MOLECULAR ANALYSIS OF CAENORHABDITIS-ELEGANS TCP-1, A GENE ENCODING ACHAPERONIN PROTEIN

A Caenorhabditis elegans (Ce) homologue to the eukaryotic tcp-1 gene ( encoding t-complex polypeptide-1) has been mapped, isolated and sequen ced. Ce tcp-1 is a single-copy gene located on chromosome II. Nucleoti de sequence analysis of the gene reveals the presence of four introns in the coding region and repetitive elements upstream from the start c odon. The predicted Ce TCP-1 protein displays more than 60% amino-acid sequence identity to other eukaryotic TCP-1, suggesting a common orig in and function for these proteins. The primary tcp-1 transcript under goes transsplicing to the spliced leader SL1 RNA, in addition to cis-s plicing, to yield a single mRNA species of 1.9 kb. Northern blot analy sis shows that unlike the evolutionarily related Hsp60 chaperonin gene s, tcp-1 is not upregulated at elevated temperatures, but instead appe ars to be down-regulated. Additionally, the overall level of the tcp-1 transcript is approximately constant throughout the development of th e nematode. The Ce chaperonin-containing TCP-1 (CCT) was identified. A protein extract made from Ce embryos was subjected to sucrose gradien t fractionation and ATP-agarose chromatography. Western blot analysis of the purified protein fractions, using anti-mouse TCP-1 monoclonal a ntibody and antibodies raised against Ce TCP-1, reveals that Ce TCP-1 is a 57-kDa protein subunit of a high-molecular-mass complex capable o f binding ATP.