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To investigate the effect of proteasome activator PSME1 on proteasome function and antioxidant stress capacity in endothelial cells.
An adenoviral vector (Ad-PSME1) was used to transduce the target gene into human umbilical vein endothelial cells (HUVEC) to establish PSME1-overexpressing endothelial cells. Western blot was used to detect the expression levels of proteasome-related proteins; dot blot was employed to assess protein carbonylation levels; co-immunoprecipitation(Co-IP) was performed to examine the interaction between PSME1 and PSME2; fluorogenic substrate assays were used to determine proteasome activity;cycloheximide(CHX) chase experiments were conducted to evaluate protein degradation rates; pulse-chase assays were applied to measure the degradation rate of GFPu; and DNPH derivatization was used to detect protein carbonylation levels.
Overexpression of PSME1 in HUVECs increased the expression and prolonged the half-life of PSME2, enhanced chymotrypsin-like (β5) proteasome activity and ATP-dependent proteolytic function, and significantly shortened the half-life of misfolded protein GFPu(P<0.05). No significant effects were observed on the stability of endogenous UPS substrates(AKT, GATA4, and PTEN) or on the abundance of the 19S and 20S proteasome complexes (RPN2, RPT6, and PSMB5) (P>0.05). Additionally, PSME1 overexpression significantly reduced hydrogen peroxide-induced protein carbonylation levels and enhanced the antioxidant stress capacity of endothelial cells(P<0.05).
Overexpression of PSME1 stabilizes PSME2, enhances the function of the 11S regulatory particle, and improves the proteasome's ability to degrade misfolded and oxidatively damaged proteins, thereby protecting endothelial cells.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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