ADP-ribosylation Factor-1 Stimulates Formation of Nascent Secretory Vesicles from the trans-Golgi Network of Endocrine Cells

doi:10.1074/jbc.271.10.5297

ADP-ribosylation Factor-1 Stimulates Formation of Nascent Secretory Vesicles from the trans-Golgi Network of Endocrine Cells (*)

From the (1)Departments of Developmental and Molecular Biology and
(2)Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461

^¶ To whom correspondence should be addressed:
Depts. of Developmental and Molecular Biology and Anatomy and Structural Biology, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10461
. Tel.: 718-430-3306; Fax: 718-430-8567.

Abstract

ADP-ribosylation factor (ARF) is a small GTP-binding protein that has been implicated in intracellular vesicular transport. ARF regulates the budding of vesicles that mediate endoplasmic reticulum to Golgi and intra-Golgi transport. It also plays an important role in maintaining the function and morphology of the Golgi apparatus. Using a permeabilized cell system derived from GH cells, we provide evidence that ARF-1 regulates the formation of nascent secretory vesicles from the trans-Golgi network. Both myristoylated and non-myristoylated forms of recombinant human ARF-1 enhanced secretory vesicle budding about 2-fold. A mutant lacking the first 17 N-terminal residues, as well as one that preferentially binds GDP (T31N) did not stimulate vesicle formation. In contrast, a mutant defective in GTP hydrolysis (Q71L) promoted vesicle budding. Strikingly, a peptide corresponding to the N terminus of human ARF-1 (amino acids 2-17) also stimulated vesicle budding from the trans-Golgi network, in marked contrast to its inhibitory effect on vesicular transport from the endoplasmic reticulum to Golgi. These data demonstrate that in endocrine cells, ARF-1 and in particular its N terminus play an essential role in the formation of secretory vesicles.

Footnotes

↵^§ Supported in part by National Institutes of Health Training Grant CA09475.
↵* This work was supported by National Institutes of Health Grant DK21860 and by a grant from the Juvenile Diabetes Foundation (to D. S.). Core support was provided by a National Institutes of Health Cancer Center Grant P30CA13330. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

ARF

ADP-ribosylation factor

ER

endoplasmic reticulum

TGN

trans-Golgi network

GH

growth hormone

PRL

prolactin

PAGE

polyacrylamide gel electrophoresis

PCR

polymerase chain reaction.
- Received December 29, 1995.