Regulation of poly(A) site use during mouse B-cell development involves a change in the binding of a general polyadenylation factor in a B-cell stage-specific manner.
During the development of mouse B cells there is a regulated shift from the production of membrane to the secretion-specific forms of immunoglobulin (Ig) mRNA, which predominate in the late-stage or plasma B cells. By DNA transfection experiments we have previously shown that there is an increase in polyadenylation efficiency accompanying the shift to secretion-specific forms of Ig mRNA (C. R. Lassman, S. Matis, B. L. Hall, D. L. Toppmeyer, and C. Milcarek, J. Immunol. 148:1251-1260, 1992). When we look in vitro at nuclear extracts prepared from early or memory versus late-stage or plasma B cells, we see cell stage-specific differences in the proteins which are UV cross-linked to the input RNAs. We have characterized one of these proteins as the 64-kDa subunit of the general polyadenylation factor cleavage-stimulatory factor (CstF) by immunoprecipitation of UV-cross-linked material. The amount of 64-kDa protein and its mobility on two-dimensional gels do not vary between the B-cell stages. However, the activity of binding of the protein to both Ig and non-Ig substrates increases four- to eightfold in the late-stage or plasma cell lines relative to the binding seen in the early or memory B-cell lines. Therefore, the binding activity of a constitutive factor required for polyadenylation is altered in a B-cell-specific fashion. The increased binding of the 64-kDa protein may lead to a generalized increase in polyadenylation efficiency in plasma cells versus early or memory B cells which may be responsible for the increased use of the secretory poly(A) site seen in vivo.
Edwalds-Gilbert
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Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pennsylvania 15261-2072, USA.
Milcarek
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C
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000502:physiology*; D002478:Cells, Cultured; D015180:Electrophoresis, Gel, Two-Dimensional; D018507:Gene Expression Regulation, Developmental; D005803:Genes, Immunoglobulin; D006825:Hybridomas; D007074:Immunoglobulin G / Q000235:genetics; D051379:Mice; D011061:Poly A / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D039221:mRNA Cleavage and Polyadenylation Factors
D007074:Immunoglobulin G; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D039221:mRNA Cleavage and Polyadenylation Factors; D011061:Poly A