Polycomb Repression

The lincRNA HOTAIR is transcribed from the HOXC locus and targets polycomb repressive complex 2 (PRC2, which comprises H3K27 methylase EZH2, SUZ12, and EED) to silence HOXD and select genes on other chromosomes (7, 11). The genomic regions flanking HOXD are also bound by CoREST/REST repressor complexes (12), which contain LSD1 (KDM1/BHC110), a demethylase that mediates enzymatic demethylation of H3K4me2 (13) and that is required for proper repression of Hox genes in Drosophila (14). We therefore hypothesized that HOTAIR may coordinately interact with both PRC2 and LSD1. Immunoprecipitation (IP) of either endogenous LSD1 or FLAG-tagged LSD1 from primary foreskin fibroblasts or HeLa cells specifically retrieved endogenous HOTAIR RNA with enrichment comparable with that of EZH2 IP, the positive control (Fig. 1A and fig. S1A) (15). IP of three other chromatin proteins did not retrieve HOTAIR (fig. S1A), and neither LSD1, EZH2, nor FLAG-LSD1 IP retrieved U1 RNA, a nuclear ncRNA that served as a negative control. Purified biotinylated HOTAIR RNA, but not green fluorescent protein (GFP) RNA or an antisense HOTAIR fragment, specifically retrieved EZH2, SUZ12, and LSD1 from HeLa cell nuclear extract (Fig. 1B and fig. S1B). LSD1 forms a complex with CoREST (16), which can bridge LSD1 to the neuronal gene silencer REST (17). REST is believed to mediate silencing through two distinct effector arms: one via LSD1-CoREST, and separately via the adaptor protein CDYL and the H3K9 KMT G9a (18). HOTAIR specifically bound to CoREST and REST but not CDYL or G9a, nor to the putative PRC1 subunit YY1 (Fig. 1B). Further, biotinylated HOTAIR bound to purified PRC2 and LSD1 complexes in vitro (Fig. 1C and fig. S1C). These results suggest that HOTAIR directly interacts with PRC2 and LSD1 complexes.

Fig. 1

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Fig. 1

5′ domain of HOTAIR binds PRC2, and 3′ domain of HOTAIR...