Wheeler E F, Roussel M F, Hampe A, Walker M H, Fried V A, Look A T, Rettenmier C W, Sherr C J
J Virol. 1986 Aug;59(2):224-33. doi: 10.1128/JVI.59.2.224-233.1986.
The nucleotide sequence of a 5' segment of the human genomic c-fms proto-oncogene suggested that recombination between feline leukemia virus and feline c-fms sequences might have occurred in a region encoding the 5' untranslated portion of c-fms mRNA. The polyprotein precursor gP180gag-fms encoded by the McDonough strain of feline sarcoma virus was therefore predicted to contain 34 v-fms-coded amino acids derived from sequences of the c-fms gene that are not ordinarily translated from the proto-oncogene mRNA. The (gP180gag-fms) polyprotein was cotranslationally cleaved near the gag-fms junction to remove its gag gene-coded portion. Determination of the amino-terminal sequence of the resulting v-fms-coded glycoprotein, gp120v-fms, showed that the site of proteolysis corresponded to a predicted signal peptidase cleavage site within the c-fms gene product. Together, these analyses suggested that the linked gag sequences may not be necessary for expression of a biologically active v-fms gene product. The gag-fms sequences of feline sarcoma virus strain McDonough and the v-fms sequences alone were inserted into a murine retroviral vector containing a neomycin resistance gene. Both constructs were biologically active when transfected into NIH 3T3 cells and produced morphologically transformed foci at equivalent efficiencies. When transfected into a cell line (psi 2) expressing complementary viral gene functions, G418-resistant (Neor) cells containing either of these vector DNAs produced high titers of transforming viruses. Analysis of proteins produced in cells containing the vector lacking gag gene sequences showed that gP180gag-fms was not synthesized, whereas normal levels of both immature gp120v-fms and mature gp140v-fms were detected. The glycoprotein was efficiently transported to the cell surface, and it retained wild-type tyrosine kinase activity. We conclude that a cryptic hydrophobic signal peptide sequence in v-fms was unmasked by gag deletion, thereby allowing the correct orientation and transport of the v-fms gene product within membranous organelles. It seems likely that the proteolytic cleavage of gP180gag-fms is mediated by signal peptidase and that the amino termini of gp140v-fms and the c-fms gene product are identical.
人类基因组原癌基因c-fms 5'端的核苷酸序列表明，猫白血病病毒与猫c-fms序列之间的重组可能发生在编码c-fms mRNA 5'非翻译区的区域。因此，预计由猫肉瘤病毒麦克多诺株编码的多蛋白前体gP180gag-fms含有34个由c-fms基因序列编码的v-fms氨基酸，而这些序列通常不会从原癌基因mRNA翻译而来。(gP180gag-fms)多蛋白在gag-fms连接处附近进行共翻译切割，以去除其gag基因编码部分。对所得v-fms编码的糖蛋白gp120v-fms的氨基末端序列的测定表明，蛋白水解位点对应于c-fms基因产物中预测的信号肽酶切割位点。这些分析共同表明，连接的gag序列对于生物活性v-fms基因产物的表达可能不是必需的。将猫肉瘤病毒株麦克多诺的gag-fms序列和单独的v-fms序列插入到含有新霉素抗性基因的鼠逆转录病毒载体中。当转染到NIH 3T3细胞中时，这两种构建体都具有生物活性，并以相同的效率产生形态转化灶。当转染到表达互补病毒基因功能的细胞系(psi 2)中时，含有这两种载体DNA之一的G418抗性(Neor)细胞产生高滴度的转化病毒。对含有缺乏gag基因序列的载体的细胞中产生的蛋白质的分析表明，没有合成gP180gag-fms，而检测到正常水平的未成熟gp120v-fms和成熟gp140v-fms。该糖蛋白被有效地转运到细胞表面，并保留野生型酪氨酸激酶活性。我们得出结论，v-fms中一个隐蔽的疏水信号肽序列因gag缺失而暴露，从而使v-fms基因产物在膜性细胞器内能够正确定向和转运。gP180gag-fms的蛋白水解切割似乎是由信号肽酶介导的，并且gp140v-fms和c-fms基因产物的氨基末端是相同的。
