Regulation of tissue-specific splicing of the calcitonin/calcitonin gene-related peptide gene by RNA-binding proteins.
Transcripts of the rat calcitonin/calcitonin gene-related peptide (CGRP) gene are alternatively spliced in a tissue-specific manner resulting in the production of calcitonin mRNA and peptide in thyroid C cells and CGRP mRNA and peptide in neurons. Transfection studies using calcitonin and chimaeric human beta-globin/calcitonin exon minigene constructs showed that the splice acceptor and exon specific to calcitonin mRNA are spliced much less efficiently in CGRP-producing cells (F9 teratocarcinomas) than in cells that preferentially make calcitonin (HeLa cells). In vitro splicing of chimaeric human beta-globin/calcitonin transcripts in HeLa nuclear extracts were inhibited by the addition of nuclear extract from CGRP-favoring cells or tissues such as rat brain. This inhibition was specific as splicing of human beta-globin first intron transcripts was not affected by comparable amounts of rat brain extract. Fractionation of rat brain nuclear extracts allowed the partial purification of two brain-specific polypeptides of apparent molecular mass of 43 and 41 kDa which preferentially bind RNA containing the calcitonin-specific splice acceptor. Since these polypeptides cofractionate with the calcitonin mRNA-specific splicing inhibition activity, we suggest that they may mediate the inhibition of splicing observed in vitro and underlie, in part, the inefficient calcitonin mRNA production observed in CGRP-favoring cells in vivo.
Roesser
J R
JR
Department of Pharmacology, Stanford University, California 94305.
Liittschwager
K
K
Leff
S E
SE
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000378:metabolism; D002116:Calcitonin / Q000235:genetics*; D015740:Calcitonin Gene-Related Peptide / Q000235:genetics*; D005786:Gene Expression Regulation; D005914:Globins / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015183:Restriction Mapping; D013724:Teratoma; D013961:Thyroid Gland / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D005914:Globins; D002116:Calcitonin; D015740:Calcitonin Gene-Related Peptide