Class-specific interactions between Sis1 J-domain protein and Hsp70 chaperone potentiate disaggregation of misfolded proteins

Wyszkowski, Hubert; Janta, Anna; Sztangierska, Wiktoria; Obuchowski, Igor; Chamera, Tomasz; Kłosowska, Agnieszka; Liberek, Krzysztof

doi:10.1073/pnas.2108163118

“…In contrast to ribonucleotide reductase whose stability depends on Ydj1 function, loss of Ydj1 does not impact Apn2 stability [ 28 ]. It is possible that Apn2 stability in yeast is additionally regulated by other semi-redundant co-chaperones such as Sis1, which has similar yet distinct roles in the cell as Ydj1 [ 39 , 40 , 41 , 42 ]. This may also explain why in our studies, DNAJA1 the mammalian homologue of Ydj1 does not appear to interact with APE2 ( Figure 3 A).…”

Section: Discussionmentioning

confidence: 99%

The APE2 Exonuclease Is a Client of the Hsp70–Hsp90 Axis in Yeast and Mammalian Cells

Omkar

¹

,

Wani

²

,

Zheng

³

et al. 2022

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Molecular chaperones such as Hsp70 and Hsp90 help fold and activate proteins in important signal transduction pathways that include DNA damage response (DDR). Previous studies have suggested that the levels of the mammalian APE2 exonuclease, a protein critical for DNA repair, may be dependent on chaperone activity. In this study, we demonstrate that the budding yeast Apn2 exonuclease interacts with molecular chaperones Ssa1 and Hsp82 and the co-chaperone Ydj1. Although Apn2 does not display a binding preference for any specific cytosolic Hsp70 or Hsp90 paralog, Ssa1 is unable to support Apn2 stability when present as the sole Ssa in the cell. Demonstrating conservation of this mechanism, the exonuclease APE2 also binds to Hsp70 and Hsp90 in mammalian cells. Inhibition of chaperone function via specific small molecule inhibitors results in a rapid loss of APE2 in a range of cancer cell lines. Taken together, these data identify APE2 and Apn2 as clients of the chaperone system in yeast and mammalian cells and suggest that chaperone inhibition may form the basis of novel anticancer therapies that target APE2-mediated processes.

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“…Previous studies have identified this region as being important for the binding of co-chaperone proteins. The VEEVD sequence at the end of Hsp70/Ssa1 is critical interaction with DNAJB-type co-chaperones including Sis1 [49–52]. However, there is also substantial evidence that the CTD is also important for the binding of Ydj1.…”

Section: Discussionmentioning

confidence: 99%

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

Knighton

¹

,

Nitika

²

,

Omkar

³

et al. 2022

Preprint

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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 DDR 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 SummaryCells 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.

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“…Previous studies have identified this region as being important for the binding of co-chaperone proteins. The VEEVD sequence at the end of Hsp70/Ssa1 is critical for interaction with DNAJB-type co-chaperones including Sis1 [ 55 – 58 ]. However, there is also substantial evidence that the CTD is also important for the binding of Ydj1.…”

Section: Discussionmentioning

confidence: 99%

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

Knighton

¹

,

Nitika

²

,

Omkar

³

et al. 2022

View full text Add to dashboard Cite

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.

show abstract

Class-specific interactions between Sis1 J-domain protein and Hsp70 chaperone potentiate disaggregation of misfolded proteins

Cited by 26 publications

References 62 publications

The APE2 Exonuclease Is a Client of the Hsp70–Hsp90 Axis in Yeast and Mammalian Cells

The APE2 Exonuclease Is a Client of the Hsp70–Hsp90 Axis in Yeast and Mammalian Cells

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

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

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