Dishevelled Associated Activator of Morphogenesis: Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References
Gene name - Dishevelled Associated Activator of Morphogenesis

Synonyms -

Cytological map position- 1F2--3

Function - cytoskeleton

Keywords - trachea, taenidial organization

Symbol - DAAM

FlyBase ID: FBgn0025641

Genetic map position - X

Classification - formin

Cellular location - cytoplasmic



NCBI links: Precomputed BLAST | EntrezGene | UniGene | HomoloGene

Recent literature
Vogler, G., Liu, J., Iafe, T. W., Migh, E., Mihaly, J. and Bodmer, R. (2014). Cdc42 and formin activity control non-muscle myosin dynamics during Drosophila heart morphogenesis. J Cell Biol 206: 909-922. PubMed ID: 25267295
Summary:

During heart formation, a network of transcription factors and signaling pathways guide cardiac cell fate and differentiation, but the genetic mechanisms orchestrating heart assembly and lumen formation remain unclear. This study shows that the small GTPase Cdc42 is essential for Drosophila melanogaster heart morphogenesis and lumen formation. Cdc42 genetically interacts with the cardiogenic transcription factor tinman; with dDAAM which belongs to the family of actin organizing formins; and with zipper, which encodes nonmuscle myosin II. Zipper is required for heart lumen formation, and its spatiotemporal activity at the prospective luminal surface is controlled by Cdc42. Heart-specific expression of activated Cdc42, or the regulatory formins dDAAM and Diaphanous caused mislocalization of Zipper and induced ectopic heart lumina, as characterized by luminal markers such as the extracellular matrix protein Slit. Placement of Slit at the lumen surface depends on Cdc42 and formin function. Thus, Cdc42 and formins play pivotal roles in heart lumen formation through the spatiotemporal regulation of the actomyosin network (Vogler, 2014).

Molnar, I., Migh, E., Szikora, S., Kalmar, T., Vegh, A. G., Deak, F., Barko, S., Bugyi, B., Orfanos, Z., Kovacs, J., Juhasz, G., Varo, G., Nyitrai, M., Sparrow, J. and Mihaly, J. (2014). DAAM is required for thin filament formation and Sarcomerogenesis during muscle development in Drosophila. PLoS Genet 10: e1004166. PubMed ID: 24586196
Summary:

During muscle development, myosin and actin containing filaments assemble into the highly organized sarcomeric structure critical for muscle function. Although sarcomerogenesis clearly involves the de novo formation of actin filaments, this process remained poorly understood. This study shows that mouse and Drosophila members of the DAAM formin family are sarcomere-associated actin assembly factors enriched at the Z-disc and M-band. Analysis of DAAM mutants revealed a pivotal role in myofibrillogenesis of larval somatic muscles, indirect flight muscles and the heart. Loss of DAAM function results in multiple defects in sarcomere development including thin and thick filament disorganization, Z-disc and M-band formation, and a near complete absence of the myofibrillar lattice. Collectively, these data suggest that DAAM is required for the initial assembly of thin filaments, and subsequently it promotes filament elongation by assembling short actin polymers that anneal to the pointed end of the growing filaments, and by antagonizing the capping protein Tropomodulin (Molnar, 2014).

