The ubiquitin proteasome pathway ‐ Repair or degradation of damaged proteins

Abstract

Abstract Purpose Accumulation of methylglyoxal (MGO), a highly reactive side‐product of glycolysis can modify proteins. Increased levels of MGO in cells have been implicated in diabetic vascular complications. In physiological conditions, proteolytic systems and chaperones together ensure maintenance of protein quality control. We hypothesize that MGO impairs the function of UPS and on molecular chaperones. Methods Rats with moderate type 2 diabetes (GK) and retinal epithelium pigment cell line were used. Protein oxidation was assessed by formation of carbonyl groups. Production of intracellular ROS was assessed in frozen sections of diabetic retinas by DHE incorporation. 20S proteasome activities were assessed by fluorogenic peptides. Ubiquitin (Ub) conjugation activity was determined by the ability of retinal extracts to conjugate 125I‐Ub to endogenous substrates. Ub conjugates, Hsp90, Hsc70, Hsp40 and CHIP levels were assessed by WB. Cell viability was determined by MTT while proliferation was assessed by BrdU‐incorporation. Results Data show that accumulation of endogenous Ub conjugates in the presence of MGO is associated with an increased ability of retinal extracts to conjugate 125I‐Ub to endogenous substrates. Moreover, MGO significantly decreases the 20S ptoteasome activity. Data further show that MGO decreases the levels of the molecular chaperones Hsp90 and Hsc70 and promotes aggregation of Hsp40 and CHIP. Moreover, these aggregates revealed immunoreactivity against Ub. Consistently, these effects are associated with increased cell protein oxidation, decreased cell proliferation and viability. Conclusion In diabetes, accumulation of MGO may impair the UPP and the protein quality control, leading to accumulation of obsolete proteins and cell injury.