HSP70 and p62/SQSTM 1 regulate differently autophagy clearance in ARPE‐19 cells

• Published in: Acta ophthalmologica (Oxford, England) Vol. 87; no. s244
• Main Authors: VIIRI, J, HYTTINEN, J, RYHäNEN, T, SALMINEN, A, KAARNIRANTA, K
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01-09-2009
• ISSN: 1755-375X; 1755-3768
• DOI: 10.1111/j.1755-3768.2009.216.x

Purpose Prior to proteolysis, heat shock proteins (HSPs) attempt to refold misfolded proteins. If this process is not successful proteins are degradated in proteasomes or in lysosomes. In the present study, the roles of the HSP70 and the p62/SQSTM 1 in autophagy clearance were evaluated in ARPE‐19 cells after proteasome inhibitor treatment.
Methods The HSP70, p62/SQSTM 1 and ubiquitin localization were analyzed by immunofluorescense. Confocal and transmission electron microscopy were used to detect cellular organelles and to analyze the morphological changes. HSP70 and p62/SQSTM 1 levels were modulated using RNA interference techniques. Cell viability was measured by colorimetric assay.
Results The proteasome inhibitor MG‐132 evoked the accumulation of perinuclear aggregates positive for HSP70, p62/SQSTM 1 and ubiquitin‐protein conjugates. We observed that the aggregation was reversible: a cessation of proteasome inhibition led to clearance of the deposits via autophagy. Interestingly, p62/SQSTM 1 mRNA depletion delayed autophagy clearance and significantly increased cell death in conjunction with proteasome inhibition. The hsp70 RNA interference did not change autophagy clearance, but increased cell death in response to proteasome inhibition.
Conclusion The HSP70 and p62/SQSTM 1 regulate differently autophagy clearance in RPE cells. Autophagy seems to be an important mechanism to clean proteasome inhibitor –induced protein aggregates.