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首页> 外文期刊>American Journal of Physiology >High NaCl increases TonEBP/OREBP mRNA and protein by stabilizing its mRNA.
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High NaCl increases TonEBP/OREBP mRNA and protein by stabilizing its mRNA.

机译：高氯化钠通过稳定其mRNA来增加TonEBP / OREBP mRNA和蛋白质。

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摘要

Hypertonicity increases mRNA and protein abundance of the transcription factor tonicity-responsive enhancer/osmotic response element binding protein (TonEBP/OREBP), contributing to increased transcription of downstream osmoprotective genes. Previously, this was attributed to increased transcription of TonEBP/OREBP because no change was found in its mRNA stability. However, there is no direct evidence for increased transcription, and the 3'-untranslated region (UTR) of TonEBP/OREBP contains numerous adenylate/uridylate-rich elements, which can modulate RNA stability. Therefore, we have reinvestigated the effect of hypertonicity on TonEBP/OREBP mRNA stability. We find that, in mouse inner medullary collecting duct cells, raising osmolality from 300 to 500 mosmol/kgH(2)O by adding NaCl increases TonEBP/OREBP mRNA to a peak of 2.3-fold after 4 h, followed by a decline. TonEBP/OREBP protein increases to a sustained peak of 3.0-fold at 8 h. To determine the stability of TonEBP/OREBP mRNA, we measured the rate of its decrease after inhibiting transcription with actinomycin D, finding that it is stabilized for 6 h after addition of NaCl. This stabilization is sufficient to explain the increase in mRNA without any change in transcription. To investigate how hypertonicity stabilizes TonEBP/OREBP mRNA, we tested luciferase reporters containing parts of the TonEBP/OREBP mRNA UTR. Inclusion of both the 5'- and 3'-UTR increases reporter activity, consistent with mRNA stabilization. Surprisingly, however, it is the 5'-UTR that stabilizes; the 3'-UTR, by itself, decreases reporter activity. We concluded that 1) hypertonicity stabilizes TonEBP/OREBP mRNA, contributing to its increase, and 2) stabilization depends on the presence of the 5'-UTR.
机译：高渗增加转录因子张力调节增强子/渗透反应元件结合蛋白（TonEBP / OREBP）的mRNA和蛋白丰度，从而促进下游渗透保护基因的转录。以前，这归因于TonEBP / OREBP转录的增加，因为未发现其mRNA稳定性发生变化。但是，尚无直接证据表明转录增加，并且TonEBP / OREBP的3'-非翻译区（UTR）包含大量富含腺苷酸/尿苷酸的元素，可调节RNA的稳定性。因此，我们已经重新研究了高渗性对TonEBP / OREBP mRNA稳定性的影响。我们发现，在小鼠内髓收集导管细胞中，通过添加NaCl将渗透压从300 mosmol / kgH（2）O提高到4 h后，TonEBP / OREBP mRNA的峰值达到2.3倍，然后下降。 TonEBP / OREBP蛋白在8小时时增加到3.0倍的持续峰值。为了确定TonEBP / OREBP mRNA的稳定性，我们测量了用放线菌素D抑制转录后TonEBP / OREBP mRNA的下降速率，发现添加NaCl后其稳定了6小时。这种稳定足以说明mRNA的增加而转录没有任何变化。为了研究高渗如何稳定TonEBP / OREBP mRNA，我们测试了萤光素酶报告基因，其中包含部分TonEBP / OREBP mRNA UTR。同时包含5'-和3'-UTR可增加报道分子活性，与mRNA稳定化相一致。然而，令人惊讶的是，稳定的是5'-UTR。 3'-UTR本身会降低报告基因的活性。我们得出的结论是1）高渗可稳定TonEBP / OREBP mRNA，并有助于其增加; 2）稳定取决于5'-UTR的存在。

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