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Molecular Endocrinology, Vol 8, 603-613, Copyright © 1994 by Endocrine Society

ARTICLES

Alternatively polyadenylated vasoactive intestinal peptide mRNAs are differentially regulated at the level of stability

LJ Chew, D Murphy and DA Carter
Neuropeptide Laboratory, National University of Singapore.

The role of cis-acting destabilizing RNA sequences in the determination of endocrine gene expression has been investigated using a novel paradigm, in which the differential regulation of two alternatively polyadenylated RNA transcripts may be observed both in vivo and in vitro. In the rat anterior pituitary gland in vivo, we have shown that, after the termination of an estrogen stimulus, a 1.7-kilobase (kb) vasoactive intestinal peptide (VIP) RNA containing an extensive 3'- untranslated region (UTR), is preferentially down-regulated with respect to a 1.0 kb VIP transcript that is uniquely abundant in this tissue. Differential regulation of the anterior pituitary VIP transcripts can be modeled in an explant culture system in which we defined both transcriptional and posttranscriptional phases of VIP gene regulation in vitro, and showed that selective down-regulation of the 1.7-kb transcript is posttranscriptional. Inhibitors of transcription and translation have also allowed us to show in vitro that differential regulation of VIP transcripts occurs through an active process that appears to involve the synthesis of a labile, destabilizing factor. In order to confirm the role of RNA destabilization as the primary mechanism of differential posttranscriptional regulation, we have also performed cell-free stability assays in which explant extracts were incubated with 32P-labeled run-off transcripts corresponding to the two alternatively polyadenylated VIP RNAs. The resultant estimates of RNA half-life showed significantly lower values for the synthetic VIP transcript containing the 3'-UTR. Our findings demonstrate the presence of functional destabilizing sequences in the 3'-UTR of the rat VIP RNA which appear to act in the physiological control of VIP gene expression.


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