Suppressor of profilin 2


Promoter Structure

The developmental pattern of expression of the genes encoding the catalytic (alpha) and accessory (beta) subunits of mitochondrial DNA polymerase (pol gamma) has been examined in Drosophila melanogaster. The steady-state level of pol gamma-beta mRNA increases during the first hours of development, reaching its maximum value at the start of mtDNA replication in Drosophila embryos. In contrast, the steady-state level of pol gamma-alpha mRNA decreases as development proceeds and is low in stages of active mtDNA replication. This difference in mRNA abundance results at least in part from differences in the rates of mRNA synthesis. The pol gamma genes are located in a compact cluster of five genes that contains three promoter regions (P1-P3). The P1 region directs divergent transcription of the pol gamma-beta gene and the adjacent rpII33 gene. P1 contains a DNA replication-related element (DRE) that is essential for pol gamma-beta promoter activity, but not for rpII33 promoter activity in Schneider's cells. A second divergent promoter region (P2) controls the expression of the orc5 and sop2 genes. The P2 region contains two DREs that are essential for orc5 promoter activity, but not for sop2 promoter activity. The expression of the pol gamma-alpha gene is directed by P3, a weak promoter that does not contain DREs. Electrophoretic mobility shift experiments demonstrate that the DRE-binding factor (DREF) regulatory protein binds to the DREs in P1 and P2. DREF regulates the expression of several genes encoding key factors involved in nuclear DNA replication. Its role in controlling the expression of the pol gamma-beta and orc5 genes establishes a common regulatory mechanism linking nuclear and mitochondrial DNA replication. Overall, these results suggest that the accessory subunit of mtDNA polymerase plays an important role in the control of mtDNA replication in Drosophila (Lefai, 2000).



To determine the role of the Arp2/3 complex during cell cycle-regulated actin reorganization in syncytial blastoderm stage Drosophila embryos, antibodies were prepared to the Drosophila Arpc1 and Arp3 subunits. The Drosophila Arpc2 protein, predicted from the genomic sequence, is very similar to human Arpc2, suggesting that antibodies directed against the human protein would react with the fly homolog. This antibody and the anti-Drosophila Arp1c and Arp3 antibodies recognize major polypeptides of the appropriate apparent molecular mass on whole-embryo Western blots. In addition, the three proteins recognized by these antibodies cosediment at approximately 9S on sucrose gradient fractionation, consistent with the behavior of the Arp2/3 complex in other systems. It is concluded that these antibodies recognize components of the Drosophila Arp2/3 complex, which is similar in native size and shape to Arp2/3 in other systems (Stevenson, 2002).

To determine the subcellular localization of the Arp2/3 complex, these antibodies were used to immunolabel syncytial embryos. The anti-Drosophila Arpc1 antibody did not produce a signal above background, but the remaining two antibodies produced similar labeling patterns. Results obtained with the anti-Drosophila Arp3 antibody are presented, since this reagent shows the highest specificity on Western blots. During the premigration nuclear divisions, f-actin is organized around centrosomes. Arp3 localizes to distinct particles that are also concentrated around the centrosomes. A subset of Arp3 particles have f-actin 'tails' reminiscent of the comet tails associated with intracellular pathogens. However, the majority of Arp3 particles do not have actin tails and only partially colocalize with f-actin (Stevenson, 2002).

Following nuclear migration and formation of the syncytial blastoderm embryo, cortical actin proceeds through well-characterized cycles of cap and pseudocleavage furrow assembly. Very early in interphase, f-actin first accumulates in small, poorly organized caps. At this time, Arp3 accumulates in cortical bands that partially surround these small actin caps. This is a transient organization, however, and Arp3 shows pronounced accumulation at the cap margins as soon as they being to expand. The majority of f-actin in the caps does not colocalize with Arp3 but is displaced toward the interior of the caps. During mitosis, Arp3 and actin are enriched in the pseudocleavage furrows. Within the furrows, most of the Arp3 particles do not precisely colocalize with f-actin. These observations are consistent with a function for Arp2/3 in cap expansion and furrow assembly (Stevenson, 2002).

