VIRAL SMALL RNPS AND CELL TRANSFORMATION

描述：（申请人的描述）正在研究在两种疱疹病毒转化的淋巴样细胞中发现的小核RNA-蛋白（snRNP）复合物的作用。 Epstein-Barr 病毒感染并转化人类 B 细胞，是传染性单核细胞增多症的病原体，并且与多种人类癌症有关。萨米尔疱疹病毒可在新世界猴中诱导致命性淋巴瘤和白血病，并在新世界猴中转化人类和猴 T 淋巴细胞和白血病，并在培养物中转化人类和猴 T 淋巴细胞。在转化细胞中表达的少数病毒基因产物包括两个 EBV 编码的 EBER 和七个 H. saimiri 编码的 HSUR。这些小核 RNA 丰富、保守，并与宿主自身抗原相关，而宿主自身抗原是细胞 snRNP 的组成部分。然而，EBER 和 HSUR 对于转型的病毒式增长来说并不是必需的。所提出的实验基于这样的假设：EBER 和 HSUR 通过隔离重要的宿主蛋白来扰乱细胞代谢，从而促进转化。 EBER1 结合宿主核糖体蛋白 L22 的很大一部分，该蛋白参与一些急性髓系白血病患者的染色体易位。将通过定位 60S 核糖体中的 L22 结合位点并询问 L22 耗尽的核糖体是否增强核抗原 (EBNA) mRNA 中上游 AUG 的通读以增加其在潜伏期间的翻译来研究 EBER1 与 L22 相互作用的后果。 HSUR 1、2 和 5 在其 5' 末端具有 AU3A 基序，可结合 HuR，HuR 是一种宿主蛋白​​，其过度表达可稳定原癌基因、细胞因子和淋巴因子通常寿命较短的 mRNA。 HUR 的核质穿梭基序及其与几种蛋白磷酸酶 2A (PP2A) 抑制剂的相互作用，其中一种据报道具有肿瘤抑制活性，将通过突变分析进行剖析。将研究 AU3A 引导的 RNA 衰减的亚细胞定位以及 HuR 和 hnRNPD 在 mRNA 稳定性中发挥相反作用的可能性。将研究人乳头瘤病毒 E6/E7 癌蛋白的表达，以确定 PP2A 抑制剂和 HuR 是否是调节 mRNA 稳定性的磷酸化级联的组成部分。将在非洲爪蟾系统中研究 AU3A 指导的 mRNA 去腺苷酸化/衰变的机制及其在早期发育过程中的调节。将通过鉴定相互作用蛋白来寻找 EBER2 和 HSUR 3、4、6 和 7 snRNP 功能的线索。

DESCRIPTION: (Applicant's Description) The roles of small nuclear RNA- protein (snRNP) complexes found in lymphoid cells transformed by two herpesviruses are being investigated. Epstein-Barr virus infects and transforms human B cells and is the causative agent of infectious mononucleosis, as well as being associated with several human cancers. Herpesvirus saimiri induces fatal lymphomas and leukemias in New World monkeys and transforms human and monkey T lymphocytes and leukemias in New World monkeys and transforms human and monkey T lymphocytes in culture. Among the few viral gene products expressed in transformed cells are the two EBV-encoded EBERs and the seven H. saimiri-encoded HSURs. These small nuclear RNAs are abundant, conserved, and associate with host auto-antigens that are components of cellular snRNPs. Yet, both EBERs and HSURs are non-essential for viral growth of transformation. Proposed experiments are based on the hypothesis that EBERs and HSURs perturb cellular metabolism by sequestering important host proteins, thereby facilitating transformation. EBER1 binds a significant fraction of the host's ribosomal protein L22, whose is involved in chromosomal translocations in some patients with acute myeloid leukemia. Consequences of EBER1 interaction with L22 will be studied by locating the L22 binding site in the 60S ribosome and asking whether L22-depleted ribosomes enhance the read-through of upstream AUGs in nuclear antigen (EBNA) mRNAs to increase their translation during latency. HSURs 1, 2, and 5 have AU3A motifs at their 5' ends that bind HuR, a host protein whose over-expression stabilizes the normally short-lived mRNAs for proto-oncogenes, cytokines and lymphokines. HUR's nucleo- cytoplasmic shuttling motif and its interactions with several protein phosphatase 2A (PP2A) inhibitors, one of which is reported to have tumor suppressor activity will be dissected by mutational analyses. The subcellular location of AU3A-directed RNA decay and the possibility that HuR and hnRNPD play opposing roles in MRNA stability will be investigated. The expression of human papillomavirus E6/E7 oncoproteins will be studied to establish whether the PP2A inhibitors and HuR are components of a phosphorylation cascade regulating mRNA stability. The mechanism of AU3A-directed mRNA deadenylation/decay and its regulation during early development will be studied in the Xenopus system. Clues to the functions of the EBER2 and HSUR 3, 4, 6 and 7 snRNPs will be sought by identifying interaction proteins.