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[SWI], the prion formed by the chromatin remodeling factor Swi1, is highly sensitive to alterations in Hsp70 chaperone system activity.

Hines JK, Li X, Du Z, Higurashi T, Li L, Craig EA - PLoS Genet. (2011)

Bottom Line: In addition, [SWI+] is lost upon overexpression of Sse nucleotide exchange factors, which act to destabilize Hsp70's interaction with client proteins.Given the plethora of genes affected by the activity of the SWI/SNF chromatin-remodeling complex, it is possible that this sensitivity of [SWI+] to the activity of Hsp70 chaperone machinery may serve a regulatory role, keeping this prion in an easily-lost, meta-stable state.Such sensitivity may provide a means to reach an optimal balance of phenotypic diversity within a cell population to better adapt to stressful environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.

ABSTRACT
The yeast prion [SWI+], formed of heritable amyloid aggregates of the Swi1 protein, results in a partial loss of function of the SWI/SNF chromatin-remodeling complex, required for the regulation of a diverse set of genes. Our genetic analysis revealed that [SWI+] propagation is highly dependent upon the action of members of the Hsp70 molecular chaperone system, specifically the Hsp70 Ssa, two of its J-protein co-chaperones, Sis1 and Ydj1, and the nucleotide exchange factors of the Hsp110 family (Sse1/2). Notably, while all yeast prions tested thus far require Sis1, [SWI+] is the only one known to require the activity of Ydj1, the most abundant J-protein in yeast. The C-terminal region of Ydj1, which contains the client protein interaction domain, is required for [SWI+] propagation. However, Ydj1 is not unique in this regard, as another, closely related J-protein, Apj1, can substitute for it when expressed at a level approaching that of Ydj1. While dependent upon Ydj1 and Sis1 for propagation, [SWI+] is also highly sensitive to overexpression of both J-proteins. However, this increased prion-loss requires only the highly conserved 70 amino acid J-domain, which serves to stimulate the ATPase activity of Hsp70 and thus to stabilize its interaction with client protein. Overexpression of the J-domain from Sis1, Ydj1, or Apj1 is sufficient to destabilize [SWI+]. In addition, [SWI+] is lost upon overexpression of Sse nucleotide exchange factors, which act to destabilize Hsp70's interaction with client proteins. Given the plethora of genes affected by the activity of the SWI/SNF chromatin-remodeling complex, it is possible that this sensitivity of [SWI+] to the activity of Hsp70 chaperone machinery may serve a regulatory role, keeping this prion in an easily-lost, meta-stable state. Such sensitivity may provide a means to reach an optimal balance of phenotypic diversity within a cell population to better adapt to stressful environments.

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[SWI+] is destabilized by growth at elevated temperatures.Wild-type [SWI+] cells were grown on glucose-based media at either 23°C, 30°C, or 37°C for 8 days to test the effect of elevated temperature on [SWI+] stability. Cells were allowed to recover for 1 day at 30°C on fresh media before assaying for the continued presence of the prion. (A) Presence or absence of [SWI+] was tested by assessing growth on raffinose-based medium. 10-fold serial dilutions of cells from each culture, or untreated control [SWI+] and [swi−] cultures, were spotted onto raffinose and glucose-based medium and grown at 30°C. The results of two independent experiments are shown. Dashed lines indicate some sections of the plate have been cropped for clarity. (B) The presence of [SWI+] in isolated colonies (n = 24) obtained from plating of cultures grown at various temperature was assessed by transformation with a plasmid carrying the Swi1NQ-YFP fusion. Resulting transformants were observed under the microscope, scoring for punctuate [SWI+] or diffuse [swi−] fluorescence. The number of transformants remaining [SWI+] is reported as a fraction of the total examined (Fraction [SWI+]).
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pgen-1001309-g009: [SWI+] is destabilized by growth at elevated temperatures.Wild-type [SWI+] cells were grown on glucose-based media at either 23°C, 30°C, or 37°C for 8 days to test the effect of elevated temperature on [SWI+] stability. Cells were allowed to recover for 1 day at 30°C on fresh media before assaying for the continued presence of the prion. (A) Presence or absence of [SWI+] was tested by assessing growth on raffinose-based medium. 10-fold serial dilutions of cells from each culture, or untreated control [SWI+] and [swi−] cultures, were spotted onto raffinose and glucose-based medium and grown at 30°C. The results of two independent experiments are shown. Dashed lines indicate some sections of the plate have been cropped for clarity. (B) The presence of [SWI+] in isolated colonies (n = 24) obtained from plating of cultures grown at various temperature was assessed by transformation with a plasmid carrying the Swi1NQ-YFP fusion. Resulting transformants were observed under the microscope, scoring for punctuate [SWI+] or diffuse [swi−] fluorescence. The number of transformants remaining [SWI+] is reported as a fraction of the total examined (Fraction [SWI+]).