During interphase of the 14th nuclear division cycle, actin-dependent invagination of the plasma membranes incorporates the cortical nuclei into individual cells. These cells subsequently undergo conventional cleavage and form f-actin-based contractile rings. During cellularization, Arp3 is enriched at the apical surface of the embryo but is not found at the leading edge of the invaginating furrows. During the postcellularization divisions, by contrast, Arp3 shows striking colocalization with f-actin at the contractile rings. These observations raise the possibility that Arp2/3 has a direct role in conventional cleavage furrow assembly but may have a secondary function during cellularization (Stevenson, 2002).


To determine the subcellular distribution of Arp2/3 complex components, both GFP fusion constructs and antibodies were generated. Transgenes encoding Arpc1 and Arp3 fused to GFP were generated, and their distribution in the germline cells of the ovary was examined. These fusion proteins were abundant in the cytoplasm of germ cells, and both GFP fusion proteins were enriched at ring canals. In addition, some enrichment on cortical membranes was present. At higher magnification, both GFP fusions precisely colocalize with F-actin at the ring canal. No enrichment was observed of either GFP fusion protein on the cytoplasmic actin bundles that form in the nurse cells at stage 10B (Hudson, 2002).

Initial attempts at immunolocalization of the Arp2/3 complex using antibodies against Arpc1 or Arp3 were unsuccessful, since antisera generated were only useful for immunoblotting. More informative results were obtained with antibodies recognizing one of the two Arpc3 subunits. The two Drosophila Arpc3 subunits are 74% identical and 83% similar to one another. Antisera against GST fusion proteins of Arpc3A and Arpc3B were generated. All rats immunized with GST-Arpc3A produced antibodies that strongly recognize a His-tagged Arpc3A fusion protein and also show some affinity for a His-tagged Arpc3B fusion protein (Hudson, 2002).

When crude Arpc3A and purified Arpc3B antibodies were used to probe blots of ovarian extracts, prominent bands of ~21 and 20 kD, respectively, were observed. These apparent molecular weights were consistent with the predicted molecular weights of the two proteins: 20.3 kD for Arpc3A and 19.9 kD for Arpc3B. The tissue distribution of these proteins was analyzed by probing blots of extracts derived from whole or dissected adult flies, revealing that the Arpc3A band is present in essentially equal amounts in all tissues tested. Antibodies raised against Drosophila Arpc1 and Arp3 show a similar ubiquitous distribution. In contrast, immunoblotting with the Arpc3B antibody shows a germline-specific pattern of expression. A prominent band is present in ovarian lysates but is absent from the female carcasses lacking ovaries and also absent from lysates of whole females that lack germ cells. No expression was observed in extracts of whole males or isolated testes (Hudson, 2002).

Immunofluorescence using Arpc3B antibodies revealed that Arpc3B is highly enriched at ring canals throughout oogenesis. Arpc3B first appears on ring canals very early in the germarium, perhaps as early as region 2A . This is the stage where ring canals first become distinct from their structural precursors, the arrested cytokinetic furrows. Arpc3B appears to colocalize with actin at ring canals. At high magnification, the distribution of Arpc3B on ring canals appears punctate compared with antibodies against phosphotyrosine, another ring canal marker (Hudson, 2002).

Thus, GFP fusions with Arpc1 and Arp3 and antibodies specific for Arpc3B were used to study the subcellular localization of the Arp2/3 complex. The GFP fusion proteins reveal a subcellular distribution not unlike what has been observed for the Arp2/3 complex in other systems: a diffuse cytoplasmic distribution and an enrichment at certain regions of active actin assembly, in this case ring canals (Hudson, 2002).