Dollar, G., Gombos, R., Barnett, A. A., Sanchez Hernandez, D., Maung, S. M., Mihaly, J. and Jenny, A. (2016). Unique and overlapping functions of formins Frl and DAAM during ommatidial rotation and neuronal development in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 26801180
Summary:
The non-canonical Frizzled/Planar cell polarity (PCP) pathway regulates establishment of polarity within the plane of an epithelium to generate diversity of cell fates, asymmetric, but highly aligned structures, or to orchestrate the directional migration of cells during convergent extension during vertebrate gastrulation. In Drosophila, PCP signaling is essential to orient actin wing hairs and to align ommatidia in the eye, in part by coordinating the movement of groups of photoreceptor cells during ommatidial rotation. Importantly, the coordination of PCP signaling with changes in the cytoskeleton is essential for proper epithelial polarity. Formins polymerize linear actin filaments and are key regulators of the actin cytoskeleton. This study shows that the Diaphanous related formin Frl, the single fly member of the FMNL (Formin related in Leukocytes/ Formin-like) formin subfamily affects ommatidial rotation in the Drosophila eye and is controlled by the Rho family GTPase Cdc42. Interestingly, it was also found that frl mutants exhibit an axon growth phenotype in the mushroom body, a center for olfactory learning in the Drosophila brain, which is also affected in a subset of PCP genes. Significantly, Frl cooperates with Cdc42 and another formin, DAAM, during mushroom body formation. This study thus suggests that different formins can cooperate or act independently in distinct tissues, likely integrating various signaling inputs with the regulation of the cytoskeleton. It furthermore highlights the importance and complexity of formin dependent cytoskeletal regulation in multiple organs and developmental contexts.
Szikora, S., Foldi, I., Toth, K., Migh, E., Vig, A., Bugyi, B., Maleth, J., Hegyi, P., Kaltenecker, P., Sanchez-Soriano, N. and Mihaly, J. (2017). The formin DAAM is required for coordination of the actin and microtubule cytoskeleton in axonal growth cones. J Cell Sci [Epub ahead of print]. PubMed ID: 28606990
Summary:
Directed axonal growth depends on proper coordination of the actin and microtubule cytoskeleton in the growth cone. However, despite the relatively large number of proteins implicated in actin-microtubule cross-talk, the mechanisms whereby actin polymerization is coupled to microtubule stabilization and advancement in the peripheral growth cone remained largely unclear. This study identified the formin DAAM as a novel factor playing a role in concerted regulation of actin and microtubule remodeling in Drosophila primary neurons. In vitro DAAM binds to F-actin as well as microtubules and it has the ability to crosslink the two filament systems. Accordingly, DAAM associates with the neuronal cytoskeleton, and a significant fraction of DAAM accumulates at places where the actin filaments overlap with that of microtubules. Loss of DAAM affects growth cone and microtubule morphology and several aspects of microtubule dynamics, whereas biochemical and cellular assays revealed a microtubule stabilization activity and binding to the microtubule tip protein EB1. Together these data suggest that besides operating as an actin assembly factor, DAAM is involved in linking filopodial actin remodeling to microtubule stabilization during axonal growth.
Vig, A. T., Foldi, I., Szikora, S., Migh, E., Gombos, R., Toth, M. A., Huber, T., Pinter, R., Talian, G. C., Mihaly, J. and Bugyi, B. (2017). The activities of the c-terminal regions of the formin protein Disheveled-associated activator of morphogenesis (Daam) in actin dynamics. J Biol Chem [Epub ahead of print]. PubMed ID: 28642367
Summary:
Disheveled-associated activator of morphogenesis (DAAM) is a diaphanous-related formin protein essential for the regulation of actin cytoskeleton dynamics in diverse biological processes. The conserved formin homology 1 and 2 (FH1-FH2) domains of DAAM catalyze actin nucleation and processively mediate filament elongation. These activities are indirectly regulated by the N-, and C-terminal regions flanking the FH1-FH2 domains. Recently, the C-terminal diaphanous-autoregulatory domain (DAD) and the C-terminus (CT) of formins have also been shown to regulate actin assembly by directly interacting with actin. To better understand the biological activities of DAAM, the role of DAD-CT regions of Drosophila DAAM in its interaction with actin was studied with in vitro biochemical and in vivo genetic approaches. The DAD-CT region was found to bind actin in vitro and that its main actin-binding element is the CT region, which does not influence actin dynamics on its own. However, it was also found that it can tune the nucleating activity and the filament end-interaction properties of DAAM in an FH2 domain-dependent manner. It was also demonstrated that DAD-CT makes the FH2 domain more efficient in antagonizing with capping protein. Consistently, in vivo data suggested that the CT region contributes to DAAM-mediated filopodia formation and dynamics in primary neurons. In conclusion, these results demonstrate that the CT region of DAAM plays an important role in actin assembly regulation in a biological context.
BIOLOGICAL OVERVIEW

Formins are involved in a wide range of cellular processes that require the remodeling of the actin cytoskeleton. This study analyzes a novel Drosophila formin, belonging to the recently described DAAM subfamily. In contrast to previous assumptions, it is shown that DAAM plays no essential role in planar cell polarity signaling, but it has striking requirements in organizing apical actin cables that define the taenidial fold pattern of the tracheal cuticle. These observations provide evidence the first time that the function of the taenidial organization is to prevent the collapse of the tracheal tubes. The results indicate that although DAAM is regulated by RhoA, it functions upstream or parallel to the non-receptor tyrosine kinases Src42A and Tec29 to organize the actin cytoskeleton and to determine the cuticle pattern of the Drosophila respiratory system (Matusek, 2006).

