The Sotos syndrome gene Nsd1 safeguards bivalent developmental enhancers in poised states through restraining excessive H3K27me3 deposition [RNA-Seq]
Germline haploinsufficiency of Nsd1 is implicated as the etiology of Sotos syndrome, a developmental genetic disorder manifesting skeletal overgrowth and intellectual disability. However, the underlying molecular mechanisms remain largely elusive to date. Here we employed mouse embryonic stem (ES) cell in vitro differentiation as a developmental model system to tackle this question. We found that Nsd1 is indispensable for faithful differentiation of mouse ES cells into three primary germ layers, particularly the meso-endoderm lineages such as heart, vascular, and skeletal systems. Time-series gene expression profiling revealed that Nsd1 facilitates not only the basal expression but also differentiation-accompanied rapid induction of certain key meso-endoderm linage-specifying transcription factor genes such as T and Gata4. Mechanistically, Nsd1 directly binds to putative distal bivalent enhancers of these genes under ES state, where it antagonizes excessive H3K27me3 through depositing H3K36me2 and conversely maintains the active H3K27ac mark, thereby safeguarding these enhancers at poised or primed states that are highly responsive to developmental cues. In agreement, rescue assays showed that Nsd1-catalyzed H3K36me2 depends on intact PHD1-4, PWWP2, and SET domains to a similar degree, disruption of either one severely abolished Nsd1's ability to revert the defective differentiation phenotype. Additionally, Nsd1 is likely continuously required to sustain the activation of certain developmental genes, as it continues to reside at these genes after differentiation induction, albeit recruited by a different set of transcription factors. Altogether, our results provide novel molecular insights into how Nsd1 perturbations derail the normal developmental pathways and cause human disease. Overall design: RNA-seq profiling and comparison of wild-type vs Nsd1-knock out mouse embryonic stem cells collected at 3 different time points (day 0, day 2, day 4) during in vitro differentiation