ABSTRACT: Polycomb proteins are classical regulators of stem cell self-renewal and cell lineage commitment and are frequently deregulated in cancer. Here we find that the non-canonical PRC1.1 complex, as identified by mass spectrometry-based proteomics, is critically important for human leukemic stem cells. Downmodulation of PRC1.1 complex members, like the DNA-binding subunit KDM2B, strongly reduces cell proliferation in vitro and delays or even abrogates leukemogenesis in vivo in humanized xenograft models. PRC1.1 components are significantly overexpressed in primary AML CD34+ cells. Besides a set of genes that is co-targeted by PRC1 and PRC2, ChIP-seq studies show that PRC1.1 also binds a distinct set of genes that are devoid of H3K27me3 suggesting a gene regulatory role independent of PRC2. This set encompasses genes involved in metabolism, which have transcriptionally active chromatin profiles. These data indicate that PRC1.1 controls distinct gene clusters involved in unique cell biological processes required for leukemic cell viability. K562 cells were transduced with lentiviral GFP-fusion vectors encoding GFP-CBX2, PCGF1-GPF, PCGF2-GFP, PCGF4-GFP, GFP-RING1A or GFP-RING1B. K562 cells expressing GFP-fusions at relatively low levels were sorted and expanded and subsequently crosslinked. ChIP reactions were performed using the following antibodies: anti-GFP (ab290, Abcam), anti-H3K27me3 (07-449, Millipore), anti-H3K4me3 (ab8580, Abcam), anti-H2AK119ub (D27C4, Cell Signaling Technology), and anti-KDM2B (ab137547, Abcam). Furthermore, ChIP reactions were performed using primary AML CD34+ and Peripheral Blood (PB) CD34+ cells using anti-H3K27me3 (07-449, Millipore), anti-H3K4me3 (ab8580, Abcam), anti-H2AK119ub (D27C4, Cell Signaling Technology), anti-KDM2B (ab137547, Abcam).