Proteins of the formin family play key roles in the regulation of the cytoskeleton. Although the connection between formins and microtubules is less well understood, formins are implicated in a large number of actin-based processes, including cell polarization, division, movement, stress fiber formation and vesicular trafficking. Recent work suggests that formins catalyze the assembly of unbranched actin structures and nucleate actin filaments directly (Matusek, 2006).

Although all members of this highly conserved family are defined by the presence of the formin homology domain 2 (FH2), which is both necessary and sufficient to nucleate actin in vitro, formins are multi-domain proteins that contain several other conserved sequences as well. The FH2 domain is usually flanked on the N-terminal side by a proline-rich FH1 domain that can serve as a docking site for the G-actin-binding protein profilin. In addition, the FH1 domain also binds WW domains and SH3 domains, including those of the Src family. A distinct formin subfamily, the DRF group (Diaphanous related formins: see Drosophila Diaphanous) has the ability to interact with an activated Rho GTPase through an N-terminal GTPase-binding domain (GBD). This binding alleviates the autoinhibitory interaction between the GBD and the short C-terminal DAD domain (Diaphanous autoregulatory domain) (Alberts, 2001). Crystallographic analysis of the N-terminal part of mouse Dia1 provides a structural basis of this regulation and has identified several other functional domains within this part of the protein, including a dimerization domain (DD), (Otomo, 2005a; Rose, 2005). In addition, the crystal structure of C-terminal formin domains reveals that the FH2 domain also forms a dimer required for actin nucleation and processive filament capping (Otomo, 2005b; Shimada, 2004; Xu, 2004), strongly suggesting that native formins act in dimeric forms (Matusek, 2006).

Recently, a novel formin subtype, DAAM (Dishevelled-associated activator of morphogenesis), has been identified and implicated in planar cell polarity signaling during Xenopus gastrulation (Habas, 2001). The polarized orientation of cells within the plane of a tissue, or planar cell polarization (PCP), is an important aspect of cellular differentiation and is often necessary to the formation of functional organs. Genes controlling PCP have been extensively studied in Drosophila, revealing a crucial role for frizzled (fz) signaling through dishevelled (dsh) during the course of PCP establishment. The downstream components of the Fz/PCP pathway include RhoA, Drok, the JNK cascade and several other genes, most of which act tissue specifically. Many of the genes involved in PCP signaling are also required for polarized morphogenetic cell movements such as convergent extension during early vertebrate embryogenesis suggested that the novel FH2 protein Daam1 is required for convergent extension is Xenopus embryos and that Daam1 might function as a bridging factor between Dsh and RhoA. Moreover, Wnt/Fz-mediated activation of RhoA appeared to depend on Dvl (a Dsh homolog) and Daam1 (Habas, 2001). However, in contrast to this model, previous work has provided evidence that formins act as Rho effectors downstream of the Rho GTPases (Matusek, 2006).

To gain further insights into the function of this novel class of FH2 proteins, the single Drosophila member of the DAAM family was analyzed. Phenotypic analysis of DAAM mutants showed that it plays either no role or possibly a redundant role in PCP establishment in Drosophila. However, evidence was found that DAAM is involved in the regulation of the actin cytoskeleton in several different tissues, including the tracheal system. The Drosophila tracheal network is one of the best characterized model systems of branching morphogenesis. During the first phase of tracheal development, the primordial cells invaginate in each embryonic hemisegment from the epidermis, migrate and undergo cell shape changes to form the primary branches. Subsequently, some tracheal branches fuse with an adjacent branch to build up a continuous tubular network. This process is mediated by fusion cells located at the branch tips, which recognize each other in the adjacent hemisegments and become doughnut shaped, forming a lumen that connects the two branches. Before the end of embryogenesis, tracheal cells secrete a cuticle on their apical, luminal surface that protects the larvae from dehydration and infection. The tracheal cuticle is distinguished from the epidermal cuticle by the presence of cuticle ridges, often called taenidial folds that are thought to prevent the collapse of tracheal tubes while allowing them to expand and contract along their length (Matusek, 2006).

