Comparisons of Proteins Associated with Duck‐Globin mRNA and Its Polyadenylated Segment in Polyribosomal and Repressed Free Messenger Ribonucleoprotein Complexes

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Abstract
EDTA dissociation of polyribosomes from duck erythroblasts allowed us to isolate the 15‐S globin messenger ribonucleoproteins (mRNP) by sucrose gradient centrifugation or affinity chromatography on poly(U)‐Sepharose or oligo(dT)‐ce1lu1ose columns. Their protein composition was compared by one and two‐dimensional electrophoresis in sodium dodecyl sulfate to the free 20‐S mRNP containing the repressed fraction of globin mRNA [Vincent, A., Civelli, O., Maundrell, K., and Scherrer, K. (1980) Eur. J. Biochem. 112, 617–633]. The protein composition of the 15‐S mRNP isolated by these methods in different ionic strength conditions, was characterized by a major 73000‐Mr polypeptide and seven minor polypeptides with Mr ranging from 45000 to 68000, all of which are slightly basic, and about five acidic ones in the 80000–130000‐Mr range. All these are retained in the 15‐S mRNP core particle isolated at 0.5 M KCl. At low ionic strength, in addition, a specific group of acidic polypeptides in the Mr range 35000–105000 was also found associated with globin mRNA. Oligo(dT)‐cellulose chromatography of mRNP digested with ribonucleases A and T1 indicated that the 73000‐Mr major protein is bound to the poly(A) segment; some other proteins resolved as minor components interact with both the poly(A) and non‐poly(A) regions of globin mRNA.Characterization of proteins interacting with the poly(A) segment of non‐polyribosomal globin mRNA in 20‐S free mRNP demonstrated the absence of the polyribosomal 73000‐Mr poly(A)‐binding protein. Furthermore, it confirmed that the protein compositions of translatable polyribosomal and repressed free globin mRNP are very different. Indeed, the respective core (0.5 M KCl) particles contain only two possibly common polypeptides. The specificity of proteins associated with globin mRNA in two different functional states shown here supports the hypothesis of a role of mRNP proteins in translational control of mRNA.