[
Worm Breeder's Gazette,
1995]
ELAV is a Drosophila protein that has been linked genetically to the differentiation of neuroblasts to neurons and is also responsible for the maintenance of terminally differentiated neurons. ELAV like proteins contain three RNA Recognition Motifs (RRMs), the first two RRMs are usually found at the N terminus of the protein separated by one or two amino acids. The third RRM is separated from the first two by a variable hinge region that appears to be alternatively spliced with the presence of miniexons that may influence the function of the protein. Several labs have successfully cloned homologs of the ELAV protein from other organisms such as rat, humans, and Xenopus. The human counterparts of ELAV have been shown to bind to AU rich stretches in the 3' untranslated regions of certain mRNAs, such as c-myc, c-fos, and gm-csf. Analysis of the Xenopus clones have shown that three of the ELAV homologs are expressed sequentially during the development of the frog embryo. While screening libraries for a C. elegans counterpart of ELAV, we found a cDNA that encoded a putative ORF encoding the first ELAV homolog. This clone corresponded to a predicted polypeptide on chromosome II sequenced by the genome sequencing project. Our screening has produced a full length clone corresponding to the predicted nucleotide sequence of the putative ORF. A western was performed using anti Human ELAV-Like Neuronal protein 1 (HEL-N1) rabbit polyclonal antisera against a protein extract of a mixed population of N2 worms and revealed a band, of the predicted size of the CEL-1 protein. Among other things we intend to determine the developmental regulation of this protein, attempt to rescue preexisting mutants, generate mutants if none now exist, determine the localization of the protein, elucidate the in vivo mRNA targets of the protein, and use the genetics of C. elegans to uncover the mechanism of action of this family of proteins.
[
Worm Breeder's Gazette,
1996]
We have recently cloned a C. elegans gene, we call cevh, that expresses an RNA-binding protein of unknown function. So far we have a full-length cDNA clone and also a cosmid that contains the complete gene. The cevh gene physically maps near the end of the left arm of chromosome I, however we have been unable to identify any existing mutants thus far. The CEVH protein is about 65 kD and is a member of the RRM (RNA Recognition Motif) protein superfamily. The protein contains 3 RRMs, two adjacent to one another in the middle of the protein followed by a 19 amino acid linker and then a third RRM. Previous work in our laboratory indicates that each RRM has the potential to bind to a specific RNA sequence, so CEVH may bind several RNAs or may bind the same RNA in several places. Preliminary in vitro selection experiments suggest the protein may prefer RNA that contains a UGGGC/U sequence. We have overexpressed and purified the CEVH protein in order to produce antibodies for immunoprecipitation of natural ligands. The CEVH antibody will also be used to look for tissue specific and developmental expression of the protein by immunofluorescence in worms. In an effort to uncover the function of this protein we will continue to characterize the gene and the protein it expresses.