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Gene name - Abl oncogene Synonyms - Cytological map position - 73B3--73B4 Function - signaling protein Keyword(s) - cytoskeleton, axonogenesis, oncogene |
Symbol - Abl FlyBase ID: FBgn0000017 Genetic map position - 3-[44] Classification - Protein tyrosine kinases Cellular location - cytoplasmic |
The Philadelphia chromosomal translocation is responsible for the fusion of two genes, ABL and BCR. Recognized as a frequently occuring aberrant chromosome in acute lymphoblastic leukemia, it was understood more than a decade ago that the Philadelphia chromosome occurs as a result of a recombination between two genes: the cellular ABL gene on chromosome 9, and BCR (breakpoint cluster region) gene located on chromosome 22. The Drosophila Abelson (Abl) protein is a homolog of mammalian c-Abl the wild-type gene coding for the ABL sequence found in the BRC/ABL hybrid protein. The BCR/ABL oncogene product, derived from this specific chromosomal translocation is present in Philadelphia chromosome-postive acute lymphoblastic leukemia. Despite its homology to c-Abl, the wild type mammalian protein, Drosophila ABL protein shows distinct properties and functional differences from c-Abl that suggest the two proteins are not strictly comparable. Notably, the Drosophila protein shows no nuclear localization; this suggests that the functions listed below, found for the mammalian protein, are absent in Drosophila:
Assuming that no other Abl type proteins are found in Drosophila, and no nuclear function can be found for Drosophila Abl, Abl can be used to illustrate the extent of an evolutionary divergence of nuclear function.
Nevertheless, the cytoplasmic functions of Abl appear to be conserved. Both fly and mammalian proteins have a conserved actin interaction domain and both proteins show genetic or physical interaction with a conserved protein known as Disabled (Dab). The Drosophila Disabled protein colocalizes with Abl in cell bodies and axons of embryonic CNS neurons. Dab is essential for normal CNS development, even in the presence of Drosophila Abl. Dab is tyrosine phosphorylated in insect cells and given its co-localization with Abl in the CNS, it has been suggested that Dab may be a physiological substrate of Abl. Identification of Abl substrates is important in building a picture of the role of Abl in the CNS. Drosophila Abl is required in disabled heterozygotes after the time of cell fate specification and during the time of axonogenesis in the embryonic CNS (Gertler, 1993). Murine Disabled is expressed in certain neuronal and hematopoietic cell lines and is localized to the growing nerves of embryonic mice. During mouse embryogenesis, murine Disabled is tyrosine phosphorylated when the nervous system is undergoing dramatic expansion, but when nerve tracts are established, murine Disabled lacks detectable phosphotyrosine. The properties of murine Disabled and genetic analysis of Disabled in Drosophila suggest that these molecules function in key signal transduction pathways involved in a function downstream of Abl in axonogenesis (Howell, 1997). The involvement of Abl in axonogenesis does not require Abl's kinase domain (Henkemeyer, 1990).
The importance of Abl in neurogenesis is also evidenced by Abl's interaction with another conserved protein, Enabled. Enabled is a member of the rapidly expanding VASP family of cytoskeletal associated proteins. The well studied migration of neuronal growth cones serves as a model for the actin-driven formation of membrane protrusions. Establishment of proper connections in the central nervous system depends on the ability of neuronal growth cones to guide neurites to their final targets. The ABL-ENA-Profilin pathway is implicated in the process of axonal outgrowth and fasciculation. Genetic screens for dominant second-site mutations that suppress the lethality of Abl mutations in Drosophila identify alleles of only one gene, enabled. The ENA protein contains proline-rich motifs and binds to ABL and Src SH3 domains. ENA is (as Disabled has been suggested to be) also a substrate for the Abl kinase. Tyrosine phosphorylation of ENA is increased when it is coexpressed in cells with human or Drosophila Abl, and endogenous ENA tyrosine phosphorylation is reduced in Abl mutant animals. Like Abl, ena is expressed at highest levels in the axons of the embryonic nervous system and ena mutant embryos show defects in axonal architecture. Therefore, it has been concluded that a critical function of Drosophila ABL is to phosphorylate and negatively regulate ENA protein during neural development (Gertler, 1995).
