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Molecular and Cellular Biology, July 2000, p. 4932-4947, Vol. 20, No. 13
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Amino- and Carboxy-Terminal PEST Domains Mediate Gastrin Stabilization of Rat L-Histidine Decarboxylase Isoforms

John V. Fleming and Timothy C. Wang*

Department of Medicine, Harvard Medical School, and Gastrointestinal Unit, Massachusetts General Hospital, Boston, Massachusetts 02114

Received 12 November 1999/Returned for modification 17 January 2000/Accepted 17 March 2000

Control of enzymatic function by peptide hormones can occur at a number of different levels and can involve diverse pathways that regulate cleavage, intracellular trafficking, and protein degradation. Gastrin is a peptide hormone that binds to the cholecystokinin B-gastrin receptor and regulates the activity of L-histidine decarboxylase (HDC), the enzyme that produces histamine. Here we show that gastrin can increase the steady-state levels of at least six HDC isoforms without affecting HDC mRNA levels. Pulse-chase experiments indicated that HDC isoforms are rapidly degraded and that gastrin-dependent increases are due to enhanced isoform stability. Deletion analysis identified two PEST domains (PEST1 and PEST2) and an intracellular targeting domain (ER2) which regulate HDC protein expression levels. Experiments with PEST domain fusion proteins demonstrated that PEST1 and PEST2 are strong and portable degradation-promoting elements which are positively regulated by both gastrin stimulation and proteasome inhibition. A chimeric protein containing the PEST domain of ornithine decarboxylase was similarly affected, indicating that gastrin can regulate the stability of other PEST domain-containing proteins and does so independently of antizyme/antizyme inhibitor regulation. At the same time, endoplasmic reticulum localization of a fluorescent chimera containing the ER2 domain of HDC was unaltered by gastrin stimulation. We conclude that gastrin stabilization of HDC isoforms is dependent upon two transferable and sequentially unrelated PEST domains that regulate degradation. These experiments revealed a novel regulatory mechanism by which a peptide hormone such as gastrin can disrupt the degradation function of multiple PEST-domain-containing proteins.

* Corresponding author. Mailing address: Gastrointestinal Unit, Massachusetts General Hospital, 32 Fruit Street, Boston, MA 02114. Phone: (617) 726-9228. Fax: (617) 726-3673. E-mail: wang{at}helix.mgh.harvard.edu.

Molecular and Cellular Biology, July 2000, p. 4932-4947, Vol. 20, No. 13
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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