Drosophila DAAM is required to organize an array of parallel running actin cables beneath the apical surface of the tracheal cells that define the taenidial fold pattern of the cuticle. The actin ring pattern corresponds exactly to that of the taenidial fold pattern, and it is proposed that the actin rings organized by DAAM define the position of taenidial fold formation. The genetic interaction and epistasis data are consistent with a model that DAAM activity is regulated by RhoA. In addition, DAAM works together with the non-receptor tyrosine kinases Src42A and Tec29 to regulate the actin cytoskeleton of the Drosophila tracheal system (Matusek, 2006).

The basic structure of the insect tracheal system is a highly conserved tubular network in every species. The most important function of this network is to allow oxygen flow to target cells. Thus, tracheal tubes need to be both rigid enough, to ensure continuous air transport, and flexible enough along the axis of the tubes, to prevent the break down of the tube system when body parts or segments move relative to each other. These requirements are mainly ensured by the tracheal cuticle, which covers the luminal surface of the tubes and displays cuticle ridges (making the overall structure similar to the corrugated hose of a vacuum cleaner). Analysis of DAAM mutants provides the first direct evidence that this hypothesis is correct. The data demonstrate that in the absence of DAAM the taenidial fold pattern is severely disrupted, often leading to the collapse of the tubes and to discontinuities in the tubular network. In addition, the analysis revealed that the remarkably ordered cuticle pattern, displayed in the wild-type trachea tubes, depends on DAAM-mediated apical actin organization. Apical actin is organized into parallel-running actin cables, much the same way teanidial folds run in the cuticle. Significantly, the formation of these actin bundles precedes the onset of cuticle secretion, and the number and phasing of the actin rings correspond exactly to that of the taenidial folds in the cuticle. Thus these studies revealed a novel aspect of apical actin organization in the tracheal cells that has not been appreciated before (Matusek, 2006).

The DAAM gene encodes a novel member of the formin family of proteins, involved in actin nucleation and polymerization. Consistent with this, DAAM is colocalized with apical actin in the tracheal cells, and the activated form of DAAM is able to induce actin assembly when expressed in tracheal cells and in other cell types. In DAAM mutant tracheal cells, apical actin is still detected, albeit at a somewhat lower level than in wild type, but the bundles formed in the mutant are much shorter and thinner than in wild type, and often appear to be crosslinked to each other. Most strikingly, global actin organization is almost completely lost, although some local order can still be detected. Remarkably, the cuticle pattern in mutant tracheal cells still follows the underlying aberrant actin pattern. Overall, in DAAM mutants, both the tracheal cuticle and the apical actin pattern resemble a mosaic of locally ordered patches that failed to be coordinated and aligned with each other and the axis of the tracheal tubes. It is thus proposed that the apical actin bundles play a key role in patterning the tracheal cuticle by defining the place of taenidial fold formation. Regarding the function of DAAM, the results suggest that the major role of this formin in the tracheal cells is to organize the actin filaments into parallel running actin rings or spirals instead of simply executing the well characterized formin function related to actin assembly. However, whether this is a direct effect on actin organization, and thus represents a novel formin function, needs to be further elucidated. An alternative model could be that DAAM is primarily required for actin polymerization but tightly coupled to an actin 'organizing' protein. In such scenario, the polymerization activity should be a redundant requirement, whereas the link to the organizing protein would be a DAAM-specific function, thereby explaining the presence of unorganized actin bundles in DAAM mutant tracheal cells (Matusek, 2006).

In the case of the main tracheal airways, which are multicellular along their periphery, it is striking that in wild type the run of the actin bundles is perfectly coordinated across cell boundaries. In addition, the run is always perpendicular to that of the tube axis. How does DAAM ensure the coordination of these two aspects of actin organization? Because the DAAM protein and the apical actin cables are both found at the level of the adherens junctions, it is possible that DAAM regulates the coordination of the actin cables at the cell boundaries through a direct interaction with junctional protein complexes. However, other explanations are also possible, and further experiments will be required to elucidate the molecular mechanism of this regulatory function. The fact that actin cables normally run perpendicular to the tube axis seems to suggest that tracheal cells are able to sense a global orientation cue and align their actin bundles accordingly. The nature and source of this cue is unknown, as is the mechanism by which DAAM is involved in the read-out of this signal. Nevertheless, it is interesting that in DAAM and btl-Gal4/UAS-C-DAAM mutant trachea, the main pattern of the cuticle phenotype is often changing from one segment to the other, suggesting that the effect of the 'global' orientation cue is limited to metameric units (Matusek, 2006).

