, other aspects of the mutant phenotype are not consistent with a common function. Somatic cells in KLP61F-deficient animals showed extensive disorganization of the nuclear lamina, including cells showing bipolar positioning of centrosomes and metaphase alignment of chromosomes. In contrast to somatic cells, the nuclear lamina appeared to collapse around bivalents near prometaphase and form micronuclei in klp61F mutant spermatocytes. These differences could reflect different functions in somatic cells and spermatocytes. Alternatively, the difference may reflect cell cycle regulation; the spindle assembly checkpoint is active in somatic cells, but inactive or severely abrogated in spermatocytes. This view is consistent with the disorganized state of the nuclear lamina in cultured clone 8 cells that were delayed in mitosis with an inhibitor of APC. Thus, disorganization of the nuclear lamina in somatic cells and formation of micronuclei in spermatocytes in KLP61F-deficient mutants could reflect a common underlying defect in different cell types (Wilson, 2004).

KLP61F shows overlapping, but differential localization during mitosis and male meiosis. In somatic cells, KLP61F is highly enriched near centrosomal asters during prophase, spindle-associated in metaphase and located in midbodies in telophase. In meiotic spermatocytes, KLP61F fails to show centrosomal enrichment or spindle association, but in late anaphase/early telophase KLP61F localizes to a sphere that bisects the entire spermatocyte and then follows the ingressing cleavage furrow. Similar localization in proliferating germ cells in telophase was found to reflect interactions, directly or indirectly, with components of fusomes. It is not possible to draw firm conclusions from the failure to detect KLP61F localization to centrosomal asters or to spindles as a small pool could escape detection methods. However, given static positioning of Eg5 in spindles assembled in Xenopus egg extracts, the failure to detect KLP61F localization to male meiotic spindles may indicate that KLP61F is not associated with spindle microtubules, but with non-microtubule binding partners. In most cell types, BimC kinesins are diffusely distributed throughout the cytoplasm during interphase. Localization of BimC kinesins to spindles in vertebrate cells has been linked to phosphorylation of a Cdk1 target site in the conserved BimC Box near the carboxyl tail region of these kinesins, postulated to elicit or strengthen intrinsic microtubule binding activity and spindle localization. However, phosphorylation of the BimC Box of Cut7 in Saccharomyces pombe is not required for spindle association or for Cut7 function in assembly of a bipolar spindle. It is possible that KLP61F and other BimC kinesins crosslink microtubules and non-microtubule binding partners during interphase. BimC Box phosphorylation may downregulate microtubule binding activity and allow interactions with non-microtubule binding partners to direct KLP61F localization during M-phase. Identification of non-microtubule binding partners and genetic analysis of BimC Box function in KLP61F localization could test these possibilities (Wilson, 2004).

Similar to Kar3 kinesins in vertebrates, Ncd is nuclear during interphase in spermatocytes and in somatic cells as well as in somatically derived clone 8 cells. It is not clear why Ncd fails to show nuclear localization in early embryos, but the rapidity of the cell cycle in early embryos may preclude complete reorganization of the nuclear envelope and nuclear entry of Ncd through nuclear pores. Two lines of cytological evidence suggest that Ncd may be associated with the nuclear matrix during interphase in somatic and male germ line cells. First, Ncd shows subnuclear enrichment near heterochromatin attached to the nuclear envelope. Given that fibrillar components of the nuclear matrix connect chromatin to the inner nuclear membrane, subnuclear enrichment could reflect localization of Ncd to fibrillar components of the nuclear matrix and/or localization to chromatin-associated material at these sites. Second, Ncd localizes to fibers extending between the poles of metaphase spindles in somatic cells and cultured clone 8 cells, reminiscent of Ncd localization in embryos (Endow, 1997). The functional significance of these fibers is not clear since pole to pole fibers have not been reported in female meiotic spindles and this study did not detect strong immunostaining of similar fibers in meiotic spermatocytes. However, there is a precedence for localization of other nuclear matrix proteins to spindle fibers. The nuclear matrix protein NuSAP is localized to spindle-associated fibers in cultured vertebrate cells and loss of its function generates defects in spindle organization and chromosome segregation. The nuclear matrix protein Skeletor also localizes to fibers in embryonic spindles although the fibers do not extend the full distance between spindle poles. Loss of Ncd function did not appreciably alter Skeletor distribution in spermatocytes, indicating that Ncd is not necessary or plays only a very limited role in Skeletor localization. Nonetheless, the findings are consistent with the view that Ncd is a component of the nuclear matrix in somatic cells and spermatocytes (Wilson, 2004).

