UBE4B-driven suppression of megakaryocyte maturation by HBV: A novel mechanism linking viral infection to thrombocytopenia via p53-ERK1/2 crosstalk

Background and aim: Hepatitis B virus (HBV) infection can lead to thrombocytopenia through its effects on hematopoiesis, although the underlying mechanisms have not been fully elucidated. Platelet production involves multiple stages, including the differentiation of mature megakaryocytes, which plays a pivotal role. In this study, we assessed the variances in megakaryocyte differentiation and maturation after HBV infection and investigated the molecular mechanism involved. Methods: Different doses of HBV virus solution extracted from HepAD38 cells were co-cultured with hematopoietic stem cells that had been isolated from fresh, full-term healthy maternal cord blood through magnetic bead sorting. Label-free quantitative proteomics was employed to analyze the differential proteins during the mature megakaryocyte stages pre- and post-HBV infection, with a focus on elucidating their respective molecular mechanisms. Results: A comparative analysis revealed that HBV impeded the differentiation of fully developed megakaryocytes. Its suppressive effect was more pronounced in the later stages of differentiation, consequently leading to impaired platelet production. Proteomic analysis revealed noteworthy disparities in UBE4B protein levels in mature megakaryocytes after HBV infection. Upon transfection with lentivirus and subsequent knockdown of UBE4B in mature megakaryocytes, a noticeable alleviation of HBV's inhibitory impact was observed, accompanied by regulation of p53 and ERK1/2 expression and phosphorylation. Conclusion: HBV upregulates UBE4B expression, which inhibits p53 expression and phosphorylation while enhancing ERK1/2 expression and phosphorylation. This cascade suppresses mature megakaryocyte differentiation, resulting in impaired platelet production.