Ubiquitin-conjugating Enzyme E2M Destabilizes Heat Shock Protein 70 and Mediates Sleep Deprivation-Induced Cognitive Impairment in Mice.

Sleep deprivation (SD) contributes to cognitive impairment. Heat shock protein 70 (HSP70) protects against SD-induced neuroinflammation and cognitive decline; however, how endogenous HSP70 is regulated under SD conditions, and in particular whether it is controlled by post-translational modifications such as ubiquitination, remains unclear. The present study examined whether the ubiquitin-conjugating enzyme E2M (UBE2M) governs HSP70 protein stability and thereby contributes to SD-related inflammatory and cognitive outcomes. A mouse model of SD was established using the modified multiple-platform water environment method. Protein and mRNA levels of HSP70 and UBE2M in the hippocampus were evaluated by Western blot and qRT-PCR, respectively. Co-immunoprecipitation was used to analyze the interaction between UBE2M and HSP70 and to assess HSP70 poly-ubiquitination. Cycloheximide (CHX) chase assays were performed to determine HSP70 protein stability. Cognitive performance was assessed using the Y-maze and the novel object recognition test, while hippocampal inflammation was evaluated by measuring pro-inflammatory cytokines (TNF-α, IL-1β, MCP-1) using ELISA. Oxidative stress was evaluated by quantifying MDA content, SOD activity and total antioxidant capacity. SD significantly downregulated HSP70 protein levels in the hippocampus without altering its mRNA expression, indicating post-translational regulation. UBE2M expression was markedly upregulated in the hippocampus during SD, and a direct interaction between UBE2M and HSP70 was confirmed. UBE2M promoted HSP70 poly-ubiquitination, leading to reduced HSP70 protein stability as shown by CHX chase assays. Mice treated with recombinant UBE2M showed aggravated cognitive impairment in the Y-maze and the novel object recognition test, together with exacerbated hippocampal oxidative stress and elevated levels of hippocampal pro-inflammatory cytokines. This study identifies UBE2M as a key regulator of HSP70 protein stability and indicates that the UBE2M/HSP70 axis links SD to hippocampal oxidative stress, neuroinflammation and cognitive impairment, thereby suggesting a potential therapeutic target for sleep-related cognitive disorders.