GENETICS OF NEGATIVE CELL CYCLE REGULATORS IN C ELEGANS

我们的长期目标是了解细胞的发育控制

Our long term goal is to understand the developmental controls on cell cycle progression. We are taking a genetic approach by studying two mutants in C. elegans, cul-1 and lin-23, that have hyperplasia in multiple tissues. In these mutants, dividing cells ignore developmental cues and continue to proliferate. The extra cells generated are smaller than normal. We have cloned both the cul-1 and lin-23 genes. cul-1 and lin-23 are homologous to the S. cerevisiae genes CDC53 and CDC4, respectively, which are required for the G1 to S phase cell cycle transition. CDC53 and CDC4 are required for the ubiquitin-mediated degradation of cell cycle regulators, including cyclins and cyclin- dependent kinase inhibitors. An inability to degrade G1 cyclins in yeast produces a shorter G1 phase, small cells, and continued cell proliferation in the presence of inhibitory external signals. These phenotypes are similar to what is observed in cul-1 and lin-23 mutants. A compelling model is that cul-1 and lin-23 are required for the degradation of key cell cycle regulators in G1 phase, in particular G1 cyclins. The goals of the proposed research are to test this hypothesis and determine how cul-1 and lin-23 function to negatively regulate cell proliferation. In the first of three aims, we will determine the expression pattern of cul-1 and lin-23. We will test our hypothesis that cul-1 and lin-23 affect the protein levels of various cell cycle regulators by determining their expression and perdurance in wild type and cul-1 or lin-23 mutant strains. Further, we will determine whether cul-1 and lin-23 mutant strains have a shorter G1 phase. In the second aim, we will identify interacting proteins through the use of a yeast two-hybrid screen. Putative interacting clones will be analyzed by biochemical, cellular, and genetic approaches to determine their in vivo relevance. In the third aim, we will isolate and molecularly characterize extragenic suppressors of cul-1 and lin-23.