Abstract
Medulloblastomas exhibit poor neuronal lineage specification. Expression of RE1 Silencing Transcription Factor (REST), a repressor of neurogenesis, is aberrantly elevated in human sonic hedgehog (SHH) medulloblastomas. Constitutive REST expression in mice (REST(TG)) drives medulloblastoma genesis and promotes tumor progression in the context of Ptch1 haploinsufficiency (Ptch(+/−)), implicating it as a driver of tumorigenesis. Tumor formation in Ptch(+/−)/REST(TG) mice showed significantly decreased latency and increased penetrance compared to that in Ptch(+/−) mice. Since REST silences gene expression by chromatin remodeling, we sought to identify cooperating epigenetic events that contributed to its oncogenic activity. To this end, we performed a loss of function screen employing a bar-coded library of short hairpin RNAs against epigenes, to identify candidates whose loss could create a proliferative vulnerability in the context of REST-elevation. This screen identified DNA methyltransferase 1 (DNMT1) as a high-priority epigenetic modifier. DNMT1 and the Ubiquitin like with PHD and Ring finger domain 1 (UHRF1) proteins are essential for methylation of hemi-methylated DNA at the replication fork during S-phase. Their expression is downregulated during neuronal differentiation. In human SHH-medulloblastoma tumors, REST and UHRF1 expression are positively correlated with higher levels of both genes noted specifically in the SHH-beta subtype, and is associated with poor prognosis. The requirement for DNMT1/UHRF1 in the context of REST elevation, was established by RNA-Seq and Reduced Representation Bisulfite Sequencing (RRBS), which revealed hypermethylation and downregulated expression of REST-target genes needed for neurogenesis. Thus, DNMT1/UHRF1 is a functional and potential therapeutic vulnerability in REST-elevated SHH medulloblastomas.