Sequence comparisons of FH2 proteins suggest a close phylogenetic relationship between the DRF, FRL and DAAM subfamilies (Higgs, 2005). Members of these three subfamilies have a high level of conservation in the FH2 domain, and importantly, also in the region of the GBD and DAD domains, suggesting that the FRL and DAAM family formins are also regulated by autoinhibition and RhoGTPases, like the DRFs. Further evidence is presented in support of this view. First, DAAM and RhoA display a strong genetic interaction. Second, C-DAAM (an N-terminally truncated form of DAAM) behaves like an activated form much the same way DRF family formins behave. Third, epistasis experiments with C-DAAM and RhoA suggest that RhoA acts upstream of DAAM. Thus, the data support the model in which DAAM, at least in the Drosophila tracheal system, is regulated by autoinhibition that can be relieved by RhoGTPases (Matusek, 2006).

This conclusion, however, contradicts the observation that human DAAM1 is required for Wnt/Fz/Dvl dependent RhoA activation in cultured cells and that xDaam1 appears to mediate Wnt-11 dependent RhoA activation in Xenopus embryos (Habas, 2001). These results suggested that DAAM functions upstream of RhoA in non-canonical Wnt/Fz-PCP signaling. An explanation for these distinct conclusions might be related to the fact that DAAM, in contrast to xDaam1, does not appear to be required for Fz/Dsh-PCP signaling. Hence, it is possible that the Drosophila ortholog is regulated in the same way as the DRF formins, while the vertebrate family members can be regulated in a different way, once bound by Dsh/Dvl and recruited into PCP signaling complexes (Matusek, 2006).

Genetic interactions with the hypomorphic DAAMEx1 allele identified two non-receptor tyrosine kinases, Src42A and Tec29, as strong interacting partners. Although both of these kinases play multiple roles during embryogenesis, single mutants for both affect the tracheal cuticle pattern in a similar way to DAAM. These results suggest that DAAM and the Src family kinases work together to regulate the actin cytoskeleton and cuticle pattern in tracheal cells. Although it is not known whether DAAM physically binds Src42A and/or Tec29, it has been established that the FH1 region of DRFs and other formins can bind SH3 domains, including those of the Src family kinases. In agreement with these data that DAAM acts upstream of Src42A and Tec29 in tracheal cells, cytoskeleton remodeling and SRF activation mediated by mouse Dia1 and mouse Dia2 requires Src activity. Moreover, a recent report suggests that RhoD and human DIA2C regulate endosome dynamics through Src activation, proposing that Src activity is stimulated via human DIA2C dependent recruitment to early endosomes (Gasman, 2003). Similarly, the Limb deformity protein (a formin) interacts with Src on the perinuclear membranes of primary mouse fibroblasts. Based on these examples, it is speculated that in Drosophila tracheal cells the RhoA/DAAM/Src module may not only be required to organize apical actin bundles, but additionally it might represent a link to secretory vesicles and to the regulation of exocytosis. Future studies will be required to test this hypothesis, and to unravel the mechanisms whereby DAAM family formins and Src family kinases contribute to cytoskeletal remodeling in the Drosophila tracheal system and in other tissues (Matusek, 2006).

The formin DAAM functions as molecular effector of the planar cell polarity pathway during axonal development in Drosophila

Recent studies established that the planar cell polarity (PCP) pathway is critical for various aspects of nervous system development and function, including axonal guidance. Although it seems clear that PCP signaling regulates actin dynamics, the mechanisms through which this occurs remain elusive. This study established a functional link between the PCP system and one specific actin regulator, the formin DAAM, which has previously been shown to be required for embryonic axonal morphogenesis and filopodia formation in the growth cone. DAAM also plays a pivotal role during axonal growth and guidance in the adult Drosophila mushroom body, a brain center for learning and memory. By using a combination of genetic and biochemical assays, it was demonstrated that Wnt5 and the PCP signaling proteins Frizzled, Strabismus, and Dishevelled act in concert with the small GTPase Rac1 to activate the actin assembly functions of DAAM essential for correct targeting of mushroom body axons. Collectively, these data suggest that DAAM is used as a major molecular effector of the PCP guidance pathway. By uncovering a signaling system from the Wnt5 guidance cue to an actin assembly factor, it is proposed that the Wnt5/PCP navigation system is linked by DAAM to the regulation of the growth cone actin cytoskeleton, and thereby growth cone behavior, in a direct way (Gombos, 2015).