Mentions: Our results described above indicate that [SWI+] is highly sensitive to various perturbations of the activity of the Hsp70 chaperone machinery brought about by ectopic expression or mutation. As it is well understood that yeast naturally encounter stressful environmental conditions known to alter chaperone expression, we wanted to ask if [SWI+] is sensitive to such conditions. We subjected our wild-type [SWI+] strain to a variety of cell stresses, including heat and ethanol shock, as well as acute exposure to severe oxidative stress. No appreciable loss of the prion was found compared to untreated control cells for any of these conditions (data not shown). However, because prion curing typically requires multiple cell divisions, we next tested whether extended growth at elevated temperatures, a condition known to cause prolonged alteration in chaperone activity altered [SWI+] stability. Indeed, cells grown for 8 days at 37°C reproducibly regained the ability to grow well on raffinose-based media, relative to cultures grown at 23°C (Figure 9A). To confirm that the improved growth on raffinose was in fact due to loss of the prion, rather than some other alteration acquired during growth at 37°C, we also assayed these cultures for Swi1 aggregation using the YFP assay. The fraction of [SWI+] cells in a culture grown at 37°C was significantly less than that from a control culture grown at 23°C (Figure 9B), confirming that unlike [PSI+] [35], [SWI+] is destabilized by prolonged growth at elevated temperatures.


[SWI], the prion formed by the chromatin remodeling factor Swi1, is highly sensitive to alterations in Hsp70 chaperone system activity.

Hines JK, Li X, Du Z, Higurashi T, Li L, Craig EA - PLoS Genet. (2011)

[SWI+] is destabilized by growth at elevated temperatures.Wild-type [SWI+] cells were grown on glucose-based media at either 23°C, 30°C, or 37°C for 8 days to test the effect of elevated temperature on [SWI+] stability. Cells were allowed to recover for 1 day at 30°C on fresh media before assaying for the continued presence of the prion. (A) Presence or absence of [SWI+] was tested by assessing growth on raffinose-based medium. 10-fold serial dilutions of cells from each culture, or untreated control [SWI+] and [swi−] cultures, were spotted onto raffinose and glucose-based medium and grown at 30°C. The results of two independent experiments are shown. Dashed lines indicate some sections of the plate have been cropped for clarity. (B) The presence of [SWI+] in isolated colonies (n = 24) obtained from plating of cultures grown at various temperature was assessed by transformation with a plasmid carrying the Swi1NQ-YFP fusion. Resulting transformants were observed under the microscope, scoring for punctuate [SWI+] or diffuse [swi−] fluorescence. The number of transformants remaining [SWI+] is reported as a fraction of the total examined (Fraction [SWI+]).
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Related In: Results  -  Collection

