, 2008 and Gu et al., click here 2009; see Supplemental Experimental Procedures).

Mouse brains were perfused, sectioned, and immunostained by using established protocols (Gray et al., 2008 and Gu et al., 2009; see Supplemental Experimental Procedures). HDL2 brain samples used in the study were described in detail before (Rudnicki et al., 2008). The following antibodies were used to stain NIs in HDL2 models: 3B5H10 (1:1000; Sigma, St. Louis, MO), 1C2 (1:3000; Chemicon, Billerica, MA), CBP (1:3000; A-22 & sc-583, Santa Cruz, Santa Cruz, CA), ubiquitin (1:1000; DakoCytomation, Carpinteria, CA), 3B5H10 (1:2000), MBNL1 antibody (A2764, 1:10000 dilution; Lin et al., 2006). Antigen retrieval for polyQ NI detection by using 3B5H10 was performed according to published protocols (Osmand et al., 2006). More details selleck kinase inhibitor on immunohistochemical methods and reagents and quantitation of NI sizes can be found in Supplemental Experimental

Procedures. Brain extracts or nuclear and cytoplasmic fractionations were performed by using established methods (Gray et al., 2008 and Gu et al., 2009; see Supplemental Experimental Procedures). Antibodies for western blot included: JPH3 exon 4 (1:1000; H. Takeshima, Tohoku University, Japan), M2-Flag (1:500), α-tubulin (1:2000), 3B5H10 (1:1000; Sigma, St. Louis, MO), and 1C2 (1:2000; Chemicon, Billerica, MA). ChIP analyses were performed by using our established method (Martinowich et al., 2003; see Supplemental Experimental Procedures) with the following antibodies: anti-CBP (sc-583, Santa Cruz) and anti-IgG (sc-66931, Santa Cruz). Real-time quantitative PCR was performed by using iQ SYBR Green Supermix (Bio-Rad). For quantification of relative level of CBP occupancy, we calculated the percentage of the immunoprecipitated DNA over whole-cell extract. Primer sequences used in ChIP-qPCR are listed in Supplemental Experimental Procedures. See details in Supplemental Experimental Procedures. All data are shown as the mean ± SEM. SPSS 14.0 statistics software (SPSS, Chicago, IL) was used

to perform all statistical analyses. The significance level was set at 0.05. See more details in Supplemental Experimental Procedures and in our published methods (Gray et al., 2008 and Gu et al., 2009). This work was generously supported by independent research grants from the Hereditary Disease Foundation to X.W.Y., R.L.M., A.O., and to D.D.R. X.W.Y. is also supported by National Institutes of Health (NIH)/National Institute of Neurological Disorders and Stroke (NINDS; R01NS049501), the David Weil Fund to the Semel Institute at University of California, Los Angeles, and Neuroscience of Brain Disorders Award from The McKnight Endowment Fund for Neuroscience. R.L.M. and D.D.R. are supported by NIH/NINDS (R21NS057516 and R01NS064138). We would like to thank N.S. Wexler and C. Johnson for their tremendous support of this project.