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Objectives:To determine the mechanisms of ubiquitination in postmenopausal osteoporosis and investigate the ubiquitinated spectrum of novel targets between healthy postmenopausal women and postmenopausal osteoporosis patients, we performed ubiquitylome analysis of the whole blood of postmenopausal women and postmenopausal osteoporosis patients. Methods:To obtain a more comprehensive understanding of the postmenopausal osteoporosis mechanism, we performed a quantitative assessment of the ubiquitylome in whole blood from seven healthy postmenopausal women and seven postmenopausal osteoporosis patients using high-performance liquid chromatography fractionation, affinity enrichment, and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). To examine the ubiquitylome data, we performed enrichment analysis using an ubiquitylated amino acid motif, Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway.Results: Altogether, 133 ubiquitinated sites and 102 proteins were quantified. A difference of more than 1.2 times is considered significant upregulation and less than 0.83 significant downregulation; 32 ubiquitinated sites on 25 proteins were upregulated and 101 ubiquitinated sites on 77 proteins were downregulated. These quantified proteins, both with differently ubiquitinated sites, participated in various cellular processes, such as cellular processes, biological regulation processes, response to stimulus processes, single-organism and metabolic processes. Ubiquitin conjugating enzyme activity and ubiquitin-like protein conjugating enzyme activity were the most highly enriched in molecular function of upregulated sites with corresponding proteins, but they were not enriched in downregulated in sites with corresponding proteins. The KEGG pathways analysis of quantified proteins with differentiated ubiquitinated sites found 13 kinds of molecular interactions and functional pathways, such as glyoxylate and decarboxylate metabolism, dopaminergic synapse, ubiquitin-mediated proteolysis, salivary secretion, coagulation and complement cascades, Parkinson's disease, and hippo signaling pathway. In addition, hsa04120 ubiquitin-mediated proteolysis was the most highly enriched in proteins with upregulated sites, hsa04610 complement and coagulation cascades was the most highly enriched in proteins with downregulated ubiquitinated sites, and hsa04114 Oocyte meiosis was the most highly enriched among all differential proteins. Conclusion:Our study expands the understanding of the spectrum of novel targets that are differentially ubiquitinated in whole blood from healthy postmenopausal women and postmenopausal osteoporosis patients. The findings will contribute toward our understanding of the underlying proteostasis pathways in postmenopausal osteoporosis and the potential identification of diagnostic biomarkers in whole blood.

Long‐term arsenic exposure is a worldwide public health problem that causes serious harm to human health. The liver is the main target organ of arsenic toxicity; arsenic induces disruption of the DNA damage repair pathway, but its mechanisms remain unclear. In recent years, studies have found that epigenetic mechanisms play an important role in arsenic‐induced lesions. In this study, we conducted experiments in vitro using normal human liver cells (L‐02) to explore the mechanism by which the histone demethylase JHDM2A regulates H3K9 dimethylation (me2) in response to arsenic‐induced DNA damage. Our results indicated that arsenic exposure upregulated the expression of JHDM2A, downregulated global H3K9me2 modification levels, increased the H3K9me2 levels at the promoters of base excision repair (BER) genes (N‐methylpurine‐DNA glycosylase [MPG], XRCC1 and poly(ADP‐ribose)polymerase 1) and inhibited their expression levels, causing DNA damage in cells. In addition, we studied the effects of overexpression and inhibition of JHDM2A and found that JHDM2A can participate in the molecular mechanism of arsenic‐induced DNA damage via the BER pathway, which may not be involved in the BER process because H3K9me2 levels at the promoter region of the BER genes were unchanged following JHDM2A interference. These results suggest a potential mechanism by which JHDM2A can regulate the MPG and XRCC1 genes in the process of responding to DNA damage induced by arsenic exposure and can participate in the process of DNA damage repair, which provides a scientific basis for understanding the epigenetic mechanisms and treatments for endemic arsenic poisoning.