The Polycomb repressive complex 2 (PRC2) catalyzes H3K27 methylation and is required for maintaining transcriptional patterns and cellular identity, but the specification and maintenance of genomic PRC2 binding and H3K27 methylation patterns remain incompletely understood. Epigenetic mechanisms have been proposed, wherein pre-existing H3K27 methylation directs recruitment and regulates the catalytic activity of PRC2 to support its own maintenance. Here we investigate if such mechanisms are required for specifying H3K27 methylation patterns in mouse embryonic stem cells (mESCs). Through re-expression of PRC2 subunits in genetic knockouts that have lost all H3K27 methylation, we demonstrate that methylation patterns can be accurately established de novo. We find that regional methylation kinetics correlate with original methylation patterns even in their absence, and specification of the genomic PRC2 binding pattern is retained and specifically dependent on the PRC2 core-subunit SUZ12. Thus, the H3K27 methylation patterns in mESCs are not dependent on self-autonomous epigenetic inheritance.

Inhibition of Hedgehog (HH)/GLI signaling in cancer is a promising therapeutic approach. Interactions between HH/GLI and other oncogenic pathways affect the strength and tumorigenicity of HH/GLI. Cooperation of HH/GLI with Epidermal Growth Factor Receptor (EGFR) signaling promotes transformation and cancer cell proliferation in vitro. However, the in vivo relevance of HH-EGFR signal integration and the critical downstream mediators are largely undefined. In this report we show that genetic and pharmacologic inhibition of EGFR signaling reduces tumor growth in mouse models of HH/GLI driven basal cell carcinoma (BCC). We describe HH-EGFR cooperation response genes including SOX2, SOX9, JUN, CXCR4 and FGF19 that are synergistically activated by HH-EGFR signal integration and required for in vivo growth of BCC cells and tumor-initiating pancreatic cancer cells. The data validate EGFR signaling as drug target in HH/GLI driven cancers and shed light on the molecular processes controlled by HH-EGFR signal cooperation, providing new therapeutic strategies based on combined targeting of HH-EGFR signaling and selected downstream target genes.

Regulation of the Hedgehog (Hh) pathway relies on an interaction of two receptors that is not fully understood. Patched1 (Ptch1) binds the Hh ligand, but is also a negative regulator of pathway activity. Binding of Hh ligand to Ptch1 leads to the relocation of the activating receptor Smoothened (Smo) to the primary cilium, which is required for the transcriptional Hh response. Besides the transcriptional response, Hh can also induce chemotaxis, and we assessed the effects of defective ciliary localization of Smo on its subcellular itineraries and chemotactic signaling capacity. We find that defective ciliary localization of Smo results in markedly different intracellular trafficking of Smo to sites not involving the primary cilium. These itineraries correlate with a decreased transcriptional signaling capacity, and an enhanced chemotactic responsiveness. These data imply that the ciliary localization machinery functions to transport Smo to sites where it can mediate transcriptional signaling, and away from locations where it can mediate chemotactic signaling. The subcellular localization of Smo is thus a crucial determinant of its signaling characteristics and implies the existence of pool of Smo dedicated to chemotaxis.