Based on immunoblotting, expression of Arpc3B in adults is restricted to the ovaries of female flies, whereas Arpc3A is ubiquitously expressed. Immunofluorescence using Arpc3B antibodies reveals that it is abundant on ring canals in the nurse cells and is present at low levels in the cytoplasm of these cells. It is speculated that Arpc3B may be a component of a specialized Arp2/3 complex that is required for actin assembly at ring canals and that Arpc3A may be a component of a more broadly distributed Arp2/3 complex. Thus, the subcellular distribution observed with the Arp3 and Arpc1 fusion proteins may represent two distinct populations of Arp2/3 complex. The ring canal Arp2/3 complex would be expected to contain Arpc3B, whereas the cytosolic Arp2/3 complex would contain Arpc3A (Hudson, 2002).

Effects of Mutation or Deletion

Wiskott-Aldrich Syndrome proteins (WASp) serve as important regulators of cytoskeletal organization and function. These modular proteins, which are well-conserved among eukaryotic species, act to promote actin filament assembly in response to cues from various signal transduction pathways. Genetic analysis has revealed a requirement for the single Drosophila homolog, WASp, in cell-fate decisions governing specific neuronal lineages. This unique developmental context was used to assess the contributions of established signaling and cytoskeletal partners of WASp. Biochemical and genetic evidence is presented that, as expected, Drosophila WASp performs its developmental role via the Arp2/3 complex (see Drosophila Arp2/3 component Suppressor of profilin 2,), indicating conservation of the cytoskeletal aspect of WASp function in vivo. In contrast, association with the key signaling molecules CDC42 and PIP2 is not an essential requirement, implying that activation of WASp function in vivo depends on additional or alternative signaling pathways (Tal, 2002).

Evidence presented in this study suggests that the role of WASp in cell fate determination in neural lineages involves established cytoskeletal partners of WASp, and in particular, the Arp2/3 protein complex. Binding studies demonstrate a capacity for WASp to directly associate with monomeric actin via WA, the C-terminal cytoskeleton-interacting domain present in all WASp and WASp-related proteins. In parallel, the WA domain of WASp is shown to interact with components of the Arp2/3 complex, the primary downstream target of signal transduction pathways operating through WASp family proteins. The in vivo significance of these associations, which are characteristic of WASp elements in general, is demonstrated by a dual genetic approach. The final 30 residues at the C-terminal end of the WA domain of WASp prove necessary for rescue of WASp mutant phenotypes, while mutations in the Arp2/3 complex subunit Arpc1 lead to cell-fate transformations and neuronal excess during sensory organ development, a distinct, WASp-like phenotype. Taken together with the binding studies, these genetic observations imply that engagement of the cytoskeletal machinery via the C-terminal WA domain is an essential aspect of WASp function in vivo (Tal, 2002).

A genetic experiment was designed to determine whether Arp2/3 function is required for the WASp-mediated process of sensory organ development. Toward this end, use was made of strong loss-of-function alleles of Arpc1, the Drosophila homolog of the gene encoding the ARPC1/p41 subunit of the Arp2/3 complex. Unlike WASp, zygotic mutants for Arpc1 do not survive to adult stages. Use was made of the eyeless-FLP-FRT system to generate mosaic Arpc1 heterozygous flies, in which head capsule structures and cuticle are derived from large homozygous mutant clones induced in the eye imaginal disc. Compared with wild type, Arpc1 mosaic heads exhibit a pronounced loss of external sensory organ structures, which include both the bristle-shafts and the socket structures from which they emanate. This smooth cuticle phenotype is a highly characteristic feature of WASp mutant animals. To establish the developmental basis for the Arpc1 bristle-loss phenotype, pupal mosaic heads were dissected and immunostained using two informative sensory organ markers: Suppressor-of-Hairless [Su(H)], which specifically accumulates in socket cells, and the neuronal-specific nuclear antigen ElaV. In wild-type flies, each sensory organ lineage gives rise to five distinct cell types, including a single neuron and a single socket cell. In Arpc1 mosaics, a clear preponderance of ElaV-positive neurons is observed, coupled with a marked decrease in the number of cells expressing the socket cell marker Su(H), highly similar to the cell-fate transformation characteristic of WASp mutants. An abnormal, WASp-like distribution of cell fates, in which excess neurons develop at the expense of other cell types, is therefore observed within the sensory organ lineages of Arpc1 mosaics. This finding significantly supports the assertion that the functional requirement for Drosophila WASp during cell-fate specification is mediated via the Arp2/3 complex, and is thus likely to involve reorganization of the actin-based cytoskeleton (Tal, 2002).