At this point, it is only possible to speculate on the relationship between spindle and nuclear defects in klp61F mutants and the functional significance of Ncd-mediated suppression. With few exceptions, cooperation between BimC and Kar3 kinesins in spindle assembly has been ascribed to application of antagonistic motive forces to spindle microtubules to establish or maintain centrosome separation. According to this view, nuclear defects in KLP61F-deficient animals could be secondary to primary defects in spindle organization; increasing the frequency of bipolar spindles in klp61F ncd mutants results in a decreased frequency of nuclear defects. However, this explanation does not easily explain formation of micronuclei at the poles of bipolar spindles in klp61F ncd spermatocytes. Moreover, collapse of the nuclear lamina about bivalents cannot be ascribed to spindle defects since similar defects are not found in meiotic spermatocytes of ß2tn mutants that lack microtubules and spindle structures owing to loss of an essential testis specific ß-tubulin. An alternative interpretation of the findings is that spindle defects are secondary; spindle defects reflect collapse of a nonmicrotubule spindle matrix that is derived from the nuclear matrix and attached to centrosomes and/or spindle microtubules. According to this view, interactions between KLP61F and nonmicrotubule binding partners prevent collapse of a compressible spindle matrix when nuclear and cytoplasmic contents mix at prometaphase, whether or not KLP61F is spindle associated (Wilson, 2004).

The results of this study are in part unexpected because they question the assumed relationship between localization and function of a microtubule-dependent motor protein. KLP61F is required for spindle bipolarity, but its function in male meiosis does not require spindle association. Conversely, KLP61F localizes to cleavage furrows, but it is not required for cytokinesis. With these contradictions in mind, further work must address the mechanism of KLP61F function in spindle organization and the functional significance of nuclear localization of Ncd (Wilson, 2004).

Functional coordination of three mitotic motors in Drosophila embryos

It is well established that multiple microtubule-based motors contribute to the formation and function of the mitotic spindle, but how the activities of these motors interrelate remains unclear. This study visualized spindle formation in living Drosophila embryos to show that spindle pole movements are directed by a temporally coordinated balance of forces generated by three mitotic motors, cytoplasmic dynein, KLP61F, and Ncd. Specifically, these findings suggest that dynein acts to move the poles apart throughout mitosis and that this activity is augmented by KLP61F after the fenestration of the nuclear envelope, a process analogous to nuclear envelope breakdown, which occurs at the onset of prometaphase. Conversely, Ncd generates forces that pull the poles together between interphase and metaphase, antagonizing the activity of both dynein and KLP61F and serving as a brake for spindle assembly. During anaphase, however, Ncd appears to have no effect on spindle pole movements, suggesting that its activity is down-regulated at this time, allowing dynein and KLP61F to drive spindle elongation during anaphase B (Sharp, 2000; . Full text of article).

A bipolar kinesin

Chromosome segregation during mitosis depends on the action of the mitotic spindle, a self-organizing, bipolar protein machine which uses microtubules (MTs) and their associated motors. Members of the BimC subfamily of kinesin-related MT-motor proteins are believed to be essential for the formation and functioning of a normal bipolar spindle. Here this study reports that KRP130 (Klp61F), a homotetrameric BimC-related kinesin purified from Drosophila melanogaster embryos, has an unusual ultrastructure. It consists of four kinesin-related polypeptides assembled into a bipolar aggregate with motor domains at opposite ends, analogous to a miniature myosin filament. Such a bipolar 'minifilament' could crosslink spindle MTs and slide them relative to one another (Kashina, 1996).

The kinesin-like protein KLP61F is essential for mitosis in Drosophila

Disruption of a recently discovered kinesin-like protein in Drosophila, KLP61F, results in a mitotic mutation lethal to the organism. In the absence of KLP61F function, spindle poles fail to separate, resulting in the formation of monopolar mitotic spindles. The resulting phenotype of metaphase arrest with polyploid cells is reminiscent of that seen in the fungal bimC and cut7 mutations, where it has also been shown that spindle pole bodies are not segregated. KLP61F is specifically expressed in proliferating tissues during embryonic and larval development, consistent with a primary role in cell division. The structural and functional homology of the KLP61F, bimC, cut7, and Eg5 kinesin-like proteins demonstrates the existence of a conserved family of kinesin-like molecules important for spindle pole separation and mitotic spindle dynamics (Heck, 1993; Full text of article).

A nonmotor microtubule binding site in Kinesin-5 is required for filament crosslinking and sliding

Kinesin-5, a widely conserved motor protein required for assembly of the bipolar mitotic spindle in eukaryotes, forms homotetramers with two pairs of motor domains positioned at opposite ends of a dumbbell-shaped molecule. It has long been assumed that this configuration of motor domains is the basis of kinesin-5's ability to drive relative sliding of microtubules. Recently, it has been suggested that in addition to the N-terminal motor domain, kinesin-5 also has a nonmotor microtubule binding site in its C terminus. However, it is not known how the nonmotor domain contributes to motor activity, or how a kinesin-5 tetramer utilizes a combination of four motor and four nonmotor microtubule binding sites for its microtubule organizing functions. This study shows, in single molecule assays, that Xenopus kinesin-5 homotetramers require the nonmotor C terminus for crosslinking and relative sliding of two microtubules. Remarkably, this domain enhances kinesin-5's microtubule binding without substantially reducing motor activity. These results suggest that tetramerization of kinesin-5's low-processivity motor domains is not sufficient for microtubule sliding because the motor domains alone are unlikely to maintain persistent microtubule crosslinks. Rather, kinesin-5 utilizes nonmotor microtubule binding sites to tune its microtubule attachment dynamics, enabling it to efficiently align and sort microtubules during metaphase spindle assembly and function (Weinger, 2011).


