Worm Breeder's Gazette 11(4): 52
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
The Case of 27 missing nucleotides: As reported earlier (WBG: 11, No.2, 63-64, 1990), we have cloned and sequenced two alpha tubulin genes from C. elegans. Tubulin alpha-1, by direct screening of a C. elegans cDNA expression library, using a monoclonal antibody (3A5), and the tubulin alpha-2 from a genomic clone obtained from Linda Gremke and Joe Culotti. The tubulin genes were subcloned in pUC vectors, and both strands were sequenced using the dideoxy method. An anomaly was observed in the sequence of the alpha-2 tubulin gene; that a stretch of 27 nucleotides corresponding to nine amino acids (#200-288) was missing from the cloned DNA sequence in the lambda-vector. We speculated this could be due to an internal deletion in the sequencing vector. Accordingly, another genomic DNA library was screened for the corresponding clones, and the sequence determined. We have now the complete sequence of the alpha-2 tubulin gene, filling the missing nine amino acids. [See Figure 1] Trans-splicing of tubulin: Is it a rule or an exception? Monica Driscoll and others (JCB:109, 2993-3003) have described the - tubulin encoded by ben-1 locus in C. elegans, that has a .splice acceptor sequence (TTTCAG) at -39, in the 5' flanking sequence, with the possibility of a trans-splicing reaction of ben-1 mRNA. We have found that both alpha-1 and alpha-2 tubulin genes also contain the typical C. elegans splice acceptor consensus sequences . Both alpha tubulin genes contain two small introns of about 50 nucleotides, typical of C. elegans intron sequences. Salient features of the sequences are given below: [See Figure 2] Expression of alpha tubulins during development: We have also compared the expression of alpha-2 tubulin with the total alpha tubulin during different developmental stages, by screening cDNA libraries (kindly provided by J. Ahringer and J. Kimble) with gene specific probes, as shown in the histogram below. These results suggest alpha-2 tubulin gene is expressed at higher levels during embryonic development as compared to the larval growth. Northern analysis using these probes is in progress. [See Figure 3] Chromosomal location; & Do we have a mutant in alpha tubulins? The alpha-1 tubulin gene (TUBA3) has been located by John Sulston and Alan Coulson on linkage group I. However, so far alpha-2 tubulin gene's location has not been identified as it lies on one of the rare large contigs, that is still unconnected (We hope by the time this WBG issue is issued, its chromosomal location will be known). There are no mutations that are known to affect alpha tubulin genes in C. elegans. All suggestions from the worm community leading to their discovery are welcome. We want to thank Johji Miwa and his colleagues Y. Tabuse, K. Higashi, K. Nishiwaki, T. Sano and H. Sasa for their help with this project