Drosophia Abl interacts with four other fly genes: prospero, failed axon connections, and Fasciclin 1 and Fasciclin 2, two genes coding for cell adhesion proteins. The interaction with prospero has not been explored further since the non-uniform cytoplasmic distribution of Prospero protein was discovered. The action of Inscuteable, a cytoskeleton associated protein, suggests an involvement of cytoskeleton in providing cues for Prospero and Numb subcellular localization. Cytochalasin D, which disrupts Actin filaments, eliminates Inscuteable crescents and results in incorrect Prospero crescent positioning (Kraut, 1996). Since the highest levels of Abl are not in neuroblasts but in postmitiotic neurons (Bennett, 1992), the relation between Prospero and Abl is currently unknown.
The interaction of Abl with Fas1, Fas2, and failed axon connections suggests the involvement of Abl in a feedback mechanism in which the cytoskeleton of the cell is given information about cell adherence events occurring during axonogenesis. This might be the Drosophila function of Abl with greatest relevance to vertebrate neurobiology. Abl also seems to be involved in signaling in early embryonic development both in Drosophila and Xenopus. The nature of this involvement has not yet been documented.
The proto-oncogenic kinase Abelson (Abl) regulates actin in response to cell signaling. Drosophila Abl is required in the nervous system, and also in epithelial cells, where it regulates adherens junction stability and actin organization. Abl acts at least in part via the actin regulator Enabled (Ena), but the mechanism by which Abl regulates Ena is unknown. A novel role is described for Abl in early Drosophila development, where it regulates the site and type of actin structures produced. In Abl's absence, excess actin is polymerized in apical microvilli, whereas too little actin is assembled into pseudocleavage and cellularization furrows. These effects involve Ena misregulation. In abl mutants, Ena accumulates ectopically at the apical cortex where excess actin is observed, suggesting that Abl regulates Ena's subcellular localization. Other actin regulators were also examined. Loss of Abl leads to changes in the localization of the Arp2/3 complex and the formin Diaphanous, and mutations in diaphanous or capping protein beta enhance abl phenotypes (Grevengoed, 2003).
Genetic analysis suggests that in the early Drosophila embryo, the primary means by which Abl influences the cytoskeleton is through Ena. Reducing the Ena dose by half profoundly suppresses ablM; it is as effective as adding back a wild-type abl transgene. Ena is also a critical target of Abl during embryonic morphogenesis. Although the data suggest that the primary effect of loss of Abl is Ena deregulation, they do not rule out Abl acting on the cytoskeleton by other mechanisms (Grevengoed, 2003).
The mechanism by which Abl regulates Ena has remained mysterious. This study demonstrates that Abl regulates Ena by regulating its intracellular localization. In the absence of Abl, Ena localizes to ectopic sites. Alterations in Ena and actin localization have been observed at the leading edge of migrating epidermal cells in abl mutants during dorsal closure. This suggests that regulation of Ena localization by Abl may be a more general mechanism. It is hypothesized that Abl targets Ena to places where it is needed to modulate actin dynamics, perhaps by excluding it from other sites where Ena activity would be detrimental (Grevengoed, 2003).
There are many ways in which Abl could restrict Ena localization. Abl's kinase activity is essential, and thus Abl phosphorylation of Ena may restrict its localization by preventing Ena binding to partners that localize to particular cortical sites, or by promoting Ena binding to partners that sequester it in the cytoplasm. Phosphorylation of Ena by Abl in vitro inhibits binding of Ena to SH3 domains, whereas Mena/VASP phosphorylation by PKA alters binding to SH3 domains and actin. However, if direct phosphorylation were the only mechanism by which Abl regulated Ena, mutating Ena's phosphorylation sites should create a protein that can no longer be regulated and thus would localize to ectopic sites. Instead, mutation of all of the Abl phosphorylation sites in Ena modestly reduced Ena function, rather than making it ectopically active as is seen in abl mutants (Grevengoed, 2003).
Thus, Abl may regulate Ena by additional mechanisms. Abl may modulate Ena localization and restrict Ena activity by direct binding (this could still require Abl kinase activity, since auto-phosphorylation or phosphorylation of other partners may regulate protein-protein interactions). Abl might sequester Ena in the cytoplasm in an inactive state, or it might recruit Ena to appropriate sites. Alternately, binding of Abl's SH3 domain to the Ena proline-rich region might prevent Ena from binding to other partners, such as profilin, which might in turn modulate both Ena localization and activity. In thinking about these different possible mechanisms, it is interesting to note that Abl localizes to the actin caps and apical pseudocleavage furrows during syncytial stages and the apical portion of the cellularization furrow, the precise places where ectopic actin accumulation occurs in its absence. Thus, it is poised to act at this location. Working out the details of the mechanism by which Abl regulates Ena localization will be one of the next challenges (Grevengoed, 2003).
