The regulatory function of dIno80 correlates with its DNA binding activity

Abstract

Running title: DNA binding activity of dIno80 Abbreviations: dIno80, Drosophila Ino80; hIno80; human Ino80; Ino80, Inositol requiring mutant 80; ETP, Enhancer of Trithorax and Polycomb; FID, fluorescence intercalator displacement; DBINO, DNA binding domain of Ino80; EMSA, electrophoretic mobility shift assay. Highlights  dIno80 has a DNA binding domain through which it recognizes a consensus sequence and interacts with DNA.  dIno80 interacts with variants of the consensus sequence with variable affinity.  dIno80 can regulate the expression of downstream as well as upstream reporter gene.  dIno80 can complement the function of Pho and rescue Pho lethality. 2 ABSTRACTThe INO80 complex, including the Ino80 protein, forms a highly conserved canonical complex that remodels chromatin in the context of multiple cellular functions. The Drosophila homologue, dIno80, is involved in homeotic gene regulation during development as a canonical Pho-dIno80 complex. Previously, we found that dIno80 regulates homeotic genes by interacting with epigenetic regulators, such as polycomb and trithorax, suggesting the occurrence of non-canonical Ino80 complexes. Here using spectroscopic methods and gel retardation assays, we identified a set of consensus DNA sequences that DNA binding domain of dIno80 (DBINO)interacts with havingdifferential affinity and high specificity. Testing these sequences in reporters showed that this interaction can positively regulate transcription. We also demonstrated that over-expressing dIno80 can rescue the pupal lethality of pleiohomeotic (pho) deficient flies. These results suggest that, in addition to its known function as a chromatin remodeler, dIno80 has sequence preference for interaction with DNA to regulate transcription.