TRIB3 Regulates PPARα Expression Via Proteasome-dependent Degradation To Inhibit AML Cell Apoptosis And Autophagy

Objectives:TRIB3(Tribbles homolog 3)a mammalian homolog of Drosophila tribbles,is emerging as a multifunctional oncoprotein associated with various cellular events in different tumors.This study aims to investigate the impact of TRIB3 on apoptosis and autophagy and uncover the molecular mechanisms in AML.Methods:1.TRIB3 mRNA expression from AML patients and normal human hematopoietic stem cells were analyzed in Blood Spot database.In addition,Western blot and q RT-PCR were performed to analyse the expression of TRIB3 in AML cells.2.Utilizing TRIB3-specific siRNA to silence TRIB3 in KG1 a and THP1 cells,Western blot(WB)were used to assess the levels of the apoptosis-related proteins(Cleaved PARP,Caspase3,Cleaved Caspase3)and the autophagy-associated proteins(LC3A/LC3 B,ATG7).At the same time,The apoptosis rates of si TRIB3-transfected cells were evaluated by flow cytometry after TRIB3 downregulation.3.The interaction of TRIB3 and PPARα was evaluated by immunofluorescence,coimmunoprecipitation.Furthermore,In vitro ubiquitination assays was applied to elucidated whether TRIB3 destabilized PPARα expression by promoting ubiquitin-proteasomal degradation.4.PPARα inhibitor(GW6471)was introduced into si TRIB3-transfected cells.Subsequently,the levels of the apoptosis-related proteins and the autophagy-associated proteins were assessed by Western blot(WB)in order to demonstrate whether the inhibition of TRIB3 promotes AML cell apoptosis and autophagy by degrading PPARα protein.Results:1.TRIB3 mRNA expression was markedly upregulated in AML patients.2.TRIB3 downregulation effectively induced apoptosis and autophagy in AML.3.TRIB3 interacted with PPARα and contributed to its destabilization by promoting its ubiquitination.4.Rescue experiments revealed that blocking PPARα expression using the PPARα inhibitor GW6471 reversed the functional influence of TRIB3 on AML cells.Conclusions:TRIB3 destabilizes tumor suppressor PPARα expression through ubiquitin-mediated proteasome degradation,thereby suppressing cell apoptosis and autophagy in acute myeloid leukemia...