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　Caenorhabditis elegans possesses two p97/VCP/Cdc48p homologues, named CDC-48.1 (C06A1.1) and CDC-48.2 (C41C4.8), and their expression patterns and levels are differently regulated. To clarify the regulatory mechanisms of differential expression of two p97 proteins of C. elegans , we performed detailed deletion analysis of their promoter regions. We found that the promoter of cdc-48.1 contains two regions necessary for embryonic and for post-embryonic expression, while the promoter of cdc-48.2 contains the single region necessary for embryonic expression. In particular, two elements (Element A and Element B) and three conserved boxes (Box a, Box b and Box c) were essential for cdc-48.1 expression in embryos and at post-embryonic stages, respectively. By using South-Western blotting and MALDI-TOF MS analysis, we identified HMG-12 and CAR-1 as proteins that bind to Element A and Element B, respectively, from the embryonic nuclear extract. Importantly, we found the decreased expression of p97 in embryos prepared from hmg-12(RNAi) or car-1(RNAi) worms. These results indicate that both HMG-12 and CAR-1 play important roles in embryonic expression of cdc-48.1 .

 

Figure 1: Promoter structures of two p97 genes of C. elegans . The cdc-48.1 promoter contains two modules: one is essential for embryonic expression [Element A (white box) and Element B (black box)] and the other for post-embryonic expression [Box a, Box b and Box c (gray boxes)]. HMG-12 and CAR-1 would bind to Element A and Element
B, respectively, and play a crucial role for embryonic expression of cdc-48.1 . The cdc-48.2promoter contains the region only involved in embryonic expression (hatched box).

Figure 2: Effects of RNAi of hmg-12 and car-1 on embryonic expression of cdc-48.1 were analyzed with GFP constructs (left) and with Western blotting (right). The level of p97 proteins in embryos was significantly reduced both hmg-12 (RNAi) and car-1 (RNAi) worms. In the case of car-1 (RNAi) , RNAi effects were more prominent.