Establishment of an assay system for the detection of translated materials of T-cell-replacing factor mRNA in Xenopus oocytes.
We established an assay system for detecting T cell-replacing factor (TRF) activity of translated materials in Xenopus oocytes of poly (A)-positive mRNA extracted from a T cell hybrid cell line, B151K12 (B151) which constitutively produces TRF. Since it was difficult to detect TRF activity of the translated products of B151-mRNA, partly because of low TRF activities, we developed the following two systems. First, RNA was prepared from B151 cells stimulated with phorbol myristate acetate and calcium ionophore A23187 because such stimulations augmented TRF production by approximately three to five-fold. Second, interleukin 2 (IL-2, 125 U/ml) was added to the culture of BCL1 cells to detect a small amount of TRF-active materials since IL-2 synergizes with a suboptimal dose of TRF to induce IgM secretion in TRF-responding BCL1 cells (chronic B cell leukemic cells). Here we describe TRF activity of translation products of B151-mRNA in Xenopus oocytes. B151-TRF mRNA was detected in the fractions sedimented between 15 and 18S by analysis using sucrose density gradient centrifugation.
D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D004357:Drug Synergism; D005260:Female; D007376:Interleukin-2 / Q000494:pharmacology; D015848:Interleukin-5; D008222:Lymphokines / Q000032:analysis* / Q000096:biosynthesis; D009865:Oocytes; D011061:Poly A / Q000378:metabolism; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014982:Xenopus laevis
D007376:Interleukin-2; D015848:Interleukin-5; D008222:Lymphokines; D012333:RNA, Messenger; D011061:Poly A; D000001:Calcimycin; D013755:Tetradecanoylphorbol Acetate