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Hsp90-Associated Immunophilin Homolog Cpr7 Is Required for the Mitotic Stability of [URE3] Prion in Saccharomyces cerevisiae.

Kumar N, Gaur D, Gupta A, Puri A, Sharma D - PLoS Genet. (2015)

Bottom Line: We show that Cpr7 interacts with Ure2 and enhances its fibrillation.The requirement of Cpr7 is specific for [URE3] as its deletion does not antagonize both strong and weak variant of another yeast prion [PSI+], suggesting a distinct role of the Hsp90 co-chaperone with different yeast prions.Our data show that, similar to the Hsp70 family, the Hsp90 chaperones also influence yeast prion maintenance, and that immunophilins could regulate protein multimerization independently of their activity as peptidyl-prolyl isomerases.

View Article: PubMed Central - PubMed

Affiliation: Council of Scientific and Industrial Research-Institute of Microbial Technology, Chandigarh, India.

ABSTRACT
The role of Hsp70 chaperones in yeast prion propagation is well established. Highly conserved Hsp90 chaperones participate in a number of cellular processes, such as client protein maturation, protein degradation, cellular signalling and apoptosis, but little is known about their role in propagation of infectious prion like aggregates. Here, we examine the influence of Hsp90 in the maintenance of yeast prion [URE3] which is a prion form of native protein Ure2, and reveal a previously unknown role of Hsp90 as an important regulator of [URE3] stability. We show that the C-terminal MEEVD pentapeptide motif, but not the client maturation activity of Hsp90, is essential for [URE3] prion stability. In testing deletions of various Hsp90 co-chaperones known to bind this motif, we find the immunophilin homolog Cpr7 is essential for [URE3] propagation. We show that Cpr7 interacts with Ure2 and enhances its fibrillation. The requirement of Cpr7 is specific for [URE3] as its deletion does not antagonize both strong and weak variant of another yeast prion [PSI+], suggesting a distinct role of the Hsp90 co-chaperone with different yeast prions. Our data show that, similar to the Hsp70 family, the Hsp90 chaperones also influence yeast prion maintenance, and that immunophilins could regulate protein multimerization independently of their activity as peptidyl-prolyl isomerases.

No MeSH data available.


Related in: MedlinePlus

Deleting the MEEVD motif of Hsp90 destabilizes [URE3].(A) Effect of deletion and overexpression of Hsp90 on [URE3]. Hsp82Δ (SY295) or Hsc82Δ (SY297) cells grown overnight were subcultured into YPAD medium and grown from O.D.600nm of 0.02 to 1.7. Cells were then plated onto ½ YPD plates and monitored after 3 days of incubation at 30°C. For overexpression studies, SY187 was transformed with plasmid (pRS426PGPD-His6-Hsp82 or pRS426PGPD-His6-Hsc82) encoding His6-Hsp82 or His6-Hsc82 and plated onto uracil deficient solid SD medium with limiting adenine. As seen by white colony color phenotype, no adverse effect on [URE3] stability was observed upon deletion or overexpression of Hsp90 isoforms. (B) The strains constructed as described in Materials and Methods express the indicated Hsp90 isoform as the sole cytosolic Hsp90. Cells were grown from O.D.600nm of 0.02 to 1.7 before plating onto ½ YPD medium. As seen by red colony color phenotype the frequency of [ure-o] colonies increases upon deletion of MEEVD motif.
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pgen.1005567.g001: Deleting the MEEVD motif of Hsp90 destabilizes [URE3].(A) Effect of deletion and overexpression of Hsp90 on [URE3]. Hsp82Δ (SY295) or Hsc82Δ (SY297) cells grown overnight were subcultured into YPAD medium and grown from O.D.600nm of 0.02 to 1.7. Cells were then plated onto ½ YPD plates and monitored after 3 days of incubation at 30°C. For overexpression studies, SY187 was transformed with plasmid (pRS426PGPD-His6-Hsp82 or pRS426PGPD-His6-Hsc82) encoding His6-Hsp82 or His6-Hsc82 and plated onto uracil deficient solid SD medium with limiting adenine. As seen by white colony color phenotype, no adverse effect on [URE3] stability was observed upon deletion or overexpression of Hsp90 isoforms. (B) The strains constructed as described in Materials and Methods express the indicated Hsp90 isoform as the sole cytosolic Hsp90. Cells were grown from O.D.600nm of 0.02 to 1.7 before plating onto ½ YPD medium. As seen by red colony color phenotype the frequency of [ure-o] colonies increases upon deletion of MEEVD motif.

Mentions: It is known that Hsp70 and Hsp90 family proteins influence folding and maturation of many cellular proteins, yet what determines chaperone specificity is not entirely clear. Whether yeast prions that are well known Hsp70 substrates also require Hsp90 remains unclear. In order to explore the role of Hsp90 chaperones on yeast prions, S. cerevisiae strains having deletion of either HSC82 or HSP82 were constructed. As seen in Fig 1A, no significant effect on [URE3] was observed suggesting that the presence of either Hsp90 is enough to support stable prion propagation. Alternatively, the Hsp90 family of proteins may be dispensable for [URE3] propagation.


