Figure 1: Suv39h1 enhances HP1α sumoylation in vivo.
From: The methyltransferase Suv39h1 links the SUMO pathway to HP1α marking at pericentric heterochromatin
(a) Schematic representation of Suv39h1 and Suv39h2. (b) SUMO-1-HP1α retrieved from NIH3T3 or Suv39h double-null cells. We used nuclear extracts prepared in presence of N-ethylmaleimide (NEM) for RNA pull-down using forward (F) major (Maj2 F) or minor (Min F) RNAs as baits. Western blot analysis using anti-HP1α antibodies revealed endogenous sumoylated HP1α (SUMO-1-HP1α) and unmodified HP1α. Input is 5% of nuclear extracts. The histogram shows the ratio between SUMO-1-HP1α (bound to RNA) and unmodified HP1α obtained, respectively, for NIH3T3 and Suv39h double-null cells. (c) Experimental scheme. (d) In vivo HP1α sumoylation assay. After anti-HA immunoprecipitation, western blot analysis using anti-GFP antibodies revealed sumoylated HP1α-HA (GFP-SUMO-1-HP1α-HA, box). Western blot using anti-Myc antibodies indicated that the four Myc-tagged proteins coimmunoprecipitated with HP1α-HA. The expression of all transfected proteins assessed in total cell extracts is shown by western blot using anti-GFP, anti-Myc and anti-HA antibodies. IgG corresponds to the immunoglobulin light chain. (e) As in (d) with the mutant versions Suv39h1-H324K and Suv39h1-ΔN40.
