Role of EB virus nuclear protein EBNA3C in B-cell growth transformation

EB病毒(Epstein-Barr Virus，EBV)感染可在体外将人原代B细胞转化为无限期增殖的淋巴母细胞样细胞系(LCLS)。EBV核蛋白EBNA3C是这种生长转化所必需的。为了研究EBNA3C如何促进LCL的生长，我们建立了表达条件活性EBNA3C的LCL。EBNA3C失活导致LCLS生长停滞。我们还发现，EBNA3C失活导致pl6(Ink4a)表达的诱导伴随着pRb磷酸化的降低。因此，EBNA3C可能通过抑制p16(Ink4a)的表达而参与LCL的生长维持。EBNA3C与序列特异性DNA结合蛋白RBP-Jkappa结合并调节RBP-Jkappa依赖的转录。我们研究了EBNA3C与RBP-Jkappa的结合是否对LCL的生长维持至关重要。对一系列EBNA3C突变体的分析表明，EBNA3C突变体通过RBP-Jkappa调控转录的能力与其维持LCL生长维持的能力密切相关。这些数据表明，EBNA3C通过RBP-Jkappa对转录的调控是维持LCL生长的关键。我们还评估了一系列EBNA3C缺失突变体支持LCL生长的能力，以确定EBNA3C的关键或可缺少的区域。我们发现EBNA3C的氨基酸残基(AA)1-506和AS-733-909是维持LCL生长的关键。这些区域在EBV和非人灵长类淋巴裂隙病毒之间具有氨基酸序列同源性和功能同源性，这表明LCL维持生长所必需的区域在进化上是保守的。

Epstein-Barr virus (EBV) infection converts primary human B cells into indefinitely proliferating lymphoblastoid cell lines (LCLs) in vitro. EBV nuclear protein EBNA3C is essential for this growth transformation. To investigate how EBNA3C contributes to LCL growth, we established LCLs that express a conditionally active EBNA3C. EBNA3C inactivation resulted in growth arrest of LCLs. We also found that EBNA3C inactivation caused the induction of pl6(INK4A) expression accompanied by the decrease of pRb phosphorylation. Thus, it is likely that EBNA3C contributes LCL growth maintenance through inhibiting p16(INK4A) expression.EBNA3C associates with the sequence-specific DNA binding protrein RBP-Jkappa and regulates RBP-Jkappa dependent transcription We examined whether EBNA3C association with RBP-Jkappa is critical for LCL growth maintenance. Analyses using a series of EBNA3C mutant revealed that the ability of EBNA3C mutant to regulate transcription through RBP-Jkappa was exactly correlated with its ability to sustain LCL growth maintenance. The data indicates that EBNA3C regulation of transcription through RBP-Jkappa is critical for maintaining LCL growth.We also evaluated a series of EBNA3C deletion mutant for the ability to support LCL growth to identify the critical or dispensable regions of EBNA3C. We found that the amino acid residues (aa) 1 to 506, and the as 733 to 909 of EBNA3C are critical for LCL growth maintenance. These regions were previously noted to have amino acid sequence homology and functional homology among EBV and nonhuman primate lymphocriptviruses, suggesting that the regions essential for LCL growth maintenance are evolutionally conserved.