Interactions of HP1 with other chromosomal proteins

Abstract

To investigate the molecular basis of HP1a binding with a number of non‐histone chromosomal proteins in D. melanogaster, we are focusing on three examples: HP2, PIWI, and dAF10. Using co‐immunoprecipitation assays we demonstrate that an HP2 region containing an LCVKI sequence is responsible for binding to the chromo shadow domain (CSD) of HP1a. HP2 co‐precipitates with native HP1, but not with mutatants in HP1 that disrupt dimerization (I191E) or disrupt the PXVXL platform (W200A); mutation of the HP2 peptide at V2468E also abrogates binding. Using fluorescence polarization binding assays, we measure the observed dissociation constant of a partner peptide interacting with the HP1a‐CSD. We find that all three peptides bind with micromolar affinity and that the HP2 peptide binds the tightest followed by PIWI and the dAF10 peptides. Using 2D NMR spectroscopy we show that binding of any of the three peptides causes similar chemical shift perturbations in the portion of HP1a encompassing the CSD and CTE (C‐terminal extension) regions. Together our studies show that different short linear hydrophobic peptides can bind to the HP1a CSD‐CTE region, and potentially influence its localization in heterochromatin.Supported by NIH GM068388‐18 to SCRE and GM064786 to SK.