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Related Concept Videos

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Purification of Hsp104, a Protein Disaggregase
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Hsp70 chaperones, Ssa1 and Ssa2, limit poly(A) binding protein aggregation.

Hannah E Buchholz1, Sean A Martin1, Jane E Dorweiler1

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Loss of Hsp70 proteins in yeast causes endogenous Pab1 protein aggregation. Overexpressing other chaperones or Hsp70 can prevent this, suggesting Hsp70

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Area of Science:

  • Molecular biology
  • Cellular homeostasis
  • Protein aggregation

Background:

  • Molecular chaperones, particularly Hsp70 proteins (Ssa1 and Ssa2 in yeast), are crucial for protein folding and preventing aggregation.
  • Loss of Ssa1 and Ssa2 in yeast leads to reduced growth and lifespan, with known formation of inclusions by misfolded proteins.
  • The behavior of endogenous wild-type proteins under limited Hsp70 conditions remains unclear.

Purpose of the Study:

  • To investigate the aggregation of endogenous wild-type Poly A binding protein (Pab1) in yeast lacking Ssa1 and Ssa2.
  • To determine the role of Hsp70 and other chaperones in preventing Pab1 aggregation.
  • To understand the impact of Pab1 inclusions and stress granules on cellular recovery after heat shock.

Main Methods:

  • Utilized wild-type yeast strains with deletions in Ssa1 and Ssa2 (ssa1Δssa2Δ).
  • Observed Pab1 aggregation using Pab1-GFP fluorescence microscopy.
  • Assessed the effects of overexpressing Ssa1, Hsp104, and Sis1 on Pab1 inclusions.
  • Monitored stress granule formation and disassembly kinetics following heat shock.

Main Results:

  • In ssa1Δssa2Δ cells, wild-type Pab1 formed large cytoplasmic inclusions in about half of the cells, even without stress.
  • Overexpression of Ssa1, Hsp104, or Sis1 significantly reduced Pab1 inclusion formation.
  • Heat shock induced stress granules (SGs) in both wild-type and ssa1Δssa2Δ cells, regardless of Pab1 inclusions.
  • Cells with SGs alone disassembled faster than wild-type; cells with both Pab1 inclusions and SGs disassembled slower.
  • Pab1 inclusions were also observed in aged wild-type yeast cultures and could be partially rescued by Ssa1 overexpression.

Conclusions:

  • Hsp70 chaperones limit the aggregation of endogenous proteins like Pab1 in unstressed and aged yeast cells.
  • Elevated levels of Ssa1, Hsp104, or Sis1 can compensate for reduced Hsp70 levels, preventing aggregation.
  • The interplay between Pab1 inclusions and stress granules affects cellular recovery dynamics.
  • Depletion of active Hsp70 may contribute to age-related protein aggregation.