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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

The loss of [URE3] upon Cpr7 deletion is not due to altered expression of other major chaperones.(A) Yeast lysates from wild type (wt) [URE3], wild type [ure-o] and cpr7Δ strains were probed with antibodies against Ydj1, Sse1 and Hsp90. The chaperones were found to be expressed at similar levels in the strains examined. Loading control is same blot stained with amido-black. (B) 5μg (1X) or 10μg (2X) of total lysate protein was loaded into each lane and probed with anti Hsp70 antibodies. As seen, increased Hsp70 level was observed in strain lacking Cpr7 as compared to wt [URE3] or wt [ure-o] strains. (C) SY187(wt) expressing additional Ssa Hsp70 from transformed plasmids pRS315PSSA2-SSA1/SSA2/SSA3/SSA4 or pRS315 empty vector were spread onto leucine deficient SD solid medium with limiting adenine. As seen by white colony color phenotype of transformants, the increased expression of Ssa Hsp70 supports stable [URE3].
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pgen.1005567.g003: The loss of [URE3] upon Cpr7 deletion is not due to altered expression of other major chaperones.(A) Yeast lysates from wild type (wt) [URE3], wild type [ure-o] and cpr7Δ strains were probed with antibodies against Ydj1, Sse1 and Hsp90. The chaperones were found to be expressed at similar levels in the strains examined. Loading control is same blot stained with amido-black. (B) 5μg (1X) or 10μg (2X) of total lysate protein was loaded into each lane and probed with anti Hsp70 antibodies. As seen, increased Hsp70 level was observed in strain lacking Cpr7 as compared to wt [URE3] or wt [ure-o] strains. (C) SY187(wt) expressing additional Ssa Hsp70 from transformed plasmids pRS315PSSA2-SSA1/SSA2/SSA3/SSA4 or pRS315 empty vector were spread onto leucine deficient SD solid medium with limiting adenine. As seen by white colony color phenotype of transformants, the increased expression of Ssa Hsp70 supports stable [URE3].

Mentions: Modulation of Hsp70 activity can alter yeast prion formation and propagation. Similarly, overexpressing the Hsp70 co-chaperones Ydj1 or nucleotide exchange factor Sse1 destabilize [URE3]. In order to explore whether the loss of [URE3] in cpr7Δ strain is due to alteration in the overall abundance of Hsp70 and/or its co-chaperones, their relative expression level was monitored in [URE3] and [ure-o] cells of wild type and cpr7Δ strains. As seen in Fig 3A, Ydj1, Sse1 and Hsp90 are expressed at similar levels in both wild type and cpr7Δ strains, regardless of [URE3] status. In contrast, the Hsp70 level was increased by about 1.5–2.0 fold in the cpr7Δ strain as compared to that of wild type strains (Fig 3B).


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)

The loss of [URE3] upon Cpr7 deletion is not due to altered expression of other major chaperones.(A) Yeast lysates from wild type (wt) [URE3], wild type [ure-o] and cpr7Δ strains were probed with antibodies against Ydj1, Sse1 and Hsp90. The chaperones were found to be expressed at similar levels in the strains examined. Loading control is same blot stained with amido-black. (B) 5μg (1X) or 10μg (2X) of total lysate protein was loaded into each lane and probed with anti Hsp70 antibodies. As seen, increased Hsp70 level was observed in strain lacking Cpr7 as compared to wt [URE3] or wt [ure-o] strains. (C) SY187(wt) expressing additional Ssa Hsp70 from transformed plasmids pRS315PSSA2-SSA1/SSA2/SSA3/SSA4 or pRS315 empty vector were spread onto leucine deficient SD solid medium with limiting adenine. As seen by white colony color phenotype of transformants, the increased expression of Ssa Hsp70 supports stable [URE3].
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4608684&req=5

pgen.1005567.g003: The loss of [URE3] upon Cpr7 deletion is not due to altered expression of other major chaperones.(A) Yeast lysates from wild type (wt) [URE3], wild type [ure-o] and cpr7Δ strains were probed with antibodies against Ydj1, Sse1 and Hsp90. The chaperones were found to be expressed at similar levels in the strains examined. Loading control is same blot stained with amido-black. (B) 5μg (1X) or 10μg (2X) of total lysate protein was loaded into each lane and probed with anti Hsp70 antibodies. As seen, increased Hsp70 level was observed in strain lacking Cpr7 as compared to wt [URE3] or wt [ure-o] strains. (C) SY187(wt) expressing additional Ssa Hsp70 from transformed plasmids pRS315PSSA2-SSA1/SSA2/SSA3/SSA4 or pRS315 empty vector were spread onto leucine deficient SD solid medium with limiting adenine. As seen by white colony color phenotype of transformants, the increased expression of Ssa Hsp70 supports stable [URE3].
Mentions: Modulation of Hsp70 activity can alter yeast prion formation and propagation. Similarly, overexpressing the Hsp70 co-chaperones Ydj1 or nucleotide exchange factor Sse1 destabilize [URE3]. In order to explore whether the loss of [URE3] in cpr7Δ strain is due to alteration in the overall abundance of Hsp70 and/or its co-chaperones, their relative expression level was monitored in [URE3] and [ure-o] cells of wild type and cpr7Δ strains. As seen in Fig 3A, Ydj1, Sse1 and Hsp90 are expressed at similar levels in both wild type and cpr7Δ strains, regardless of [URE3] status. In contrast, the Hsp70 level was increased by about 1.5–2.0 fold in the cpr7Δ strain as compared to that of wild type strains (Fig 3B).

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