This work provides an in vivo test of the current model for Ena function, and allows extension of this model. The excess growth of microvilli seen when Ena is ectopically localized in early embryos fits well with work on Ena/VASP function in mammalian fibroblasts, where forced localization of Ena/VASP proteins to the leading edge promotes the formation of long, unbranched filaments. Ena also localizes to the ends of filopodia and microspikes, suggesting that Ena's role in promoting long unbranched actin structures is broadly conserved. Earlier experiments in fibroblasts artificially altered Ena localization. This study demonstrates that Ena localization is a normal regulatory point in vivo, and that Abl is a critical player in this process. Finally, in vitro experiments have suggested that Ena promotes filament elongation by antagonizing capping protein. Mutations in cpb enhance the effects of mutations in abl in the CNS and probably during oogenesis. These data are consistent with Ena and capping protein playing antagonistic roles in vivo, with Abl potentially influencing the outcome of this antagonism. However, Abl and capping protein may also work together independently of Ena in the regulation of actin dynamics (Grevengoed, 2003).
Different actin regulators play fundamentally different biochemical roles. Models often picture all of these regulators modulating actin assembly and disassembly at a single site, but of course individual cells target different actin regulators to distinct sites, creating actin structures with diverse functions. Syncytial embryos provide an excellent example. During interphase, they assemble actin-based microvillar caps above each nucleus. As they enter prophase, caps are disassembled and actin polymerization is redirected to the pseudocleavage furrows. This is likely to require new machinery: both Arp2/3 and the formin Dia are required for pseudocleavage furrow formation, but not for actin caps. Cellularization also requires distinct machinery to polymerize/disassemble apical microvilli and to recruit and modulate actin at the cellularization front. For transitions to occur smoothly, two fundamental changes have to occur: the location at which actin polymerization occurs must change, and a different constellation of actin regulators must be deployed to produce the distinct actin structures observed (Grevengoed, 2003).
The data support a hypothesis in which the balance of activity of different actin regulators at distinct sites is tightly regulated, influencing the nature of the actin structures produced. One regulator is Abl. In its absence, Ena localizes ectopically to the cortical region, upsetting the temporal and spatial balance of actin regulators. This leads to a change in both the location and nature of actin polymerization during mitosis. Excess actin is polymerized into microvillar projections that extend from the apical region of the furrows, whereas insufficient actin is directed to the pseudocleavage furrows. Similarly, during cellularization in ablM mutants, actin polymerization continues to be directed to apical microvilli, whereas in a wild-type embryo this ceases early in cellularization (Grevengoed, 2003).
The data also suggest that there is cross-talk between different modulators of actin polymerization, and that the balance of their activities determines the outcome. Although many actin modulators are unaffected in ablM mutants, both the Arp2/3 complex and Dia are recruited to sites of ectopic actin polymerization. However, genetic analysis suggests that although Ena mislocalization plays a critical role in the actin alterations seen in ablM mutants, Dia and Arp2/3 mislocalization may not. In fact, reduction of the dose of Dia enhanced the ablM phenotype. Dia normally promotes actin polymerization lining the furrows. In ablM mutants, the balance of actin polymerization is already shifted to the apical microvilli because of ectopic Ena localization. Reduction in the dose of Dia might further reduce actin polymerization in pseudocleavage furrows, resulting in the observed enhancement of the ablM phenotype. The abnormal recruitment of Dia to the apical regions in ablM mutants may also reduce pseudocleavage furrow formation (Grevengoed, 2003).
It will now be important to investigate how the cell regulates the distinct types of actin polymerization required for distinct cellular and developmental processes. One mechanism of cross-talk may involve direct or indirect recruitment of one type of actin modulator by another. Abl's ability to interact with both Ena and the Arp2/3 regulator WAVE1 is interesting in this regard. However, the recruitment of Arp3 and Dia to ectopic actin structures observed in ablM mutants may have a more simple explanation. Both are thought to have a higher affinity for newly polymerized, ATP-bound actin, which is likely to be increased where ectopic actin polymerization appears to occur (Grevengoed, 2003).
