ARFAPTIN-1, A PUTATIVE CYTOSOLIC TARGET PROTEIN OF ADP-RIBOSYLATION FACTOR, IS RECRUITED TO GOLGI MEMBRANES

ADP-ribosylation factors (ARFs) have been implicated in vesicle transp ort in the Golgi complex. Employing yeast two-hybrid screening of an H L60 cDNA library using a constitutively active mutant of ARF3 (ARF3 . Q71L), 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 arfa ptin 1. The mRNA of arfaptin 1 was ubiquitously expressed, and recombi nant 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 me mbranes 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 yielde d another clone, which encoded a putative protein, which we have named arfaptin 2. This consisted of the same number of amino acids as arfap tin 1 and was 60% identical to it, Arfaptin 2 was also ubiquitously ex pressed and bound to the GTP-, but not GDP-liganded form of class I AR Fs, 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.