This study has shown that DAAM plays an important role in the regulation of axonal growth and guidance of the Drosophila MB neurons. Several lines of evidence suggest that DAAM acts in concert with Wnt5 and the core PCP proteins to ensure correct targeting of the KC axons. DAAM functions downstream of Dsh and Rac1, and its ability to promote actin assembly is absolutely required for neural development in the MB. These data suggest a simple model in which axon guidance cues, such as Wnt5, signal through the PCP pathway to activate DAAM to control actin filament formation in the neuronal growth cone. Thus, PCP signaling appears to be linked to cytoskeleton regulation in a direct way, and these results provide compelling experimental evidence suggesting that, at least in neuronal cells, the major cellular target of PCP signaling is the actin cytoskeleton (Gombos, 2015).

Formins are highly potent actin assembly factors that are under tight regulation in vivo. The major mechanism of controlling the activity of the Diaphanous-related formin (DRF) subfamily involves an intramolecular autoinhibitory interaction between the N-terminal diaphanous inhibitory domain (DID) and the C-terminal Diaphanous autoinhibitory domain (DAD). This inhibition can be relieved upon binding of an activated Rho family GTPase that interacts with the GBD (GTP-ase binding domain)/DID region and also by proteins that bind to the DAD domain. Consistently, this study found that the Rac1 GTPase and the DAD domain binding Dsh protein both play role in DAAM activation in MB neurons. With this regard, it is notable that, despite that dsh1 is considered a PCP-null allele, the DAAMEx1, dsh1 double hemizygous mutants exhibit a stronger MB phenotype than dsh1 mutants alone, suggesting that DAAM must receive Dsh-independent regulatory inputs for which Rac1 is a prime candidate. Although previous work indicated that Rho GTPases might function downstream of Dsh in a linear pathway), the data suggest that Dsh and Rac1 act in parallel pathways in the MB. As the impairment of GTPase binding severely, but not completely, abolishes DAAM activity, it is concluded that Rac1 is likely to have a stronger contribution to DAAM activation in vivo; nonetheless, the simultaneous binding of Dsh appears to be required for full activation (Gombos, 2015).

Presumably, the most remarkable feature of the PCP system relies in its ability to create subcellular asymmetries. Therefore, it is a tempting idea that, upon guidance signaling, the PCP proteins are involved in the generation of molecular asymmetries within axonal growth cones, yet recent attempts failed to reveal such polarized distributions in MB neurons. Interestingly, however, it was shown that Fz and Vang display a differential requirement during development of the MBs, with Fz predominantly acting in the dorsal lobes and Vang predominantly acting in the medial lobes (MLs). This study found that, in contrast to Fz and Vang, DAAM plays a crucial role in both lobes of the MBs. Additionally, it was demonstrated that Fz promotes the formation of membrane-associated Dsh-DAAM complexes in S2 cells. This result, together with genetic data, suggests that DAAM acts as the downstream effector of a Fz/Dsh module, which is required for the correct growth and guidance of the dorsal MB axon branches (Gombos, 2015).

In addition to their potential connection to Fz signaling in the dorsal lobe, DAAM and Dsh were linked to Vang- and Wnt5-dependent ML development as well. Wnt5 and Vang have an identical effect on ML development when overexpressed, and this GOF phenotype can be suppressed by the same set of mutations (DAAM, dsh, Rac1). In particular, the putative PCP-null dsh1 allele and heterozygosity for Rac1 cause an almost equally strong, yet partial, suppression with regard to the ML fusion phenotype. This is best explained by assuming that Wnt5 and Vang signal both in a Dsh-dependent and in a Dsh-independent, but Rac-dependent, manner. With regard to DAAM, this study has shown that DAAM nearly completely suppresses the GOF of Wnt5 and Vang, and Dsh and Rac1 both contribute to DAAM activation. Collectively, these data suggest a model in which Wnt5 and Vang promote β lobe extension by signaling to Dsh and Rac1 that will activate DAAM in parallel to each other. The colocalization of Vang and DAAM, observed in S2 cells, indicates that they may bind each other directly, which would be in good accordance with genetic data suggesting a close functional link between DAAM and Vang during β lobe development. However, formins are not known to bind Vang proteins; therefore, an indirect interaction, mediated by Rac1, which has recently been shown to be bound and redistributed by Vangl2 in epithelial cell lines, appears a more likely possibility (Gombos, 2015).

