Figure 8.

Loss of Id2 results in direct Hes1-mediated repression of Hes1 and Mash1 promoters. Chromatin immunoprecipitation (ChIP) was conducted to determine whether the loss of Id2 resulted in increased Hes1 function. A, PCR analysis of a known Hes1 auto-regulatory domain was conducted following preclearing and ChIP with Hes1 antisera in WT and Id2^−/− neurospheres. B, Further analysis of Hes1 immunoprecipitated DNA fragments using primers directed at a putative Hes1 binding site within the Mash1 promoter. In all ChIP experiments specificity was demonstrated in all trials by amplification of the housekeeping genes β-actin and cyclophilin in addition to target genes in both input and post-IP material. C, A novel model of Id2 function by which Id2 blocks the auto-repression of Hes1. Loss of Id2 results in both increased Hes1 auto-repressive function as well as inhibiting the expression of Mash1, a gene important in a prodopaminergic cell-fate program required during adult neurogenesis.