The Regulatory Function Of Deubiquitinase USP50 Through NF-?B Signaling Pathway In Talaromyces Marneffei Infection

Objectives:To investigate the effect and regulation of deubiquitinase USP50 upon related signaling pathway in Talaromyces marneffei(TM)infected-THP-1 macrophages and understand the innate immune mechanism of host cells,so as to provide the evidence for the treatment of TM infection in clinical.Methods:TM was identified by Matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF MS).THP-1 cells were induced and differentiated into THP-1 macrophages by phorbol 12-myristate 13-acetate(PMA).PCR array technique was used to screen differential gene expressions of 88 deubiquitinases in TM infected-THP-1 macrophages.Meanwhile,the activation of the associated signaling pathways were also screened by transcriptome sequencing.The expression levels of deubiquitinase USP50 and TLR signaling pathway molecules were analyzed by Western blot.In addition,the high and low expressions of USP50 gene were constructed by lentivirus mediated overexpression and RNA interference technology in THP-1 cells.Subsequently,cocultured with TM spores,the expression levels of downstream signaling pathway molecules were analyzed by Western blot.The concentrations of TNF-? and IL-8 inflammatory cytokines were detected by ELISA Results:The expression of deubiquitinase USP50 was significantly up-regulated in TM-stimulated THP-1 macrophages by PCR-Array,while TLR signaling pathways were remarkably activated by transcriptome sequencing.The expression of TLR4,USP50 and TRAF6 proteins were significantly up-regulated in TM-infected THP-1 macrophages.Western blot analysis showed that the overexpression of USP50 upregulated the phosphorylation of NF-?B p65,while RNA interference inhibited the phosphorylation of NF-?B p65.Stimulated by TM spores in THP-1 macrophages,the levels of TNF-? and IL-8 inflammatory factors in cell culture supernatants increased significantly at 8h and 18h,respectively.However,the secretion of TNF-? and IL-8 were also inhibited by USP50 downregulation(P<0.01).Conclusions:The expressions of TLR4,USP50 and TRAF6 are upregulated in TM-stimulated THP-1 macrophages.Whereas,deubiquitinase USP50 may positively regulate the phosphorylation of NF-?B.It is postulated that deubiquitinase USP50 regulates the inflammatory cytokine secretions through the TLR4/TRAF6/NF-?B signaling pathway under TM infection.