Broadening the functionality of a J-protein/Hsp70 molecular chaperone system SchilkeBrenda A. CiesielskiSzymon J. ZiegelhofferThomas KamiyaErina TonelliMarco LeeWoonghee CornilescuGabriel K. HinesJustin MarkleyJohn L. A. CraigElizabeth 2017 <div><p>By binding to a multitude of polypeptide substrates, Hsp70-based molecular chaperone systems perform a range of cellular functions. All J-protein co-chaperones play the essential role, via action of their J-domains, of stimulating the ATPase activity of Hsp70, thereby stabilizing its interaction with substrate. In addition, J-proteins drive the functional diversity of Hsp70 chaperone systems through action of regions outside their J-domains. Targeting to specific locations within a cellular compartment and binding of specific substrates for delivery to Hsp70 have been identified as modes of J-protein specialization. To better understand J-protein specialization, we concentrated on <i>Saccharomyces cerevisiae SIS1</i>, which encodes an essential J-protein of the cytosol/nucleus. We selected suppressors that allowed cells lacking <i>SIS1</i> to form colonies. Substitutions changing single residues in Ydj1, a J-protein, which, like Sis1, partners with Hsp70 Ssa1, were isolated. These gain-of-function substitutions were located at the end of the J-domain, suggesting that suppression was connected to interaction with its partner Hsp70, rather than substrate binding or subcellular localization. Reasoning that, if <i>YDJ1</i> suppressors affect Ssa1 function, substitutions in Hsp70 itself might also be able to overcome the cellular requirement for Sis1, we carried out a selection for <i>SSA1</i> suppressor mutations. Suppressing substitutions were isolated that altered sites in Ssa1 affecting the cycle of substrate interaction. Together, our results point to a third, additional means by which J-proteins can drive Hsp70’s ability to function in a wide range of cellular processes—modulating the Hsp70-substrate interaction cycle.</p></div>