Limits...
The GTPase Arf1p and the ER to Golgi cargo receptor Erv14p cooperate to recruit the golgin Rud3p to the cis-Golgi.

Gillingham AK, Tong AH, Boone C, Munro S - J. Cell Biol. (2004)

Bottom Line: Bornens. 2004.Cell. 118:323-335).In contrast, we find that this region binds to the Golgi in a GRAB domain-dependent manner, suggesting that GMAP-210 may not link the Golgi to gamma-tubulin and centrosomes.

View Article: PubMed Central - PubMed

Affiliation: Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 2QH, England, UK.

ABSTRACT
Rud3p is a coiled-coil protein of the yeast cis-Golgi. We find that Rud3p is localized to the Golgi via a COOH-terminal domain that is distantly related to the GRIP domain that recruits several coiled-coil proteins to the trans-Golgi by binding the small Arf-like GTPase Arl1p. In contrast, Rud3p binds to the GTPase Arf1p via this COOH-terminal "GRIP-related Arf-binding" (GRAB) domain. Deletion of RUD3 is lethal in the absence of the Golgi GTPase Ypt6p, and a screen of other mutants showing a similar genetic interaction revealed that Golgi targeting of Rud3p also requires Erv14p, a cargo receptor that cycles between the endoplasmic reticulum and Golgi. The one human protein with a GRAB domain, GMAP-210 (CEV14/Trip11/Trip230), is known to be on the cis-Golgi, but the COOH-terminal region that contains the GRAB domain has been reported to bind to centrosomes and gamma-tubulin (Rios, R.M, A. Sanchis, A.M. Tassin, C. Fedriani, and M. Bornens. 2004. Cell. 118:323-335). In contrast, we find that this region binds to the Golgi in a GRAB domain-dependent manner, suggesting that GMAP-210 may not link the Golgi to gamma-tubulin and centrosomes.

Show MeSH

Related in: MedlinePlus

Erv14p is required for the Golgi localization of Rud3p. (A) Fluorescent micrographs of live yeast (BY4741) expressing GFP-Rud3p from plasmid pE1, with the indicated genes deleted. (B) Fluorescent micrographs of an erv14Δ strain with the RUD3 ORF NH2-terminally GFP tagged in the genome and containing either an empty CEN plasmid or a similar plasmid encoding Erv14p-HA as indicated. (C) Fluorescent micrographs of a yeast strain lacking ERV14 and with USO1 COOH-terminally GFP tagged in the genome and containing a CEN plasmid expressing RFP-Rud3p as in Fig. 1 A. (D) Distribution of the Golgi markers Gos1p and Vrg4p expressed as GFP-fusions in either wild-type cells or a mutant lacking ERV14 as indicated. GFP-tagged proteins expressed as in A.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2172552&req=5

fig5: Erv14p is required for the Golgi localization of Rud3p. (A) Fluorescent micrographs of live yeast (BY4741) expressing GFP-Rud3p from plasmid pE1, with the indicated genes deleted. (B) Fluorescent micrographs of an erv14Δ strain with the RUD3 ORF NH2-terminally GFP tagged in the genome and containing either an empty CEN plasmid or a similar plasmid encoding Erv14p-HA as indicated. (C) Fluorescent micrographs of a yeast strain lacking ERV14 and with USO1 COOH-terminally GFP tagged in the genome and containing a CEN plasmid expressing RFP-Rud3p as in Fig. 1 A. (D) Distribution of the Golgi markers Gos1p and Vrg4p expressed as GFP-fusions in either wild-type cells or a mutant lacking ERV14 as indicated. GFP-tagged proteins expressed as in A.

Mentions: Because the RUD3 gene displays a synthetic lethal interaction with the RIC1 gene, it seemed likely that genes encoding proteins involved in Rud3p targeting will also be synthetically lethal when deleted in combination with the RIC1 gene. Thus, we examined the localization of GFP-Rud3p in deletion strains for all of the 113 genes whose deletion had been found to be synthetically lethal with ric1Δ by synthetic genetic array analysis (Tong et al., 2004; Fig. 5 A and Table S1, available at http://www.jcb.org/cgi/content/full/jcb.200407088/DC1). GFP-Rud3p showed a typical punctate distribution in all of the mutants, with one exception. In the mutant lacking the gene ERV14, GFP-Rud3p was instead found diffusely distributed throughout the cytosol (Fig. 5 B). This phenotype could be rescued by expression of untagged Erv14p or Erv14p tagged at the COOH terminus with two copies of the HA epitope, indicating that it was caused by loss of Erv14p and not due to an indirect effect of the deletion (Fig. 5 B). Moreover, the mislocalization of Rud3p did not reflect a general disturbance of the Golgi in the erv14Δ strain, as Uso1p-GFP remained largely localized to puncta (Fig. 5 C). In addition, the distributions of the Golgi SNARE protein Gos1p and the Golgi GDP-mannose transporter Vrg4p were also unaffected in the erv14Δ strain (Fig. 5 D).


