Swanson M E, Elste A M, Greenberg S M, Schwartz J H, Aldrich T H, Furth M E
J Cell Biol. 1986 Aug;103(2):485-92. doi: 10.1083/jcb.103.2.485.
We have cloned a DNA fragment from the marine mollusc Aplysia californica, which contains sequences homologous to mammalian ras genes, by screening a genomic library with a viral Ha-ras oncogene probe under conditions of low stringency hybridization. Nucleotide sequencing revealed a putative exon that encodes amino acids sharing 68% homology with residues 5 to 54 of mammalian p21ras polypeptides, and which therefore is likely to encode a ras-like Aplysia protein. The cloned locus, designated Apl-ras, is distinct from the Aplysia rho (ras-homologue) gene and appears to be more closely related to mammalian ras. We used a panel of monoclonal antibodies raised against v-Ha-ras p21 to precipitate an Mr 21,000 protein from extracts of Aplysia nervous tissue, ovotestis, and, to a much lesser degree, buccal muscle. Fluorescence immunocytochemistry revealed that ras-like protein is most abundant in neuronal cell bodies and axon processes, with staining most prominent at plasma membranes. Much less was present in other tissues. The prominence of ras protein in neurons, which are terminally differentiated and non-proliferating, indicates that the control of cell division is not the sole function of this proto-oncogene. The large identified neurons of Aplysia offer the opportunity to examine how ras protein might function in mature nerve cells.
我们通过在低严格度杂交条件下用病毒Ha-ras癌基因探针筛选基因组文库,从海洋软体动物加州海兔中克隆了一个DNA片段,该片段含有与哺乳动物ras基因同源的序列。核苷酸测序揭示了一个推定的外显子,其编码的氨基酸与哺乳动物p21ras多肽的第5至54位残基具有68%的同源性,因此可能编码一种类似ras的海兔蛋白。克隆的基因座命名为Apl-ras,与海兔rho(ras同源物)基因不同,似乎与哺乳动物ras的关系更为密切。我们使用一组针对v-Ha-ras p21产生的单克隆抗体,从海兔神经组织、卵睾以及程度较轻的颊肌提取物中沉淀出一种分子量为21,000的蛋白质。荧光免疫细胞化学显示,类似ras的蛋白在神经元细胞体和轴突中最为丰富,在质膜处染色最为明显。在其他组织中含量则少得多。ras蛋白在终末分化且不增殖的神经元中的突出存在表明,细胞分裂的控制并非该原癌基因的唯一功能。海兔中已鉴定的大型神经元为研究ras蛋白在成熟神经细胞中的功能提供了机会。