Previous work (Fyrberg, 1993; Fyrberg, 1994) and the sequencing of the Drosophila genome has revealed apparent homologs of each of the seven subunits of the Arp2/3 complex. Single isoform homologs of each subunit are encoded, except for Arpc3 (p21) for which two Drosophila genes display a high degree of homology. Following the nomenclature adopted by the field (Higgs, 2001), these two genes have been designated Arpc3A and Arpc3B (Hudson, 2002).

Loss of function mutations were identified in the genes encoding the Arpc1 and Arp3 subunits. Identification of Arpc1 mutations was facilitated by the location of Arpc1 near the Adh gene, which resides within an extensively characterized region on the left arm of chromosome 2. The saturation mutagenesis and genetic mapping studies allowed the mapping of the Arpc1 gene to a region containing several lethal complementation groups within a genetic interval defined by Df(2L)b84A9. A genomic transgene containing the Arpc1+ genomic region, constructed for use in an independent study, was used to rescue the lethality associated with the l(2)34DdCH60 lethal mutation. Sequencing of the l(2)34DdCH60 allele revealed a 207-bp genomic deletion that removes the last 62 codons of Arpc1; therefore, this mutation was designated as Arpc1CH60 (Hudson, 2002).

Since Arpc1CH60 was the only allele of Arpc1 extant at the time of this study, and this allele was likely to be hypomorphic, an ethyl methane sulfonate noncomplementation screen was performed to isolate additional Arpc1 alleles. Five new Arpc1 alleles were identified. Genomic DNA from the mutant chromosomes was sequenced, and molecular lesions for each allele were identified. Three alleles, Q25sd, Q25st, and W82st, were predicted to truncate the protein before the second WD repeat; these alleles behaved as null mutations. Another allele, W108R, had a missense mutation changing Trp108 to Arg, and this allele exhibited phenotypes of moderate strength. The weakest allele, R337st, truncated the protein in the last WD repeat (Hudson, 2002).

A mutation in the Drosophila Arp3 gene was identified based on the Berkeley Drosophila Genome Project P-element disruption project. The EP element in EP(3)3640 was inserted 138 bp upstream of the predicted initiating ATG methionine codon. The lethality associated with the EP-element insertion could be reverted by excising the P element. In addition, the lethality of the EP(3)3640 insertion could be rescued by ubiquitous expression of the Gal4 transcriptional activator; since the EP(3)3640 insertion is oriented with the Gal4 UAS directed toward Arp3, ubiquitous Gal4 expression presumably restores expression of Arp3 to viable levels. Based on these observations, the EP(3)3640 insertion has been designated as Arp3EP(3)3640 (Hudson, 2002).

The spectrum of phenotypes exhibited by Arpc1 and Arp3 mutations is identical with respect to several different actin structures in several cell types (Hudson, 2002).

All of the alleles in Arp2/3 complex genes result in lethality before the adult stage; therefore, the phenotypes of Arp2/3 complex mutations were examined in clones of mutant cells in specific tissue types. To learn whether the Arp2/3 complex was required during oogenesis, germline clones were generated using the FLP/FRT/DFS technique, which allows the identification of egg chambers with mutant germline cells after stage 8 of oogenesis. Germline mosaics using Arp2/3 mutations revealed a requirement for the Arp2/3 complex in cytoplasm transport. Late stage egg chambers contained abnormally small oocytes and large residual nurse cells (Hudson, 2002).