Drosophila Abl also functions in other contexts. It has a role in embryonic morphogenesis, where it also acts, at least in part, via Ena. However, in this context Abl also affects AJ stability. Since Ena is normally highly enriched in AJs, it is hypothesized that Abl helps restrict Ena localization to AJs, and thus helps initiate the proper organization of actin underlying AJs. In Abl's absence, Ena may localize to sites other than AJs, leading to ectopic actin polymerization at those sites and reduction in actin polymerization at AJs (analogous to the divergent effects on apical actin and pseudocleavage/cellularization furrows). Since the cortical actin belt underlying the AJ plays an important role in its stability, this could explain the phenotype of abl mutants. A similar model may help explain the roles of Abl and Ena in axon outgrowth. The network of actin filaments in the growth cone is complex, with different types of actin in filopodia and in the body of the growth cone. By regulating Ena localization, Abl may influence the balance of the different types of actin, thus influencing growth cone motility. Likewise, in fibroblasts, where Ena/VASP proteins regulate motility, the Arp2/3 regulators N-WASP and WAVE localize to sites at the leading edge distinct from those where Mena is found. Whether Abl or Arg regulate the localization of Ena/VASP family proteins in mammals remains to be determined. Likewise, it is possible that deregulation of Ena/VASP proteins underlie some of the alterations in cell behavior in Bcr-Abl–transformed lymphocytes. Experiments to test whether Ena/VASP activity is important for either mammalian Abl's normal function or for the pathogenic effects of Bcr-Abl will help answer these questions (Grevengoed, 2003).
Exons - 10
Bases in 3' UTR - 1611
Abelson (Abl) gene consists of 10 exons extending over 26 kilobase pairs of genomic DNA. The DNA sequence encodes a protein of 1,520 amino acids with strong sequence similarity to the human c-abl proto-oncogene beginning in the type lb 5' exon and extending through the region essential for tyrosine kinase activity. When the tyrosine kinase homologous region is expressed in Escherichia coli, phosphorylation of proteins on tyrosine residues is observed. These results show that the abl gene is highly conserved through evolution and encodes a functional tyrosine protein kinase required for Drosophila development (Henkemeyer, 1988). There are five different 3' ends to ABL mRNA sequences, due to multiple polyadenylation sites separated from one another by as much as one kb (Telford, 1985).
The amino-terminal region of Abl tyrosine kinase related proteins shares several common features with Src-family members. These include a myristoylation site, and the arrangement of Src-homology domains in the primary sequence. Characteristic of the Abl-family members, however, is a large C-terminal extention beyond the kinase domain. The C-terminal segments are not well conserved among Abl-family members. For example, Drosophila Abl and the mammalian c-Abl are between 16-32% identical in this region. The divergent C-terminal segments are not interchangeable between Drosophila Abl and c-Abl, while the kinase domains are interchangeable. These results indicate that the C-terminal segments are important in specifying the biological functions of the Abl tyrosine kinases, and suggest that Drosophila Abl and c-Abl may perform different functions. Nevertheless, a C-terminal actin binding domain appears to be conserved. A presumed DNA binding domain and nuclear transport sequence is present in a region in c-Abl that is not conserved in Drosophila Abl (Wang, 1993 and references).
The Drosophila melanogaster abl and the murine v-abl genes encode tyrosine protein kinases (TPKs) whose amino acid sequences are highly conserved. To assess functional conservation between the two gene products, Drosophila abl/v-abl-chimeric Abelson murine leukemia viruses were constructed. In these chimeric Abelson murine leukemia viruses, the TPK and carboxy-terminal regions of v-Abl were replaced with the corresponding regions of Drosophila Abl. The chimeric Abelson murine leukemia viruses are able to mediate morphological and oncogenic transformation of NIH 3T3 cells and are able to abrogate the interleukin-3 dependence of a lymphoid cell line. A virus that contains both TPK and carboxy-terminal Drosophila Abl regions has no in vitro transforming activity for primary bone marrow cells and lacks the ability to induce tumors in susceptible mice. A virus that replaces only a portion of the v-Abl TPK region with that of Drosophila Abl has low activity in in vitro bone marrow transformation and tumorigenesis assays. These results indicate that the transforming functions of Abl TPKs are only partially conserved through evolution. These results also imply that the TPK region of v-Abl is a major determinant of its efficient lymphoid cell-transforming activity (Holland, 1990).
date revised: 15 October 2005
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