Porcu P, Ferber A, Pietrzkowski Z, Roberts C T, Adamo M, LeRoith D, Baserga R
Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Mol Cell Biol. 1992 Nov;12(11):5069-77. doi: 10.1128/mcb.12.11.5069-5077.1992.
We have used a plasmid expressing a temperature-sensitive (ts) mutant of simian virus 40 (SV40) T antigen, stably transfected into 3T3 cells, to study the role of insulinlike growth factor 1 (IGF-1) and its receptor in T-antigen-mediated growth. While 3T3 cells do not grow in serum-free medium, in 1% serum, or with the sole addition of either platelet-derived growth factor (PDGF) or IGF-1, cells expressing the tsA T antigen (BALB 58 cells) grow at 34 degrees C in either PDGF or 1% serum but not in IGF-1. At the restrictive temperature (39.6 degrees C), these cells can only grow in 10% serum. We show that BALB 58 cells, at 34 degrees C, have a markedly increased expression of IGF-1 and IGF-1 mRNA and that their growth in 1% serum (at 34 degrees C) is inhibited by an antisense oligodeoxynucleotide to the IGF-1 receptor RNA. When this tsA plasmid is stably transfected into cells constitutively overexpressing the human IGF-1 receptor cDNA, the resulting cell lines show a constitutively phosphorylated IGF-1 receptor and grow in serum-free medium at 34 degrees C (but not at 39.6 degrees C). A functional SV40 T antigen also increases the expression of a plasmid in which the reporter luciferase gene is under the control of a rat IGF-1 promoter. We conclude (i) that the SV40 T antigen induces the expression of IGF-1 and IGF-1 mRNA, at least in part by a transcriptional mechanism, thus altering the growth factors requirements, and (ii) that, in BALB/c3t3 cells, the SV40 T antigen necessitates a functional IGF-1 receptor for its growth-stimulating effect in low serum (or PDGF).
我们使用了一种表达猿猴病毒40（SV40）T抗原温度敏感（ts）突变体的质粒，将其稳定转染到3T3细胞中，以研究胰岛素样生长因子1（IGF-1）及其受体在T抗原介导的生长中的作用。虽然3T3细胞在无血清培养基、1%血清中，或仅添加血小板衍生生长因子（PDGF）或IGF-1时都不生长，但表达tsA T抗原的细胞（BALB 58细胞）在34℃下，在PDGF或1%血清中能生长，但在IGF-1中不能生长。在限制温度（39.6℃）下，这些细胞只能在10%血清中生长。我们发现，BALB 58细胞在34℃时，IGF-1和IGF-1 mRNA的表达明显增加，并且它们在1%血清中（34℃）的生长受到针对IGF-1受体RNA的反义寡脱氧核苷酸的抑制。当这种tsA质粒稳定转染到组成性过表达人IGF-1受体cDNA的细胞中时，所得细胞系显示出组成性磷酸化的IGF-1受体，并在34℃的无血清培养基中生长（但在39.6℃下不生长）。功能性SV40 T抗原还增加了一种质粒的表达，其中报告荧光素酶基因受大鼠IGF-1启动子的控制。我们得出结论：（i）SV40 T抗原至少部分通过转录机制诱导IGF-1和IGF-1 mRNA的表达，从而改变生长因子需求；（ii）在BALB/c3t3细胞中，SV40 T抗原在低血清（或PDGF）中发挥生长刺激作用需要功能性IGF-1受体。
