The C-terminal domain of Hsp70 is responsible for paralog-specific regulation of ribonucleotide reductase.

The Hsp70 family of molecular chaperones is well-conserved and expressed in all organisms. In budding yeast, cells express four highly similar cytosolic Hsp70s Ssa1, 2, 3 and 4 which arose from gene duplication. Ssa1 and 2 are constitutively expressed while Ssa3 and 4 are induced upon heat shock. Recent evidence suggests that despite their amino acid similarity, these Ssas have unique roles in the cell. Here we examine the relative importance of Ssa1-4 in the regulation of the enzyme ribonucleotide reductase (RNR). We demonstrate that cells expressing either Ssa3 or Ssa4 as their sole Ssa are compromised for their resistance to DNA damaging agents and activation of DNA damage response (DDR)-regulated transcription. In addition, we show that the steady state levels and stability of RNR small subunits Rnr2 and Rnr4 are reduced in Ssa3 or Ssa4-expressing cells, a result of decreased Ssa-RNR interaction. Interaction between the Hsp70 co-chaperone Ydj1 and RNR is correspondingly decreased in cells only expressing Ssa3 and 4. Through studies of Ssa2/4 domain swap chimeras, we determined that the C-terminal domain of Ssas are the source of this functional specificity. Taking together, our work suggests a distinct role for Ssa paralogs in regulating DNA replication mediated by C-terminus sequence variation. Author summary: Cells require molecular chaperones to fold proteins into their active conformation. A major mystery however is why cells express so many highly-related and apparently redundant Hsp70 paralogs. We examined the role of four Hsp70 paralogs in budding yeast (Ssa1, 2, 3 and 4) on the activity of the ribonucleotide reductase (RNR complex). Importantly, we demonstrate there is selectivity of RNR subunits for Ssa1 and Ssa2 subunits, which is dictated by the co-chaperone Ydj1. Taken together, our work provides new insight into the functional specificity of Hsp70 paralogs using a native client protein. [ABSTRACT FROM AUTHOR]