Search PubMed for articles about Drosophila Klp61f

Feng, C., Thyagarajan, P., Shorey, M., Seebold, D. Y., Weiner, A. T., Albertson, R. M., Rao, K. S., Sagasti, A., Goetschius, D. J. and Rolls, M. M. (2019). Patronin-mediated minus end growth is required for dendritic microtubule polarity. J Cell Biol 218(7): 2309-2328. PubMed ID: 31076454

Feng, C., Cleary, J. M., Kothe, G. O., Stone, M. C., Weiner, A. T., Hertzler, J. I., Hancock, W. O. and Rolls, M. M. (2021). Trim9 and Klp61F promote polymerization of new dendritic microtubules along parallel microtubules. J Cell Sci. PubMed ID: 33988240

Heck, M. M., et al. (1993). The kinesin-like protein KLP61F is essential for mitosis in Drosophila. J. Cell Biol. 123(3): 665-79. PubMed ID: 8227131

Kapitein, L. C. et al. (2005). The bipolar mitotic kinesin Eg5 moves on both microtubules that it crosslinks. Nature 435: 114-118. PubMed ID: 15875026

Kashina, A. S., et al. (1996). A bipolar kinesin. Nature 379: 270-272. PubMed ID: 8538794

Kwok, B. H., et al. (2006). Allosteric inhibition of kinesin-5 modulates its processive directional motility. Nat. Chem. Biol. 2: 480-485. PubMed ID: 16892050

Liang, X., Kokes, M., Fetter, R. D., Sallee, M. D., Moore, A. W., Feldman, J. L. and Shen, K. (2020). Growth cone-localized microtubule organizing center establishes microtubule orientation in dendrites. Elife 9. PubMed ID: 32657271

Nadar, V. C., Ketschek, A., Myers, K. A., Gallo, G. and Baas, PW. (2009). Kinesin-5 is essential for growth-cone turning. Curr. Biol. 18(24): 1972-7. PubMed ID: 19084405

Sawin, K. E. and Mitchison, T. J. (1995). Mutations in the kinesin-like protein Eg5 disrupting localization to the mitotic spindle. Proc. Natl. Acad. Sci. 92: 4289-4293. PubMed ID: 7753799

Scholey, J. M. (2009). Kinesin-5 in Drosophila embryo mitosis: sliding filament or spindle matrix mechanism? Cell Motil. Cytoskeleton 66(8): 500-8. PubMed ID: 19291760

Sharp, D. J., McDonald, K. L., Brown, H. M., Matthies, H. J., Walczak, C., Vale, R. D., Mitchison, T. J. and Scholey, J. M. (1999). The bipolar kinesin, KLP61F, cross-links microtubules within interpolar microtubule bundles of Drosophila embryonic mitotic spindles. J. Cell Biol. 144(1): 125-38. PubMed ID: 9885249

Sharp, D. J., Brown, H. M., Kwon, M., Rogers, G. C., Holland, G. and Scholey, J. M. (2000). Functional coordination of three mitotic motors in Drosophila embryos. Mol. Biol. Cell. 11(1): 241-53. PubMed ID: 10637305

Tao, L., Mogilner, A., Civelekoglu-Scholey, G., Wollman, R., Evans, J., Stahlberg, H. and Scholey, J. M. (2006). A homotetrameric kinesin-5, KLP61F, bundles microtubules and antagonizes Ncd in motility assays. Curr. Biol 16: 2293-2302. PubMed ID: 17141610

van den Wildenberg, S. M., Tao, L., Kapitein, L. C., Schmidt, C. F., Scholey, J. M. and Peterman, E. J. (2008). The homotetrameric kinesin-5 KLP61F preferentially crosslinks microtubules into antiparallel orientations. Curr. Biol. 18(23): 1860-4. PubMed ID: 19062285

Weiner, A. T., Seebold, D. Y., Torres-Gutierrez, P., Folker, C., Swope, R. D., Kothe, G. O., Stoltz, J. G., Zalenski, M. K., Kozlowski, C., Barbera, D. J., Patel, M. A., Thyagarajan, P., Shorey, M., Nye, D. M. R., Keegan, M., Behari, K., Song, S., Axelrod, J. D. and Rolls, M. M. (2020). Endosomal Wnt signaling proteins control microtubule nucleation in dendrites. PLoS Biol 18(3): e3000647. PubMed ID: 32163403

Weinger, J. S., et al. (2011). A nonmotor microtubule binding site in Kinesin-5 is required for filament crosslinking and sliding. Curr. Biol. 21: 154-160. PubMed ID: 21236672

Wilson, P. G. (1999). BimC motor protein KLP61F cycles between mitotic spindles and fusomes in Drosophila germ cells. Curr. Biol. 9(16): 923-6. PubMed ID: 10469596

Wilson, P. G., Simmons, R. and Saighal, S. (2004). Novel nuclear defects in KLP61F-deficient mutants in Drosophila are partially suppressed by loss of Ncd function. J. Cell Sci. 117(Pt 21): 4921-33. PubMed ID: 15367580

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date revised: 17 December 2021

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