Closer examination of the Arpc1 and Arp3 mutant egg chambers reveals defective ring canals. Visualization of actin with fluorescent phalloidin and ring canals with Hts-RC antibodies shows that ring canal size and integrity are severely compromised; this phenotype is 100% penetrant. The effect on ring canals is heterogeneous; often, the four ring canals connecting the oocyte to the nurse cells appear less severely affected than ring canals connecting nurse cells to other nurse cells. Quantification of the effect on ring canal size reveals that at stage 10A, ring canals between the oocyte and adjacent nurse cells are on average 30% smaller in diameter in Arpc1 or Arp3 mutants than in wild type. Ring canals connecting pairs of nurse cells are <50% the size of wild-type ring canals in Arpc1 and Arp3 germline clones (Hudson, 2002).

Given the striking effect on ring canal integrity observed in late stage egg chambers, egg chambers at earlier stages were examined. Since the FLP/FRT/DFS method does not allow examination of mutant egg chambers before stage 8 of oogenesis, clones were generated using green fluorescent protein (GFP) as a marker. In this system, mutant cells were identified by loss of GFP expression. Surprisingly, ring canal development appears to initiate normally. In the germarium, ring canals in Arpc1 mutant clones are indistinguishable from wild type, and it is not possible to detect differences between mutant and wild type through at least stage 4 of oogenesis. By stages 5-6, differences between ring canals in Arpc1 mutant egg chambers and those in comparably staged wild-type egg chambers could be discerned. Actin associated with Arpc1 ring canals is less robust, and ring canals appear smaller in diameter. By stage 8, there appears to be a reduction in ring canal actin relative to wild type, and some ring canals were misshapen. By stages 9-10, ring canals appear identical to those seen in germline clones generated using the FLP/FRT/DFS technique. Thus, the Arp2/3 complex is not required for the initial recruitment of actin to ring canals but rather becomes necessary for later stages of ring canal growth (Hudson, 2002).

In later stages, ring canals sometimes are detached from the nurse cell membranes, and the nurse cell subcortical actin becomes destabilized. This apparently results in the formation of multinucleate cells. Multinucleate cells are never observed in younger egg chambers and thus are unlikely to result from defective cytokinesis. Furthermore, all clonal egg chambers contain 16 germline cells, indicating that Arp2/3 is not required for germline cell division (Hudson, 2002).

Completion of cytoplasm transport from the nurse cells to the oocyte is dependent on the formation of dense networks of parallel hexagonally packed actin bundles that form in nurse cell cytoplasm just before the contraction that drives final cytoplasm transport. Germline clones of null alleles of Arpc1 or the strong hypomorph Arp3EP(3)3640 produce abundant cytoplasmic actin bundles. The relative abundance of bundles is comparable to wild type. The bundles present in mutant nurse cells are also striated in appearance as they are in wild type. The distribution of the bundles are sometimes irregular: in contrast to wild type, where the bundles are evenly distributed around the nuclei, the bundles in Arpc1 and Arp3 mutant nurse cells tend to be unevenly clustered. This may be a secondary consequence of severely damaged ring canals and an unstable plasma membrane cytoskeleton (Hudson, 2002).

Proper extension of the mechanosensory bristles is also dependent on the polymerization of parallel hexagonally packed actin bundles. To examine the consequences of loss of Arp2/3 activity in developing bristles, null alleles of Arpc1 were used to generate mosaic flies lacking Arpc1 in bristle precursor cells. The mature Arpc1 bristles are indistinguishable from wild type at low magnification. However, at higher magnification, alterations in bristle morphology are apparent. Compared with wild type, the Arpc1 mutant bristles have approximately twice the number of longitudinal ridges. This phenotype is observed in all mutant macrochaetes. Arp3EP(3)3640 homozygous animals die as pharate adults so the Arp3 bristle phenotype was examined after dissecting Arp3 adults from their pupal cases. Bristle morphology appears normal at lower magnification as is the case for Arpc1 mutant bristles. At higher magnification, alterations in the ridging pattern identical to those observed for Arpc1 are observed. The penetrance of this phenotype is ~80% (Hudson, 2002).