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pgen-1001309-g009: [SWI+] is destabilized by growth at elevated temperatures.Wild-type [SWI+] cells were grown on glucose-based media at either 23°C, 30°C, or 37°C for 8 days to test the effect of elevated temperature on [SWI+] stability. Cells were allowed to recover for 1 day at 30°C on fresh media before assaying for the continued presence of the prion. (A) Presence or absence of [SWI+] was tested by assessing growth on raffinose-based medium. 10-fold serial dilutions of cells from each culture, or untreated control [SWI+] and [swi−] cultures, were spotted onto raffinose and glucose-based medium and grown at 30°C. The results of two independent experiments are shown. Dashed lines indicate some sections of the plate have been cropped for clarity. (B) The presence of [SWI+] in isolated colonies (n = 24) obtained from plating of cultures grown at various temperature was assessed by transformation with a plasmid carrying the Swi1NQ-YFP fusion. Resulting transformants were observed under the microscope, scoring for punctuate [SWI+] or diffuse [swi−] fluorescence. The number of transformants remaining [SWI+] is reported as a fraction of the total examined (Fraction [SWI+]).
Mentions: Our results described above indicate that [SWI+] is highly sensitive to various perturbations of the activity of the Hsp70 chaperone machinery brought about by ectopic expression or mutation. As it is well understood that yeast naturally encounter stressful environmental conditions known to alter chaperone expression, we wanted to ask if [SWI+] is sensitive to such conditions. We subjected our wild-type [SWI+] strain to a variety of cell stresses, including heat and ethanol shock, as well as acute exposure to severe oxidative stress. No appreciable loss of the prion was found compared to untreated control cells for any of these conditions (data not shown). However, because prion curing typically requires multiple cell divisions, we next tested whether extended growth at elevated temperatures, a condition known to cause prolonged alteration in chaperone activity altered [SWI+] stability. Indeed, cells grown for 8 days at 37°C reproducibly regained the ability to grow well on raffinose-based media, relative to cultures grown at 23°C (Figure 9A). To confirm that the improved growth on raffinose was in fact due to loss of the prion, rather than some other alteration acquired during growth at 37°C, we also assayed these cultures for Swi1 aggregation using the YFP assay. The fraction of [SWI+] cells in a culture grown at 37°C was significantly less than that from a control culture grown at 23°C (Figure 9B), confirming that unlike [PSI+] [35], [SWI+] is destabilized by prolonged growth at elevated temperatures.

Bottom Line: In addition, [SWI+] is lost upon overexpression of Sse nucleotide exchange factors, which act to destabilize Hsp70's interaction with client proteins.Given the plethora of genes affected by the activity of the SWI/SNF chromatin-remodeling complex, it is possible that this sensitivity of [SWI+] to the activity of Hsp70 chaperone machinery may serve a regulatory role, keeping this prion in an easily-lost, meta-stable state.Such sensitivity may provide a means to reach an optimal balance of phenotypic diversity within a cell population to better adapt to stressful environments.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.

ABSTRACT
The yeast prion [SWI+], formed of heritable amyloid aggregates of the Swi1 protein, results in a partial loss of function of the SWI/SNF chromatin-remodeling complex, required for the regulation of a diverse set of genes. Our genetic analysis revealed that [SWI+] propagation is highly dependent upon the action of members of the Hsp70 molecular chaperone system, specifically the Hsp70 Ssa, two of its J-protein co-chaperones, Sis1 and Ydj1, and the nucleotide exchange factors of the Hsp110 family (Sse1/2). Notably, while all yeast prions tested thus far require Sis1, [SWI+] is the only one known to require the activity of Ydj1, the most abundant J-protein in yeast. The C-terminal region of Ydj1, which contains the client protein interaction domain, is required for [SWI+] propagation. However, Ydj1 is not unique in this regard, as another, closely related J-protein, Apj1, can substitute for it when expressed at a level approaching that of Ydj1. While dependent upon Ydj1 and Sis1 for propagation, [SWI+] is also highly sensitive to overexpression of both J-proteins. However, this increased prion-loss requires only the highly conserved 70 amino acid J-domain, which serves to stimulate the ATPase activity of Hsp70 and thus to stabilize its interaction with client protein. Overexpression of the J-domain from Sis1, Ydj1, or Apj1 is sufficient to destabilize [SWI+]. In addition, [SWI+] is lost upon overexpression of Sse nucleotide exchange factors, which act to destabilize Hsp70's interaction with client proteins. Given the plethora of genes affected by the activity of the SWI/SNF chromatin-remodeling complex, it is possible that this sensitivity of [SWI+] to the activity of Hsp70 chaperone machinery may serve a regulatory role, keeping this prion in an easily-lost, meta-stable state. Such sensitivity may provide a means to reach an optimal balance of phenotypic diversity within a cell population to better adapt to stressful environments.

Show MeSH
Related in: MedlinePlus