Stiles C D, Pledger W J, Tucker R W, Martin R G, Scher C D
J Supramol Struct. 1980;13(4):489-99. doi: 10.1002/jss.400130408.
The platelet-derived growth factor (PDGF), which is found in serum but not in plasma, has been purified to homogeneity; it stimulates replication at a concentration of 10(-10) M. Brief treatment with PDGF causes density-inhibited Balb/c-3T3 cells to become competent to synthesize DNA; pituitary fibroblast growth factor (FGF) or precipitates of calcium phosphate also induce competence. Continuous treatment with plasma allows competent, but not incompetent, cells to synthesize DNA. A critical component of plasma is somatomedin, a group of hormones with insulin-like activity; multiplication-stimulating activity (MSA) or insulin replace plasma somatomedin in promoting DNA synthesis. We have studied the molecular correlates of competence and the role of SV40 gene A products in regulating DNA synthesis. Treatment of quiescent cells with pure PDGF or FGF causes the preferential synthesis of five cytoplasmic proteins (approximate molecular weight 29,000, 35,000, 45,000, 60,000, and 72,000 detected by SDS-PAGE under reducing conditions). Two of these competence-associated proteins (29,000 and 35,000 daltons) are insulin, or epidermal growth factor (EGF). PDGF, FGF, or calcium phosphate induce an ultrastructure change within the centriole of 3T3 cells; this ultrastructural modification of the centriole is detectable by immunofluorescence within 2 h or PDGF treatment. Plasma, EGF, or MSA do not modify the centriole. SV40 induces replicative DNA synthesis in growth-arrested 3T3 cells but does not cause this alteration in centriole structure. Gene A variants of SV40, including a mutant with temperature-sensitive (ts) T-antigen (ts A209), a deletion in t-antigen (dl 884), and several ts A209 strains containing t-antigen deletions were used to induce DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells. Like wild type SV40, all strains induced DNA synthesis equally well under permissive or nonpermissive conditions. Addition of PDGF or plasma had little effect on SV40-induced DNA synthesis. Thus, the viral function that induces replicative DNA synthesis in Balb/c-3T3 cells is not t and is not temperature sensitive. This SV40 gene function overrides the cellular requirement for hormonal growth factors. It does not induce transient centriole deciliation, a hormonally regulated event.
血小板衍生生长因子（PDGF）存在于血清而非血浆中，已被纯化至同质；它在浓度为10⁻¹⁰ M时刺激细胞复制。用PDGF短暂处理会使密度抑制的Balb/c - 3T3细胞有能力合成DNA；垂体成纤维细胞生长因子（FGF）或磷酸钙沉淀也能诱导细胞获得这种能力。用血浆持续处理可使有能力但无此能力的细胞合成DNA。血浆的一个关键成分是生长调节素，这是一组具有胰岛素样活性的激素；增殖刺激活性（MSA）或胰岛素在促进DNA合成方面可替代血浆生长调节素。我们研究了细胞获得能力的分子关联以及SV40基因A产物在调节DNA合成中的作用。用纯PDGF或FGF处理静止细胞会导致优先合成五种细胞质蛋白（在还原条件下通过SDS - PAGE检测，分子量约为29,000、35,000、45,000、60,000和72,000）。其中两种与获得能力相关的蛋白（29,000和35,000道尔顿）不是胰岛素，也不是表皮生长因子（EGF）。PDGF、FGF或磷酸钙会诱导3T3细胞中心粒内的超微结构变化；在PDGF处理后2小时内通过免疫荧光可检测到这种中心粒的超微结构改变。血浆、EGF或MSA不会改变中心粒。SV40在生长停滞的3T3细胞中诱导复制性DNA合成，但不会引起中心粒结构的这种改变。SV40的基因A变体，包括具有温度敏感（ts）T抗原的突变体（ts A209）、T抗原缺失（dl 884）以及几种含有T抗原缺失的ts A209菌株，被用于在Balb/c - 3T3细胞中诱导DNA合成。与野生型SV40一样，所有菌株在允许或非允许条件下诱导DNA合成的效果相同。添加PDGF或血浆对SV40诱导的DNA合成影响很小。因此，在Balb/c - 3T3细胞中诱导复制性DNA合成的病毒功能不是T抗原，也不是温度敏感的。这种SV40基因功能超越了细胞对激素生长因子的需求。它不会诱导短暂的中心粒去纤毛，这是一个受激素调节的事件。