Therefore, the phenotypes of the Arp3EP(3)3640 mutation mirror those seen with the Arpc1 mutations. The simplest interpretation of these observations is that in either case the consequences of loss of Arp2/3 complex activity are being examined and that neither subunit is required for functions apart from their shared function in the Arp2/3 complex. Reconstitution experiments have recently been reported, showing that Arp2/3 complexes lacking Arp3 or Arpc1 have little or no nucleation activity (Gournier, 2001), supporting the notion that mutations in these subunits abolish Arp2/3 activity (Hudson, 2002).

The onset of ring canal defects in Arp2/3 mutant egg chambers occurs relatively late in oogenesis. Interestingly, the stage at which defects are first apparent coincides with a transition in the mode of actin filament addition. The early mode of actin filament addition to ring canals involves both the expansion of the ring canal diameter and an increase in the number of filaments in cross-section. This pattern persists until stage 5 of oogenesis where the ring canals are ~5 µm in diameter and reach a maximum of ~700 filaments in cross-section. Beginning at stage 5, the pattern of actin filament addition shifts so that the diameter of the ring canal continues to expand, whereas the number of filaments in cross-section remains constant (Hudson, 2002).

One possible explanation for the late onset of the Arp2/3 complex phenotypes is that the shift in the pattern of actin filament addition reflects an underlying mechanistic difference. An Arp2/3-independent mechanism may contribute to actin filament accumulation early in oogenesis when the number of filaments in cross-section is increasing. This Arp2/3-independent mechanism could be related to the process by which actin filaments are recruited to cleavage furrows where preexisting filaments from the cortical actin cytoskeleton are recruited to the rings to support their expansion. From stage 5 of oogenesis onward, the Arp2/3 complex may be required to provide the additional F-actin required to support expansion of the ring canal diameter (Hudson, 2002).

Localization of Arp2/3 components to the ring canals is consistent with a requirement for the Arp2/3 complex during ring canal expansion. However, it is curious that Arp2/3 complex components are found localized to or enriched at ring canals much earlier than the loss of function phenotypes becomes evident. One possible explanation for this is that the Arp2/3 complex does contribute to actin accumulation during earlier stages of oogenesis, but the Arp2/3-independent mechanism is functionally redundant with the Arp2/3 complex during this time. Alternatively, the Arp2/3 complex may associate with ring canals in an inactive state and become activated when required later in oogenesis (Hudson, 2002).

The only known product of Arp2/3 nucleation is an orthogonal network of branching filaments reminiscent of the filaments at the leading edge of a lamellipodium. However, the actin filaments in ring canals have been described at the ultrastructural level as loosely packed parallel actin bundles. A branching F-actin network could be transformed into parallel bundles through the action of F-actin bundling proteins present at the ring canal; these include Filamin, the product of the cheerio gene and Kelch, which binds and bundles F-actin (Hudson, 2002).

The requirement for Arp2/3 during ring canal growth raises the interesting possibility that new actin polymerization at the ring canal plasma membrane drives ring canal growth. There are numerous examples of actin filament-driven membrane movement, including the motility of Listeria and viruses within cells and the advancement of the leading edge membrane in motile cells. By analogy, the ring canal with its dynamic actin cytoskeleton may be equivalent to a circular leading edge membrane. Instead of being confined to one end of a cell, the Arp2/3-dependent actin is positioned between two cells in a continuous expanding ring. To drive expansion of the ring canal diameter, new actin polymerization must be directed circumferentially around the ring (Hudson, 2002).

