Project description:The Retinoblastoma-like pocket proteins p130 and p107 act as gatekeepers of the cell cycle through their activity within the DREAM (Dp/Rb-like/E2F/MuvB) transcriptional repressor complex. The goal of this study was to address how the pocket protein contributes to DREAM complex assembly and function on chromatin by utilizing a protein null mutant of the only C. elegans pocket protein LIN-35. We performed ChIP-seq of C. elegans DRM subunits in wild-type and lin-35 null late embryos to assess the effect on their chromatin localization following loss of LIN-35.
Project description:The highly conserved DREAM transcriptional repressor complex contains an RB-like pocket protein, an E2F-DP transcription factor heterodimer, and the 5-subunit MuvB complex. Using CRISPR/Cas9 targeted mutagenesis, we disrupted the interaction between the sole Caenorhabditis elegans pocket protein LIN-35 and the MuvB subunit LIN-52. A triple alanine substitution of LIN-52's LxCxE motif (3A) severed LIN-35-MuvB association and caused classical DREAM mutant phenotypes, including synthetic multiple vulvae, high-temperature arrest, and ectopic expression of germline genes in the soma. We performed RNA-seq in lin-52(3A) mutant late embryos (4 replicates) compared to lin-52(WT) wild-type late embryos (4 replicates) to assess the genome-wide effects on gene expression that result from severing LIN-35-MuvB association.
