PDGF-RECEPTOR NEGATIVE CELLS

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The treatment of density-arrested BALB/c-3T3 cells with electrophoretically homogenous or highly purified preparations of the platelet-derived growth factor (PDGF) stimulated the rapid and selective accumulation of several species of abundant mRNA identified by cell-free translation. These translatable mRNAs appeared long before entry into the S phase. Less PDGF was required for selective mRNA accumulation than for PDGF-modulated DNA synthesis. The translatable mRNAs also accumulated after addition of the epidermal growth factor but not after addition of insulin or platelet-poor plasma. Their selective accumulation was blocked by addition of actinoimycin D. Three classes of PDGF-modulated mRNAs were defined. An early (primary) RNA appeared within 30 to 60 min of PDGF addition; its accumulation was not blocked by cycloheximide. Another early mRNA also appeared within 60 min, but treatment with both PDGF and cycloheximide was required for optimal accumulation. A third class, secondary RNAs, began to accumulate later at 90 to 120 min; the appearance of this class was inhibited by cycloheximide. One-\and two-dimensional gel electrophoresis of translation products demonstrated that a spontaneously transformed BALB/c-3T3 (ST2-3T3) cell line, which does not require PDGF or EGF for growth, constitutively accumulated the secondary growth factor-regulated mRNAs. The accumulation of these translatable mRNAs may be required for PDGF-modulated DNA synthesis. We have recently utilized a cDNA probe of a PDGF-regulated lysosomal protein (termed MEP) to directly quantify a PDGF-modulated mRNA. PDGF began to stimulate MEP mRNA accumulation 240 min after addition in a dose dependent fashion. Other growth factors, including EGF, IGF-1, insulin, and platelet-poor plasma did not have this effect. The PDGF modulated accumulation of MEP mRNA was inhibitable by cycloheximide demonstrating that PDGF modulated protein synthesis is required. A spontaneously transformed variant of BALB/c-3T3 cells which does not require PDGF for growth accumulated MEP mRNA in a constitutive fashion. Thus the MEP transcript is an example of a PDGF-modulated secondary RNA. (J)