Assembly of maximally crosslinked parallel bundles of actin filaments is a conserved process that occurs in tissues requiring the structural support of the actin cytoskeleton. These actin bundles are highly ordered and are assembled into a paracrystalline state through the action of multiple actin crosslinking proteins. Intestinal microvilli, stereocilia, developing Drosophila bristles, and Drosophila ovarian nurse cells all require parallel actin bundles. In the case of Drosophila bristles and nurse cells, the long actin bundles are made up of short, 2-3 µm modules of actin filament bundles; the striated appearance of the nurse cell bundles in wild-type and mutant nurse cells in reflects the regions where there is little overlap in these short bundles. The growing ends of the filaments in both nurse cell and bristle bundles are located at the plasma membrane, and it has been proposed that these filaments are nucleated by factors present at the plasma membrane analogous to the dense material at the tip of microvilli. It is striking that the nurse cell cytoplasmic actin bundles are able to form in Arp2/3 complex mutants, and the striated appearance of the phalloidin-labeled bundles indicates that the mutant bundles are likely wild-type in ultrastructural organization. In addition, the finding that bristles are able to extend in Arp2/3 mutants provides further evidence that parallel actin bundles do not require Arp2/3 complex function. It is concluded that the Arp2/3 complex is not involved in the nucleation of actin filaments for microvillus-type actin filament bundles; rather, an alternative nucleation factor may be involved or other mechanisms, such as filament uncapping, may play a role (Hudson, 2002).

The phenotype observed in Arp2/3 mutant bristles is intriguing. Although Arp2/3 mutant cells can produce bristle extensions, the mutant bristles have twice as many ridges as in wild type. The ridges are formed between the actin bundles in the developing bristle; thus, the valleys on a mature bristle mark the location of the actin bundles during bristle development. In Arp2/3 mutants, there is an apparent increase in the number of F-actin bundles, suggesting that bundle organization requires the Arp2/3 complex. Actin filaments for the parallel actin bundles are nucleated at the tips of growing bristles, and the newly polymerized filaments are initially organized into ~50 small bundles near the bristle tip. These bundles subsequently aggregate into 7 to 10 larger bundles that mark the positions of the ridges in the mature bristle. In Arp2/3 mutants, the number of ridges is increased, suggesting a specific defect in the bundle aggregation process. Perhaps a distinct population of actin filaments nucleated by the Arp2/3 complex is required to facilitate the aggregation of the parallel actin bundles in developing bristles (Hudson, 2002).

The observations presented in this study begin to define the specific types of actin rearrangements that require the Arp2/3 complex in metazoan cells. Strikingly, no evidence is found for a requirement of the Arp2/3 complex in the formation of parallel actin bundles. This result suggests that other cytosolic factors are required to nucleate actin polymerization for such structures. Alternatively, these results may highlight the importance of other mechanisms for initiating new polymerization, such as severing or uncapping of existing filaments (Hudson, 2002).

The Arp2/3 complex is required in the development of ovarian ring canals and mechanosensory bristle morphology. Both of these developmental processes offer excellent model systems in which to study the function of actin regulatory proteins, and further studies of how the Arp2/3 complex interacts with other actin binding proteins to shape the actin cytoskeleton in vivo are being pursued (Hudson, 2002).


Balasubramanian, M. K., et al. (1996). Fission yeast Sop2p: a novel and evolutionarily conserved protein that interacts with Arp3p and modulates profilin function. EMBO J. 15(23): 6426-37. 8978670

Feoktistova, A., McCollum, D., Ohi, R. and Gould. K. L. (1999). Identification and characterization of Schizosaccharomyces pombe asp1+, a gene that interacts with mutations in the Arp2/3 complex and actin. Genetics 152(3): 895-908. 10388810

Fyrberg, C. and Fyrberg, E. (1993). A Drosophila homologue of the Schizosaccharomyces pombe act2 gene. Biochem. Genet. 31: 329-341. 8274139

Fyrberg, C., et al. (1994). Genes encoding actin-related proteins of Drosophila melanogaster. J. Mol. Biol. 241: 498-503. 8064864

Gournier, H., et al. (2001). Reconstitution of human Arp2/3 complex reveals critical roles of individual subunits in complex structure and activity. Mol. Cell. 8: 1041-1052. 11741539

