Impact of the segment-specific region of the 3'-untranslated region of the influenza A virus PB1 segment on protein expression
Impact of the segment-specific region of the 3'-untranslated region of the influenza A virus PB1 segment on protein expression
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DOI：
10.1007/s11262-013-0969-0
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发表时间：
2013-12-01
期刊：
影响因子：
1.6
通讯作者：
Cao, Yongchang
中科院分区：
文献类型：
--
作者：
Ma, Jun;Liu, Kang;Cao, Yongchang
甲型流感病毒基因组的3 '-和5'-非翻译区（UTR）中的12和13个末端核苷酸分别对于病毒RNA的转录和mRNA的翻译是重要的。然而，UTR的片段特异性区域的功能并不为人所知。我们利用增强型绿色荧光蛋白（eGFP）作为报告基因，评估这些UTR在体外对蛋白表达的影响。结果显示，NP-eGFP、NS-eGFP和HA-eGFP的蛋白表达水平高于其他报告基因的蛋白表达水平，并且PB 1-eGFP的蛋白水平在转染后48小时保持在相对低的量。结果表明，各区段的UTR对蛋白表达水平的影响不同，其中PB 1区段的UTR对蛋白表达的影响显著。在PB 1 - 3 '-UTR中缺失“UAAA”和“UAAACU”基序使蛋白表达水平分别显著增加49.8%和142.6%。这一发现表明，PB 1 - 3 '-UTR中的“UAAACU”基序至少部分地负责低蛋白表达水平。通过在相似位置将“UAAACU”基序引入其他3 '-UTR（PA、NS、NP和HA）中，eGFP表达如预期分别降低56、61、22和22%。这一结果进一步证实了PB 1 - 3 '-UTR的“UAAACU”基序可以抑制蛋白质表达。我们的研究结果表明，在UTR中的片段特异性区域，而不仅仅是UTR的保守区域在病毒蛋白表达中起重要作用。此外，报告的发现也可能揭示甲型流感病毒基因组的新调控机制。
The 12 and 13 terminal nucleotides in the 3'- and 5'-untranslated regions (UTRs) of the influenza A virus genome, respectively, are important for the transcription of the viral RNA and the translation of mRNA. However, the functions of the segment-specific regions of the UTRs are not well known. We utilized an enhanced green fluorescent protein (eGFP) flanked at both ends by different UTRs (from the eight segments of H1N1 PR8/34) as a reporter gene to evaluate the effects of these UTRs on protein expression in vitro. The results showed that the protein expression levels of NP-eGFP, NS-eGFP, and HA-eGFP were higher than those of the other reporters and that the protein level of PB1-eGFP remained at a relatively low amount 48-h post-transfection. The results revealed that the UTRs of all segments differently affected the protein expression levels and that the effect of the UTRs of PB1 segment on protein expression was significant. The deletion of "UAAA" and "UAAACU" motifs in the PB1-3'-UTR significantly increased the protein expression level by 49.8 and 142.6 %, respectively. This finding suggests that the "UAAACU" motif in the PB1-3'-UTR is at least partly responsible for the low protein expression level. By introducing the "UAAACU" motif into other 3'-UTRs (PA, NS, NP, and HA) at similar locations, the eGFP expression was reduced as expected by 56, 61, 22, and 22 %, respectively. This result further confirmed that the "UAAACU" motif of the PB1-3'-UTR can inhibit protein expression. Our findings suggest that the segment-specific regions in the UTRs and not just the conserved regions of the UTRs play an important role in the viral protein expression. Additionally, the reported findings may also shed light on novel regulatory mechanism for the influenza A virus genome.