Chardin P et al. (1996) Nature "A human exchange factor for ARF contains Sec7- and pleckstrin-homology domains."

Chardin P, Chabre M, Antonny B, Beraud-Dufour S, Jackson CL, Paris S, Robineau S

[Nature, 1996]

The small G protein ARF1 is involved in the coating of vesicles that bud from the Golgi compartments. Its activation is controlled by as-yet unidentified guanine-nucleotide exchange factors. Gea1, the first ARF exchange factor to be discovered in yeast, is a large protein containing a domain of homology with Sec7, another yeast protein that is also involved in secretion. Here we characterized a smaller human protein (relative molecular mass 47K) named ARNO, which contains a central Sec7 domain that promotes guanine-nucleotide exchange on ARF1. ARNO also contains an amino-terminal coiled-coil motif and a carboxy-terminal pleckstrin-homology (PH) domain. The PH domain mediates an enhancement of ARNO exchange activity by negatively charged phospholipid vesicles supplemented with phosphatidylinositol bisphosphate. The exchange activity of ARNO is not inhibited by brefeldin A, an agent known to block vesicular transport and inhibit the exchange activity on ARF1 in cell extracts. This suggests that a regulatory component which is sensitive to brefeldin A associates with ARNO in vivo, possibly through the amino-terminal coiled-coil. We propose that other proteins with a Sec7 domain regulate different members of the ARF family.

Roelens B et al. (2019) MicroPubl Biol

Roelens B, Zawadzki KA, Villeneuve AM

[MicroPubl Biol, 2019]

The me101 allele was isolated in a genetic screen for mutants with an altered number of GFP::COSA-1 foci, which mark the sites of crossovers in C. elegans germ cells (Rosu et al. 2013). After multiple rounds of outcrossing, we confirmed that me101 mutants were defective in some aspects of meiotic prophase, as late pachytene me101 mutant meiocytes failed to form the six GFP::COSA-1 foci observed in wild-type late pachytene meiocytes (Fig 1.A). We also observed structural defects ranging from chromosome fragmentation to the formation of chromosome aggregates in me101 diakinesis-stage oocytes (Fig. 1B), suggesting a defect in some aspect of the DNA damage response. Further, 100% of eggs laid by me101 mutant hermaphrodites are inviable. We then assessed the localization of the recombinase RAD-51, an essential component of the homologous recombination machinery that is required for the repair of DNA breaks and the maintenance of genome integrity during meiosis (Rinaldo et al. 2002; Alpi et al. 2003); no RAD-51 foci were observed in the gonads of me101 mutants (Fig 1C). Sequencing of the rad-51 locus in the me101 mutant revealed a single G to A substitution (IV:10283785 from WS269), leading to an Arginine to Histidine substitution in a conserved residue (Fig. 1D). Failure to detect RAD-51 foci in the me101 mutant indicates that this residue is important for the loading and/or the stability of the RAD-51 protein.