Interaction analysis of the AAA+ domain of the heterochromatin protein Sir3

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The silent information regulator 2/3/4 (SIR) complex is an archetypal heterochromatin complex re-quired for gene silencing at the silent mating-type loci and the telomeres in the yeast Saccharomyces cerevisiae. The Sir3 component consists of an N-terminal bromo-adjacent homology (BAH) domain and a C-terminal AAA+ ATPase-like domain. In a recent study, we have determined the crystal struc-ture of the AAA+ domain and, through an exhaustive mutational analysis, have identified several sur-face regions on this domain required for Sir3 silencing function, one of which we have shown repre-sents a site of interaction with Sir4. Based on these results, we here will determine the interaction partners of three other surface domains using a combination of genetics, molecular biology and bio-chemistry. Furthermore, we will define the region on Sir4 that interacts with Sir3. This detailed molecu-lar information about protein interactions within the SIR complex and with chromatin will be comple-mentary to structural information on the SIR complex and will for the first time provide molecular in-sight into a heterochromatin complex.