Higgs, H. N., and Pollard, T. D. (2001). Regulation of actin filament network formation through the Arp2/3 complex: activation by a a diverse array of proteins. Annu. Rev. Biochem. 70: 649-676. 11395419

Hudson, A. M. and Cooley, L. (2002). A subset of dynamic actin rearrangements in Drosophila requires the Arp2/3 complex. J. Cell Biol. 156: 677-687. 11854308

Lefai, E., Fernandez-Moreno, M. A., Alahari, A., Kaguni, L. S. and Garesse, R. (2000). Differential regulation of the catalytic and accessory subunit genes of Drosophila mitochondrial DNA polymerase. J. Biol. Chem. 275(42): 33123-33. 10930405

Machesky, L. M., et al. (1994). Purification of a cortical complex containing two unconventional actins from Acanthamoeba by affinity chromatography on profilin-agarose. J. Cell Biol. 127(1): 107-15. 7929556

Machesky, L. M., et al. (1997). Mammalian actin-related protein 2/3 complex localizes to regions of lamellipodial protrusion and is composed of evolutionarily conserved proteins. Biochem J. 328: 105-112. 9359840

Mullins, R. D., Stafford, W. F. and Pollard, T. D. (1997). Structure, subunit topology, and actin-binding activity of the Arp2/3 complex from Acanthamoeba. J. Cell Biol. 136(2): 331-43. 9015304

Robinson, R. C., et al. (2001). Crystal structure of Arp2/3 complex. Science 294(5547): 1679-84. 11721045

Sawa, M., et al. (2003). Essential role of the C. elegans Arp2/3 complex in cell migration during ventral enclosure. J. Cell Sci. 116: 1505-1518. 12640035

Stevenson, V. A., Kramer, J., Kuhn, J., and Theurkauf, W. E. (2001). Centrosomes and the Scrambled protein coordinate microtubule- independent actin reorganization. Nat. Cell Biol. 3: 68-75. 11146628

Stevenson, V., Hudson, A., Cooley, L. and Theurkauf, W. E. (2002). Arp2/3-Dependent Psuedocleavage Furrow Assembly in Syncytial Drosophila Embryos. Curr. Biol. 12: 705-711. 12007413

Tal, T., Vaizel-Ohayon, D. and Schejter, E. D. (2002). Conserved interactions with cytoskeletal but not signaling elements are an essential aspect of Drosophila WASp function. Dev. Biol. 243(2): 260-71. 11884035

Weaver, A. M., et al. (2002). Interaction of cortactin and N-WASp with Arp2/3 complex. Curr. Biol. 12(15): 1270-8. 12176354

Welch, M. D., DePace, A. H., Verma, S., Iwamatsu, A. and Mitchison, T J. (1997). The human Arp2/3 complex is composed of evolutionarily conserved subunits and is localized to cellular regions of dynamic actin filament assembly. J. Cell Biol. 138(2): 375-84. 9230079

Winter, D., Podtelejnikov, A. V., Mann, M. and Li, R. (1997). The complex containing actin-related proteins Arp2 and Arp3 is required for the motility and integrity of yeast actin patches. Curr. Biol. 7(7): 519-29. 9210376

Winter, D. C., Choe, E. Y. and Li, R. (1999). Genetic dissection of the budding yeast Arp2/3 complex: a comparison of the in vivo and structural roles of individual subunits. Proc. Natl. Acad. Sci. 96: 7288-7293. 10377407

Zalevsky, J., Grigorova, I. and Mullins, R. D. (2000). Activation of the Arp2/3 complex by the Listeria acta protein. Acta binds two actin monomers and three subunits of the Arp2/3 complex. J. Biol. Chem. 276(5): 3468-75. 11029465

Zallen, J. A., et al. (2002). SCAR is a primary regulator of Arp2/3-dependent morphological events in Drosophila. J. Cell. Biol. 156: 689-701. 11854309

Suppressor of profilin 2 : Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation

date revised: 20 January 2004

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