Curran Kristen L, Allen Latoya, Porter Brittany Bronson, Dodge Joseph, Lope Chelsea, Willadsen Gail, Fisher Rachel, Johnson Nicole, Campbell Elizabeth, VonBergen Brett, Winfrey Devon, Hadley Morgan, Kerndt Thomas
University of Wisconsin-Whitewater, Department of Biological Sciences, Whitewater, Wisconsin, United States of America.
PLoS One. 2014 Sep 19;9(9):e108266. doi: 10.1371/journal.pone.0108266. eCollection 2014.
We have been investigating whether xBmal1 and xNocturnin play a role in somitogenesis, a cyclic developmental process with an ultradian period. Previous work from our lab shows that circadian genes (xPeriod1, xPeriod2, xBmal1, and xNocturnin) are expressed in developing somites. Somites eventually form the vertebrae, muscles of the back, and dermis. In Xenopus, a pair of somites is formed about every 50 minutes from anterior to posterior. We were intrigued by the co-localization of circadian genes in an embryonic tissue known to be regulated by an ultradian clock. Cyclic expression of genes involved in Notch signaling has been implicated in the somite clock. Disruption of Notch signaling in humans has been linked to skeletal defects in the vertebral column. We found that both depletion (morpholino) and overexpression (mRNA) of xBMAL1 protein (bHLH transcription factor) or xNOCTURNIN protein (deadenylase) on one side of the developing embryo led to a significant decrease in somite number with respect to the untreated side (p<0.001). These manipulations also significantly affect expression of a somite clock component (xESR9; p<0.05). We observed opposing effects on somite size. Depletion of xBMAL1 or xNOCTURNIN caused a statistically significant decrease in somite area (quantified using NIH ImageJ; p<0.002), while overexpression of these proteins caused a significant dose dependent increase in somite area (p<0.02; p<0.001, respectively). We speculate that circadian genes may play two separate roles during somitogenesis. Depletion and overexpression of xBMAL1 and NOCTURNIN both decrease somite number and influence expression of a somite clock component, suggesting that these proteins may modulate the timing of the somite clock in the undifferentiated presomitic mesoderm. The dosage dependent effects on somite area suggest that xBMAL1 and xNOCTURNIN may also act during somite differentiation to promote myogenesis.
我们一直在研究xBmal1和xNocturnin在体节发生过程中是否发挥作用，体节发生是一个具有超日周期的周期性发育过程。我们实验室之前的研究表明，昼夜节律基因（xPeriod1、xPeriod2、xBmal1和xNocturnin）在发育中的体节中表达。体节最终形成椎骨、背部肌肉和真皮。在非洲爪蟾中，从前向后大约每50分钟形成一对体节。我们对昼夜节律基因在已知受超日时钟调节的胚胎组织中的共定位很感兴趣。参与Notch信号通路的基因的周期性表达与体节时钟有关。人类Notch信号通路的破坏与脊柱骨骼缺陷有关。我们发现，在发育中的胚胎一侧对xBmal1蛋白（bHLH转录因子）或xNocturnin蛋白（去腺苷酸酶）进行缺失（吗啉代）和过表达（mRNA）处理，相对于未处理的一侧，体节数量显著减少（p<0.001）。这些操作也显著影响体节时钟成分（xESR9）的表达（p<0.05）。我们观察到对体节大小有相反的影响。xBmal1或xNocturnin的缺失导致体节面积在统计学上显著减小（使用NIH ImageJ进行量化；p<0.002），而这些蛋白的过表达导致体节面积显著的剂量依赖性增加（分别为p<0.02；p<0.001）。我们推测昼夜节律基因可能在体节发生过程中发挥两个不同的作用。xBmal1和Nocturnin的缺失和过表达都会减少体节数量并影响体节时钟成分的表达，这表明这些蛋白可能在未分化的前体节中胚层调节体节时钟的时间。对体节面积的剂量依赖性影响表明，xBmal1和xNocturnin也可能在体节分化过程中发挥作用以促进肌发生。