Hsp90-Associated Immunophilin Homolog Cpr7 Is Required for the Mitotic Stability of [URE3] Prion in Saccharomyces cerevisiae.

Kumar N, Gaur D, Gupta A, Puri A, Sharma D - PLoS Genet. (2015)

Deleting the MEEVD motif of Hsp90 destabilizes [URE3].(A) Effect of deletion and overexpression of Hsp90 on [URE3]. Hsp82Δ (SY295) or Hsc82Δ (SY297) cells grown overnight were subcultured into YPAD medium and grown from O.D.600nm of 0.02 to 1.7. Cells were then plated onto ½ YPD plates and monitored after 3 days of incubation at 30°C. For overexpression studies, SY187 was transformed with plasmid (pRS426PGPD-His6-Hsp82 or pRS426PGPD-His6-Hsc82) encoding His6-Hsp82 or His6-Hsc82 and plated onto uracil deficient solid SD medium with limiting adenine. As seen by white colony color phenotype, no adverse effect on [URE3] stability was observed upon deletion or overexpression of Hsp90 isoforms. (B) The strains constructed as described in Materials and Methods express the indicated Hsp90 isoform as the sole cytosolic Hsp90. Cells were grown from O.D.600nm of 0.02 to 1.7 before plating onto ½ YPD medium. As seen by red colony color phenotype the frequency of [ure-o] colonies increases upon deletion of MEEVD motif.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4608684&req=5

pgen.1005567.g001: Deleting the MEEVD motif of Hsp90 destabilizes [URE3].(A) Effect of deletion and overexpression of Hsp90 on [URE3]. Hsp82Δ (SY295) or Hsc82Δ (SY297) cells grown overnight were subcultured into YPAD medium and grown from O.D.600nm of 0.02 to 1.7. Cells were then plated onto ½ YPD plates and monitored after 3 days of incubation at 30°C. For overexpression studies, SY187 was transformed with plasmid (pRS426PGPD-His6-Hsp82 or pRS426PGPD-His6-Hsc82) encoding His6-Hsp82 or His6-Hsc82 and plated onto uracil deficient solid SD medium with limiting adenine. As seen by white colony color phenotype, no adverse effect on [URE3] stability was observed upon deletion or overexpression of Hsp90 isoforms. (B) The strains constructed as described in Materials and Methods express the indicated Hsp90 isoform as the sole cytosolic Hsp90. Cells were grown from O.D.600nm of 0.02 to 1.7 before plating onto ½ YPD medium. As seen by red colony color phenotype the frequency of [ure-o] colonies increases upon deletion of MEEVD motif.
Mentions: It is known that Hsp70 and Hsp90 family proteins influence folding and maturation of many cellular proteins, yet what determines chaperone specificity is not entirely clear. Whether yeast prions that are well known Hsp70 substrates also require Hsp90 remains unclear. In order to explore the role of Hsp90 chaperones on yeast prions, S. cerevisiae strains having deletion of either HSC82 or HSP82 were constructed. As seen in Fig 1A, no significant effect on [URE3] was observed suggesting that the presence of either Hsp90 is enough to support stable prion propagation. Alternatively, the Hsp90 family of proteins may be dispensable for [URE3] propagation.

Bottom Line: We show that Cpr7 interacts with Ure2 and enhances its fibrillation.The requirement of Cpr7 is specific for [URE3] as its deletion does not antagonize both strong and weak variant of another yeast prion [PSI+], suggesting a distinct role of the Hsp90 co-chaperone with different yeast prions.Our data show that, similar to the Hsp70 family, the Hsp90 chaperones also influence yeast prion maintenance, and that immunophilins could regulate protein multimerization independently of their activity as peptidyl-prolyl isomerases.

View Article: PubMed Central - PubMed

Affiliation: Council of Scientific and Industrial Research-Institute of Microbial Technology, Chandigarh, India.

ABSTRACT
The role of Hsp70 chaperones in yeast prion propagation is well established. Highly conserved Hsp90 chaperones participate in a number of cellular processes, such as client protein maturation, protein degradation, cellular signalling and apoptosis, but little is known about their role in propagation of infectious prion like aggregates. Here, we examine the influence of Hsp90 in the maintenance of yeast prion [URE3] which is a prion form of native protein Ure2, and reveal a previously unknown role of Hsp90 as an important regulator of [URE3] stability. We show that the C-terminal MEEVD pentapeptide motif, but not the client maturation activity of Hsp90, is essential for [URE3] prion stability. In testing deletions of various Hsp90 co-chaperones known to bind this motif, we find the immunophilin homolog Cpr7 is essential for [URE3] propagation. We show that Cpr7 interacts with Ure2 and enhances its fibrillation. The requirement of Cpr7 is specific for [URE3] as its deletion does not antagonize both strong and weak variant of another yeast prion [PSI+], suggesting a distinct role of the Hsp90 co-chaperone with different yeast prions. Our data show that, similar to the Hsp70 family, the Hsp90 chaperones also influence yeast prion maintenance, and that immunophilins could regulate protein multimerization independently of their activity as peptidyl-prolyl isomerases.

No MeSH data available.


Related in: MedlinePlus