Flying through the Drosophila Cytoskeletal Genome

The well-developed genetics of Drosophila make it an excellent system to understand mechanisms of cytoskeletal function and organization and how the cytoskeleton is coupled to development and differentiation. Many years of work have revealed that Drosophila has a set of cytoskeletal structural proteins and protein motors qualitatively similar to most other eukaryotes. The full toolbox of cytoskeletal elements, and the extent to which they resemble cytoskeletal proteins in other organisms has, however, remained unknown owing to the lack of a complete genome sequence. With the recent arrival of the complete sequence of the euchromatic genome of Drosophila (Adams et al., 2000; Rubin et al., 2000), we can begin to compile a complete list of cytoskeletal components. We can also answer once and for all the question of what cytoskeletal proteins discovered in mammals and other eukaryotes are present in the fly, and what variations on these known proteins exist. This article provides a brief review of the cytoskeletal genes of Drosophila , and highlights several interesting features of their nature and organization not previously known. As described (Adams et al., 2000) several different methods were used to predict the set of proteins encoded in the Drosophila genome. These methods relied on a combination of coupling the sequence to known genetic loci, EST sequences, and gene prediction programs. The predicted proteins were searched using greater than 1,000 cytoskeletal query sequences (see Table SI at http:// www.jcb.org/cgi/content/full/150/2/F63/DC1) from vertebrates, C. elegans , and S. cerevisiae . Queries were chosen by selecting representative sequences from each class of known cytoskeletal proteins described in Kreis and Vale (1999), as well as selections of proteins classified as cytoskeletal on the C. elegans and S. cerevisiae proteome Web sites. In addition, consensus sequence elements from the BLOCKS database were also used (Henikoff et al., 1999a,b). Searches were done primarily using BLASTP against the set of predicted proteins. In a few cases, TBLASTN was used to search the entire nucleotide sequence when an exhaustive classification of protein families was needed, or where a rigorous case for absence of a particular protein, or protein class was being established. G proteins and kinases were not analyzed since other workers were analyzing these proteins separately. Therefore, attention was effectively restricted to cytoskeletal structural proteins, non-enzymatic regulatory proteins, or proteins that moved, bound, cross-linked, or severed cytoskeletal polymers. Search results were inspected and evaluated manually; many false positive hits were found and eliminated from the classification as cytoskeletal. Overall, 262 genes (see Table SII at http://www.jcb.org/ cgi/content/full/150/2/F63/DC1) were found to give moderate to completely convincing homology with some member of the query set. This result corresponds to z 2% of the estimated 13,000 proteins encoded in the Drosophila genome. This 2% estimate is probably an underestimate since there are no doubt many proteins not yet recognized to have cytoskeletal functions, and it is also possible that some weakly homologous proteins were incorrectly eliminated from consideration. This 2% estimate is about half that reported for C. elegans (Chervitz et al., 1998) although the precise source of the estimate in this case is unclear. A few systematic sources of potential error were also identified in these analyses. In particular, proteins having coiled-coil, WD-40, or ankyrin repeat regions often gave significant BLAST scores with a variety of cytoskeletal queries, but examination of the actual sequence alignments revealed that they were not true homologues or relatives. For example, searches with the myosin II tail or intermediate filament proteins (see below) yield apparently significant hits with clearly unrelated a -helical coiled domains of myosins, kinesins, lamins, restin, and other unrelated a -helical coiled-coil containing proteins. Among the z 261 genes encoding cytoskeletal structural or motor proteins 95 genes encode polypeptides belonging to the kinesin, dynein, or myosin motor superfamilies, or accessory/regulatory polypeptides known to interact with the motor polypeptide subunits. Approximately 80 genes encode actin-binding proteins of various types, including proteins belonging to the spectrin/ a -actinin/dystrophin ✪ The online version of this article contains supplemental material. Address correspondence to Dr. Lawrence S.B. Goldstein, HHMI/ CMM Room 334, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0683. Tel.: (858)-534-9702. Fax: (858)-534-9701. E-mail: [email protected] on M ay 1, 2017 D ow nladed fom Published July 24, 2000