ALTERNATIVE SPLICING OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 MESSENGER-RNA MODULATES VIRAL PROTEIN EXPRESSION, REPLICATION, AND INFECTIVITY
ALTERNATIVE SPLICING OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 MESSENGER-RNA MODULATES VIRAL PROTEIN EXPRESSION, REPLICATION, AND INFECTIVITY
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DOI：
10.1128/jvi.67.11.6365-6378.1993
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发表时间：
1993-11-01
影响因子：
5.4
通讯作者：
MARTIN, MA
中科院分区：
文献类型：
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作者：
PURCELL, DFJ;MARTIN, MA
人类免疫缺陷病毒 1 型 (HIV-1) 基因组 RNA 内存在多个 RNA 剪接位点，这些位点能够为多种病毒蛋白中的每一种合成许多 mRNA。我们评估了外周血淋巴细胞和人类 T 细胞系的 HIV-1 感染过程中可变剪接 mRNA 种类的生物学意义，以确定可变 RNA 剪接在调节 HIV-1 复制和感染中的潜在作用。首先，我们使用放射性标记的 cDNA 进行半定量聚合酶链式反应进行凝胶分析，以确定不同阵列的选择性剪接的 HIV-1 mRNA 的相对丰度。主要的 rev、tat、vpr 和 env RNA 含有最少的非编码序列，但主要的 nef mRNA 剪接不完全，并且总是包含非编码外显子。其次，在对 HIV-1NL4-3 的主要 5' 剪接供体和几个隐秘的、组成性的和竞争性 3' 剪接受体基序进行诱变后，测量了 RNA 加工改变的影响。 消除组成型剪接位点的突变导致新的隐藏位点的激活；其中一些保留了生物功能。消除竞争性剪接受体位点的突变导致病毒衍生的 mRNA 库发生显着改变，在某些情况下，还导致病毒感染性和/或病毒蛋白谱发生显着改变。 HIV-1基因组中的冗余RNA剪接信号和选择性剪接的mRNA提供了一种调节HIV-1蛋白相对比例以及在某些情况下病毒感染性的机制。
Multiple RNA splicing sites exist within human immunodeficiency virus type 1 (HIV-1) genomic RNA, and these sites enable the synthesis of many mRNAs for each of several viral proteins. We evaluated the biological significance of the alternatively spliced mRNA species during productive HIV-1 infections of peripheral blood lymphocytes and human T-cell lines to determine the potential role of alternative RNA splicing in the regulation of HIV-1 replication and infection. First, we used a semiquantitative polymerase chain reaction of cDNAs that were radiolabeled for gel analysis to determine the relative abundance of the diverse array of alternatively spliced HIV-1 mRNAs. The predominant rev, tat, vpr, and env RNAs contained a minimum of noncoding sequence, but the predominant nef mRNAs were incompletely spliced and invariably included noncoding exons. Second, the effect of altered RNA processing was measured following mutagenesis of the major 5' splice donor and several cryptic, constitutive, and competing 3' splice acceptor motifs of HIV-1NL4-3. Mutations that ablated constitutive splice sites led to the activation of new cryptic sites; some of these preserved biological function. Mutations that ablated competing splice acceptor sites caused marked alterations in the pool of virus-derived mRNAs and, in some instances, in virus infectivity and/or the profile of virus proteins. The redundant RNA splicing signals in the HIV-1 genome and alternatively spliced mRNAs provides a mechanism for regulating the relative proportions of HIV-1 proteins and, in some cases, viral infectivity.