As discussed above, and contrary to Vang, Fz does not appear to be required for ML development, or if anything, it might play an opposite role, as loss of fz leads to ML fusion in 16.1% of the lobes. This is a surprising observation at first glance as Wnt proteins are thought to activate members of the Fz receptor family, but former analysis of Wnt5 signaling during MB development also failed to reveal a Fz requirement in the β lobes. Instead, Wnt5 has been linked to other type of Wnt receptors, the Ryk/Derailed atypical tyrosine kinase receptors, which are known to be involved in axonal guidance in flies and vertebrates. In light of these results, it will be of future interest to analyze the Wnt5-Vang connection in the MB in more details and identify the Wnt5 receptor in this context (Gombos, 2015).

Consistent with the lack of lobe-specific requirement for dsh and DAAM, the current studies revealed that Dsh, DAAM, and Rac1 are used as common effector elements of a dorsal lobe-specific Fz-dependent signal and a Vang-dependent ML-specific signal. It follows that Dsh and DAAM are likely to take part in two types of PCP complexes. Although, in vitro, Dsh has the ability to interact with both Fz and Vang, the conclusion that Dsh functions downstream of Vang in the β lobes is markedly different from the classical PCP regulatory context in which the Fz/Dsh and Vang/Pk complexes have opposing effects. Thus, this result, together with the Wnt5-Vang data, substantiates the earlier findings that the PCP system operates at least partly differently in neurons than during tissue polarity signaling (Gombos, 2015).

During PCP signaling, the vertebrate DAAM orthologs control convergence and extension movements, polarized cell movements during vertebrate gastrulatio. In contrast, DAAM is dispensable for classical planar polarity establishment in flies, suggesting that the tissue polarity function of DAAM might be restricted only to vertebrates. Despite the lack of direct function in establishing tissue polarization, this study provides evidence that DAAM is linked to the PCP pathway in another important regulatory context, notably directed neuronal development in the adult brain. Consistent with the results, recent studies revealed that PCP signaling and DAAM regulate neural development in planarians and in Xenopus embryos. Given that the vertebrate PCP proteins are known to be involved in multiple aspects of CNS development, and the vertebrate DAAM orthologs are strongly expressed in the CNS, it is conceivable that the PCP/DAAM module represents a highly conserved regulatory system that is used to regulate various aspects of neuronal development throughout evolution (Gombos, 2015).


GENE STRUCTURE

cDNA clone length - 5369 (isoform b)

Bases in 5' UTR - 1168

Exons - 8

Bases in 3' UTR - 739

PROTEIN STRUCTURE

Amino Acids - 1153 (isoform b)

Structural Domains

A recently published phylogenetic analysis (based on sequence comparisons of 101 FH2 domains from 26 eukaryotic species) has concluded that metazoan formins can be grouped into seven major subclasses (Higgs, 2005). This analysis suggested that the DAAM subfamily clearly represents a distinct class. This is consistent with the previous finding that the DAAM subfamily exhibits extensive sequence similarity both within and outside of their highly conserved FH1 and FH2 domains (Habas, 2001). At present, however, very little is known about the in vivo function of the DAAM subfamily. Sequence analysis revealed the presence of a single DAAM ortholog in the Drosophila genome corresponding to the annotated gene CG14622. Although FlyBase predicts that this gene might code for several different transcript classes, this work focused on the predicted CG14622-RB transcript that appears to be encoded by the RE67944 EST clone. Sequencing of this full-length cDNA clone indicated a protein consisting of 1153 amino acids. Further analysis revealed that the ORF contains several conserved domains including an FH1, FH2, GBD and a putative DAD domain characteristic of formins. Based on the overall homology level and the phylogenetic analysis of FH2 domains (Higgs, 2005), the DAAM subfamily appears to be most related to the Dia subfamily, raising the possibility that similar to the DRFs, DAAM formins are also regulated by an autoinhibitory mechanism that can be relieved upon RhoGTP binding (Matusek, 2006).


Dishevelled Associated Activator of Morphogenesis: Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References

date revised: 11 September 2006

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