The GTPase Arf1p and the ER to Golgi cargo receptor Erv14p cooperate to recruit the golgin Rud3p to the cis-Golgi.

Gillingham AK, Tong AH, Boone C, Munro S - J. Cell Biol. (2004)

Erv14p is required for the Golgi localization of Rud3p. (A) Fluorescent micrographs of live yeast (BY4741) expressing GFP-Rud3p from plasmid pE1, with the indicated genes deleted. (B) Fluorescent micrographs of an erv14Δ strain with the RUD3 ORF NH2-terminally GFP tagged in the genome and containing either an empty CEN plasmid or a similar plasmid encoding Erv14p-HA as indicated. (C) Fluorescent micrographs of a yeast strain lacking ERV14 and with USO1 COOH-terminally GFP tagged in the genome and containing a CEN plasmid expressing RFP-Rud3p as in Fig. 1 A. (D) Distribution of the Golgi markers Gos1p and Vrg4p expressed as GFP-fusions in either wild-type cells or a mutant lacking ERV14 as indicated. GFP-tagged proteins expressed as in A.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172552&req=5

fig5: Erv14p is required for the Golgi localization of Rud3p. (A) Fluorescent micrographs of live yeast (BY4741) expressing GFP-Rud3p from plasmid pE1, with the indicated genes deleted. (B) Fluorescent micrographs of an erv14Δ strain with the RUD3 ORF NH2-terminally GFP tagged in the genome and containing either an empty CEN plasmid or a similar plasmid encoding Erv14p-HA as indicated. (C) Fluorescent micrographs of a yeast strain lacking ERV14 and with USO1 COOH-terminally GFP tagged in the genome and containing a CEN plasmid expressing RFP-Rud3p as in Fig. 1 A. (D) Distribution of the Golgi markers Gos1p and Vrg4p expressed as GFP-fusions in either wild-type cells or a mutant lacking ERV14 as indicated. GFP-tagged proteins expressed as in A.
Mentions: Because the RUD3 gene displays a synthetic lethal interaction with the RIC1 gene, it seemed likely that genes encoding proteins involved in Rud3p targeting will also be synthetically lethal when deleted in combination with the RIC1 gene. Thus, we examined the localization of GFP-Rud3p in deletion strains for all of the 113 genes whose deletion had been found to be synthetically lethal with ric1Δ by synthetic genetic array analysis (Tong et al., 2004; Fig. 5 A and Table S1, available at http://www.jcb.org/cgi/content/full/jcb.200407088/DC1). GFP-Rud3p showed a typical punctate distribution in all of the mutants, with one exception. In the mutant lacking the gene ERV14, GFP-Rud3p was instead found diffusely distributed throughout the cytosol (Fig. 5 B). This phenotype could be rescued by expression of untagged Erv14p or Erv14p tagged at the COOH terminus with two copies of the HA epitope, indicating that it was caused by loss of Erv14p and not due to an indirect effect of the deletion (Fig. 5 B). Moreover, the mislocalization of Rud3p did not reflect a general disturbance of the Golgi in the erv14Δ strain, as Uso1p-GFP remained largely localized to puncta (Fig. 5 C). In addition, the distributions of the Golgi SNARE protein Gos1p and the Golgi GDP-mannose transporter Vrg4p were also unaffected in the erv14Δ strain (Fig. 5 D).

Bottom Line: Bornens. 2004.Cell. 118:323-335).In contrast, we find that this region binds to the Golgi in a GRAB domain-dependent manner, suggesting that GMAP-210 may not link the Golgi to gamma-tubulin and centrosomes.

View Article: PubMed Central - PubMed

Affiliation: Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 2QH, England, UK.

ABSTRACT
Rud3p is a coiled-coil protein of the yeast cis-Golgi. We find that Rud3p is localized to the Golgi via a COOH-terminal domain that is distantly related to the GRIP domain that recruits several coiled-coil proteins to the trans-Golgi by binding the small Arf-like GTPase Arl1p. In contrast, Rud3p binds to the GTPase Arf1p via this COOH-terminal "GRIP-related Arf-binding" (GRAB) domain. Deletion of RUD3 is lethal in the absence of the Golgi GTPase Ypt6p, and a screen of other mutants showing a similar genetic interaction revealed that Golgi targeting of Rud3p also requires Erv14p, a cargo receptor that cycles between the endoplasmic reticulum and Golgi. The one human protein with a GRAB domain, GMAP-210 (CEV14/Trip11/Trip230), is known to be on the cis-Golgi, but the COOH-terminal region that contains the GRAB domain has been reported to bind to centrosomes and gamma-tubulin (Rios, R.M, A. Sanchis, A.M. Tassin, C. Fedriani, and M. Bornens. 2004. Cell. 118:323-335). In contrast, we find that this region binds to the Golgi in a GRAB domain-dependent manner, suggesting that GMAP-210 may not link the Golgi to gamma-tubulin and centrosomes.

Show MeSH
Related in: MedlinePlus