Regulation of mRNA abundance in activated T lymphocytes: identification of mRNA species affected by the inhibition of protein synthesis.
Inhibition of protein synthesis has often been observed to increase the concentration of mRNAs that encode proteins associated with the regulation of cell division. As two-dimensional gel electrophoresis permits the simultaneous monitoring of individual elements in large populations of gene products, we have used this technique to assess the effect of cycloheximide treatment on the mRNA complement of activated mouse T cells in an objective fashion. Two-dimensional gels of proteins generated by cell-free translation of mRNA from T-cell blasts display about 400 spots; only 5 of these are reproducibly enhanced by cycloheximide treatment and about 4 are diminished. The cDNA cloning vector lambda jac allows analysis of large arrays of molecular clones by cell-free expression, and we have used it in a sibling selection scheme to isolate a clone of one of the prominently induced mRNA species, which we refer to as chx1. chx1 mRNA concentration is increased by cycloheximide treatment of activated B cells, as well as T cells, and it is rapidly and transiently induced, in a cycloheximide-enhanced manner, upon serum stimulation of resting 3T3 fibroblastoid cells. The chx1 protein is hydrophilic, is slightly basic, and has patches of homology with the Jun-D gene product. The chx1 gene is remarkable in its lack of detectable introns and of strong bias against CpG dinucleotides.
Coleclough
C
C
Department of Immunology, Saint Jude Children's Research Hospital, Memphis, TN 38101.
Kuhn
L
L
Lefkovits
I
I
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003513:Cycloheximide / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D007438:Introns; D008213:Lymphocyte Activation; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D013154:Spleen / Q000276:immunology; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*
D012333:RNA, Messenger; D003513:Cycloheximide