Gan Zhuohui, Powell Frank L, Zambon Alexander C, Buchholz Kyle S, Fu Zhenxing, Ocorr Karen, Bodmer Rolf, Moya Esteban A, Stowe Jennifer C, Haddad Gabriel G, McCulloch Andrew D
School of Basic Medical Sciences, Wenzhou Medical University, Wenzhou, 325035, Zhejiang, China.
Department of Bioengineering, University of California San Diego, La Jolla, CA, 92093, USA.
J Physiol. 2017 Sep 1;595(17):5797-5813. doi: 10.1113/JP274556. Epub 2017 Jul 27.
Changes in gene expression that occur within hours of exposure to hypoxia in in vivo skeletal muscles remain unexplored. Two hours of hypoxia caused significant down-regulation of extracellular matrix genes followed by a shift at 6 h to altered expression of genes associated with the nuclear lumen while respiratory and blood gases were stabilized. Enrichment analysis of mRNAs classified by stability rates suggests an attenuation of post-transcriptional regulation within hours of hypoxic exposure, where PI3K-Akt signalling was suggested to have a nodal role by pathway analysis. Experimental measurements and bioinformatic analyses suggested that the dephosphorylation of Akt after 2 h of hypoxic exposure might deactivate RNA-binding protein BRF1, hence resulting in the selective degradation of mRNAs.
The effects of acute hypoxia have been widely studied, but there are few studies of transcriptional responses to hours of hypoxia in vivo, especially in hypoxia-tolerant tissues like skeletal muscles. We used RNA-seq to analyse gene expression in plantaris muscles while monitoring respiration, arterial blood gases, and blood glucose in mice exposed to 8% O for 2 or 6 h. Rapid decreases in blood gases and a slower reduction in blood glucose suggest stress, which was accompanied by widespread changes in gene expression. Early down-regulation of genes associated with the extracellular matrix was followed by a shift to genes associated with the nuclear lumen. Most of the early down-regulated genes had mRNA half-lives longer than 2 h, suggesting a role for post-transcriptional regulation. These transcriptional changes were enriched in signalling pathways in which the PI3K-Akt signalling pathway was identified as a hub. Our analyses indicated that gene targets of PI3K-Akt but not HIF were enriched in early transcriptional responses to hypoxia. Among the PI3K-Akt targets, 75% could be explained by a deactivation of adenylate-uridylate-rich element (ARE)-binding protein BRF1, a target of PI3K-Akt. Consistent decreases in the phosphorylation of Akt and BRF1 were experimentally confirmed following 2 h of hypoxia. These results suggest that the PI3K-Akt signalling pathway might play a role in responses induced by acute hypoxia in skeletal muscles, partially through the dephosphorylation of ARE-binding protein BRF1.
体内骨骼肌在暴露于低氧环境数小时内发生的基因表达变化仍未得到充分研究。两小时的低氧导致细胞外基质基因显著下调，随后在6小时时转变为与核腔相关基因的表达改变，而呼吸和血气则保持稳定。根据稳定性速率分类的mRNA富集分析表明，低氧暴露数小时内转录后调控减弱，通路分析表明PI3K-Akt信号传导起关键作用。实验测量和生物信息学分析表明，低氧暴露2小时后Akt的去磷酸化可能使RNA结合蛋白BRF1失活，从而导致mRNA的选择性降解。
急性低氧的影响已得到广泛研究，但对体内数小时低氧的转录反应研究较少，尤其是在骨骼肌等耐低氧组织中。我们使用RNA测序分析了在暴露于8%氧气环境2或6小时的小鼠中，比目鱼肌的基因表达，同时监测呼吸、动脉血气和血糖。血气快速下降和血糖缓慢降低表明存在应激，同时伴随着基因表达的广泛变化。与细胞外基质相关的基因早期下调，随后转变为与核腔相关的基因。大多数早期下调的基因mRNA半衰期超过2小时，表明转录后调控起作用。这些转录变化在信号通路中富集，其中PI3K-Akt信号通路被确定为枢纽。我们的分析表明，PI3K-Akt而非HIF的基因靶点在低氧早期转录反应中富集。在PI3K-Akt靶点中，75%可由富含腺苷酸-尿苷酸元件（ARE）结合蛋白BRF1（PI3K-Akt的一个靶点）的失活来解释。实验证实，低氧2小时后Akt和BRF1的磷酸化持续下降。这些结果表明，PI3K-Akt信号通路可能在骨骼肌急性低氧诱导的反应中起作用，部分是通过ARE结合蛋白BRF1的去磷酸化实现的。
