DSIR2 and dHDAC6: Two novel, inhibitor-resistant deacetylases in Drosophila melanogaster

We have identified new members of the histone deacetylase enzyme family in Drosophila melanogaster. dHDAC6 is a class II deacetylase with two active s ites, and dSIR2 is an NAD-dependent histone deacetylase, These proteins, to gether with two class I histone deacetylases, dHDAC1 and dHDAC3, have been expressed and characterized as epitope-tagged recombinant proteins in Schne ider SL2 cells. All these proteins have in vitro deacetylase activity and a re able to deacetylate core histone H4 at all four acetylatable lysine resi dues (5, 8, 12, and 16), Recombinant dHDAC6 and dSIR2 are both insensitive to TSA and HC toxin and resistant, relative to dHDAC1 and dHDAC3, to inhibi tion by sodium butyrate, Indirect immunofluorescence microscopy of stably t ransfected SL2 lines reveals that dHDAC1 and dSIR2 are nuclear, dHDAC6 is c ytosolic, and dHDAC3 is detectable in both cytosol and nucleus. dHDAC6 and dSIR2 elute from Superose 6 columns with apparent molecular weights of 90 a nd 200 kDa, respectively. In contrast, dHDAC1 and dHDAC3 elute at 800 and 7 00 kDa, respectively, suggesting that they are components of multiprotein c omplexes. Consistent with this, recombinant dHDAC1 coimmunoprecipitates wit h components of the Drosophila NuRD complex and dHDAC3 with an as yet unkno wn 45-kDa protein. (C) zool Academic Press.