Gilmartin G M, Hung S L, DeZazzo J D, Fleming E S, Imperiale M J
Department of Microbiology and Molecular Genetics, Markey Center for Molecular Genetics, University of Vermont, Burlington 05405, USA.
J Virol. 1996 Mar;70(3):1775-83. doi: 10.1128/JVI.70.3.1775-1783.1996.
The adenovirus major late transcription unit (MLTU) encodes five families of mRNAs, L1 to L5, each distinguished by a unique poly(A) site. Use of the promoter-proximal L1 poly(A) site predominates during early infection, whereas poly(A) site choice shifts to the promoter-distal sites during late infection. A mini-MLTU containing only the L1 and L3 poly(A) sites has been shown to reproduce this processing switch. In vivo analysis has revealed that sequences extending 5' and 3' of the L1 core poly(A) site are required for efficient processing as well as for regulated expression. By replacement of the L1 core poly(A) site with that of the ground squirrel hepatitis virus poly(A) site, we now demonstrate that the L1 flanking sequences can enhance the processing of a heterologous poly(A). Upon recombination of the chimeric L1-ground squirrel hepatitis virus poly(A) site onto the viral chromosome, the L1 flanking sequences were also found to be sufficient to reproduce the processing switch during the course of viral infection. Subsequent in vitro analysis has shown that the L1 flanking sequences function to enhance the stability of binding of cleavage and polyadenylation specificity factor to the core poly(A) site. The impact of L1 flanking sequences on the binding of cleavage and polyadenylation specificity factor suggests that the regulation of the MLTU poly(A) site selection is mediated by the interaction of constitutive processing factors.
腺病毒主要晚期转录单位（MLTU）编码五个mRNA家族，即L1至L5，每个家族都由一个独特的聚腺苷酸化（poly(A)）位点区分。在早期感染期间，启动子近端的L1 poly(A)位点的使用占主导地位，而在晚期感染期间，poly(A)位点的选择则转向启动子远端的位点。已证明仅包含L1和L3 poly(A)位点的微型MLTU可重现这种加工转换。体内分析表明，L1核心poly(A)位点5'和3'端延伸的序列对于有效加工以及调控表达都是必需的。通过将L1核心poly(A)位点替换为地松鼠肝炎病毒的poly(A)位点，我们现在证明L1侧翼序列可以增强异源poly(A)的加工。将嵌合的L1-地松鼠肝炎病毒poly(A)位点重组到病毒染色体上后，还发现L1侧翼序列足以在病毒感染过程中重现加工转换。随后的体外分析表明，L1侧翼序列的作用是增强切割和聚腺苷酸化特异性因子与核心poly(A)位点结合的稳定性。L1侧翼序列对切割和聚腺苷酸化特异性因子结合的影响表明，MLTU poly(A)位点选择的调控是由组成型加工因子的相互作用介导的。
