Ishikawa Kosuke, Azuma Sakura, Ikawa Shuntaro, Morishita Yasuyuki, Gohda Jin, Akiyama Taishin, Semba Kentaro, Inoue Jun ichiro
Department of Cancer Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Tokyo 108-8639, Minato, Japan.
Gene. 2003 Dec 11;322:105-12. doi: 10.1016/j.gene.2003.08.016.
The developmentally regulated GTP-binding protein (DRG) subfamily is an uncharacterized member of the Obg family, an evolutional branch of GTPase superfamily proteins. GTPases act as molecular switches regulating diverse cellular processes. DRG2 and DRG1 comprise the DRG subfamily in eucaryotes. Although drg1 was first identified as a gene predominantly expressed during early development of the mouse central nervous system, comparative analysis of drg2 and drg1 expression during embryogenesis has never been reported, and the biochemical properties of the DRG family proteins remain to be elucidated. Thus, we first cloned Xenopus drg2 (Xdrg2) and examined the temporal and spatial expression patterns of Xdrg2 mRNA in comparison to those of Xdrg1. Both Xdrg2 and Xdrg1 are induced at late gastrula and subsequently increased during later stages of embryos (stage 13-41). Whole-mount in situ hybridization showed that Xdrg2 and Xdrg1 expression patterns are almost identical except that only Xdrg2 expression is detected in the stage 22 pronephric anlage. Strong transcripts of both genes are also observed at this stage in neural crest cells, blood islands, and developing eyes, and in brain, eyes, otic vesicle, branchial arches, pronephroses, spinal cord, notochord, head mesenchyme, and somites at stages 27 and 32. Northern blot analysis of adult tissues revealed that both genes are expressed highly in ovary and testis and rather moderately in other organs, except that Xdrg1 transcripts are scarcely detected in heart, lung, and liver. Accordingly, transcription or stability of Xdrg2 and Xdrg1 mRNAs may be regulated by different mechanisms. In addition, by generating recombinant XDRG2 and XDRG1 proteins, we found the RNA binding activity of these proteins in vitro. Our results suggest that the DRG proteins may play their physiological roles via RNA binding.
发育调控型GTP结合蛋白（DRG）亚家族是Obg家族中一个未被充分研究的成员，Obg家族是GTP酶超家族蛋白的一个进化分支。GTP酶作为分子开关，调控着多种细胞过程。DRG2和DRG1构成了真核生物中的DRG亚家族。尽管drg1最初被鉴定为在小鼠中枢神经系统早期发育过程中主要表达的基因，但尚未有关于胚胎发育过程中drg2和drg1表达的比较分析报道，DRG家族蛋白的生化特性也有待阐明。因此，我们首先克隆了非洲爪蟾的drg2（Xdrg2），并与Xdrg1相比较，研究了Xdrg2 mRNA的时空表达模式。Xdrg2和Xdrg1均在原肠胚晚期被诱导表达，随后在胚胎后期（13 - 41期）表达量增加。整体原位杂交显示，Xdrg2和Xdrg1的表达模式几乎相同，只是在22期的原肾原基中仅检测到Xdrg2的表达。在这个阶段，神经嵴细胞、血岛和发育中的眼睛以及在27期和32期的脑、眼睛、耳泡、鳃弓、原肾、脊髓、脊索、头部间充质和体节中也观察到这两个基因的强转录本。对成年组织的Northern印迹分析表明，这两个基因在卵巢和睾丸中高表达，在其他器官中表达程度适中，只是在心脏、肺和肝脏中几乎检测不到Xdrg1的转录本。因此，Xdrg2和Xdrg1 mRNA的转录或稳定性可能受不同机制调控。此外，通过生成重组XDRG2和XDRG1蛋白，我们在体外发现了这些蛋白的RNA结合活性。我们的结果表明，DRG蛋白可能通过RNA结合发挥其生理作用。
