Figure 2.
Lsd1 functions with CoREST as a H3K4 demethylase. (A) GAL4∷GFP variegation is suppressed by knocking down CoRest expression (left) or reducing CoRest gene dosage (right). For control, see Figure 1B. (B) Quantitation of variegation levels in A. N = 20 CoRestRNAi and 20 CoRestEY14216/+ ovarioles). (C) LSD1 and CoREST form a complex in adult ovaries. Immunoprecipitates (IP) were prepared from ovarian lysates using anti-LSD1 or anti-CoREST antibodies (Dallman et al. 2004) and then immunoblotted (IB) with the indicated antibodies. N = 3 experiments. (D) Point mutations in the Lsd1 enzymatic domain (i.e., lsd11 and lsd121; see the diagram above) suppress GAL4 variegation. N = 44 control, 35 lsd11/+, 22 lsd21/+, and 51 lsd1ΔN/+ ovarioles. (E) Schematic (shown above) of methylases (red) and demethylases (green and blue) that control histone H3K4 methylation. The effects of reducing the dosage of these genes on GAL4∷GFP variegation is shown. N = 51 lsd1ΔN/+, 29 ash1B1/+, 27 Trx1/+, 51 (LidK0681/+, and 33 Lid10424/+ ovarioles. (F) Trx and ash1 dominantly counteract lsd1-mediated suppression of GAL4∷ GFP variegation. N = 46 lsd1ΔN/+, Trx1/+ and 29 lsd1ΔN/+, ash1B1/+ ovarioles. (G,H) The su(var)3-9 gene encoding an H3K9 methylase does not affect GAL4∷GFP variegation (G) or suppress the effects of lsd1ΔN/+ (H). N = 20 su(var)3-91, 20 su(var)3-92, 34 su(var)3-91, lsd1ΔN/+, and 36 su(var)3-92, lsd1ΔN/+ ovarioles. Bars, 20 μm. Error bars indicate standard errors. (*) P < 0.05; (**) P < 0.01; (NS) nonsignificant difference, Student’s t-test.