Arfaptin 1, a Putative Cytosolic Target Protein of ADP-ribosylation Factor, Is Recruited to Golgi Membranes

Abstract

ADP-ribosylation factors (ARFs) have been implicated in vesicle transport in the Golgi complex. Employing yeast two-hybrid screening of an HL60 cDNA library using a constitutively active mutant of ARF3 (ARF3⅐Q71L), as a probe, we have identified a cDNA encoding a novel protein with a calculated molecular mass of 38.6 kDa, which we have named arfaptin 1. The mRNA of arfaptin 1 was ubiquitously expressed, and recombinant arfaptin 1 bound preferentially to class I ARFs, especially ARF1, but only in the GTP-bound form. The interactions were independent of myristoylation of ARF. Arfaptin 1 in cytosol was recruited to Golgi membranes by ARF in a guanosine 5 -O-(3-thiotriphosphate)-dependent and brefeldin A-sensitive manner. When expressed in COS cells, arfaptin 1 was localized to the Golgi complex. The yeast two-hybrid system yielded another clone, which encoded a putative protein, which we have named arfaptin 2. This consisted of the same number of amino acids as arfaptin 1 and was 60% identical to it. Arfaptin 2 was also ubiquitously expressed and bound to the GTP-, but not GDP-liganded form of class I ARFs, especially ARF1. These results suggest that arfaptins 1 and 2 may be direct target proteins of class 1 ARFs. Arfaptin 1 may be involved in Golgi function along with ARF1.

Activation of exocytosis by cross-linking of the IgE receptor is dependent on ADP-ribosylation factor 1-regulated phospholipase D in RBL-2H3 mast cells: evidence that the mechanism of activation is via regulation of phosphatidylinositol 4,5-bisphosphate synthesis

“…ARF1 can interact directly with several effectors, including PLD1, ARFAPTIN and the β-subunit of coatomer proteins [15,36,37]. In the present study of regulated exocytosis, the data support the possibility that part of the function of ARF1 is mediated via PLD.…”

A Family of ADP-Ribosylation Factor Effectors That Can Alter Membrane Transport through thetrans-Golgi

“…Eleven of these positive clones fell into one group and have been described in a separate publication (Boman et al, 1999). A single clone was identified as POR1/Arfaptin2 (Van Aelst et al, 1996;Kanoh et al, 1997). Of the remaining positive clones, four encode the first two members of a new protein family, which are described in more detail below.…”

Islet Cell Autoantigen of 69 kDa Is an Arfaptin-related Protein Associated with the Golgi Complex of Insulinoma INS-1 Cells

“…Back PSI-BLAST searches with human arfaptin 1 identified the ICA69 family with an expect value of 9e-73 at the third iteration. Arfaptin 1 and 2 are cytosolic proteins that bind small GTPases of the ARF family (8) and Rac1 (7). Arfaptin 1, in particular, is recruited to Golgi membranes by ARF1-GTP and acts as negative regulator of ARF1 function (8,58,59).…”

“…Arfaptin 1 and 2 are cytosolic proteins that bind small GTPases of the ARF family (8) and Rac1 (7). Arfaptin 1, in particular, is recruited to Golgi membranes by ARF1-GTP and acts as negative regulator of ARF1 function (8,58,59). Arfaptin is composed of three ␣-helices that dimerize to form the binding interface for small GTPases.…”

“…ICA69 is similar to arfaptins, which act as effectors of the small GTPases ARF and Rac (7,8). Thus, ICA69 is likely to play a role in vesicular transport regulated by small GTP-binding proteins at the Golgi complex and ISGs in ␤-cells and secretory cells in general.…”