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      The p24 Complex Contributes to Specify Arf1 for COPI Coat Selection

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          Abstract

          Golgi trafficking depends on the small GTPase Arf1 which, upon activation, drives the assembly of different coats onto budding vesicles. Two related types of guanine nucleotide exchange factors (GEFs) activate Arf1 at different Golgi sites. In yeast, Gea1 in the cis-Golgi and Gea2 in the medial-Golgi activate Arf1 to form COPI­coated vesicles for retrograde cargo sorting, whereas Sec7 generates clathrin/adaptor­coated vesicles at the trans-Golgi network (TGN) for forward cargo transport. A central question is how the same activated Arf1 protein manages to assemble different coats depending on the donor Golgi compartment. A previous study has postulated that the interaction between Gea1 and COPI would channel Arf1 activation for COPI vesicle budding. Here, we found that the p24 complex, a major COPI vesicle cargo, promotes the binding of Gea1 with COPI by increasing the COPI association to the membrane independently of Arf1 activation. Furthermore, the p24 complex also facilitates the interaction of Arf1 with its COPI effector. Therefore, our study supports a mechanism by which the p24 complex contributes to program Arf1 activation by Gea1 for selective COPI coat assembly at the cis-Golgi compartment.

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          Most cited references45

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          Coatomer is essential for retrieval of dilysine-tagged proteins to the endoplasmic reticulum.

          Dilysine motifs in cytoplasmic domains of transmembrane proteins are signals for their continuous retrieval from the Golgi back to the endoplasmic reticulum (ER). We describe a system to assess retrieval to the ER in yeast cells making use of a dilysine-tagged Ste2 protein. Whereas retrieval was unaffected in most sec mutants tested (sec7, sec12, sec13, sec16, sec17, sec18, sec19, sec22, and sec23), a defect in retrieval was observed in previously characterized coatomer mutants (sec21-1, sec27-1), as well as in newly isolated retrieval mutants (sec21-2, ret1-1). RET1 was cloned by complementation and found to encode the alpha subunit of coatomer. While temperature-sensitive for growth, the newly isolated coatomer mutants exhibited a very modest defect in secretion at the nonpermissive temperature. Coatomer from beta'-COP (sec27-1) and alpha-COP (ret1-1) mutants, but not from gamma-COP (sec21) mutants, had lost the ability to bind dilysine motifs in vitro. Together, these results suggest that coatomer plays an essential role in retrograde Golgi-to-ER transport and retrieval of dilysine-tagged proteins back to the ER.
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            Secretory protein biogenesis and traffic in the early secretory pathway.

            The secretory pathway is responsible for the synthesis, folding, and delivery of a diverse array of cellular proteins. Secretory protein synthesis begins in the endoplasmic reticulum (ER), which is charged with the tasks of correctly integrating nascent proteins and ensuring correct post-translational modification and folding. Once ready for forward traffic, proteins are captured into ER-derived transport vesicles that form through the action of the COPII coat. COPII-coated vesicles are delivered to the early Golgi via distinct tethering and fusion machineries. Escaped ER residents and other cycling transport machinery components are returned to the ER via COPI-coated vesicles, which undergo similar tethering and fusion reactions. Ultimately, organelle structure, function, and cell homeostasis are maintained by modulating protein and lipid flux through the early secretory pathway. In the last decade, structural and mechanistic studies have added greatly to the strong foundation of yeast genetics on which this field was built. Here we discuss the key players that mediate secretory protein biogenesis and trafficking, highlighting recent advances that have deepened our understanding of the complexity of this conserved and essential process.
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              Membrane traffic within the Golgi apparatus.

              Newly synthesized secretory cargo molecules pass through the Golgi apparatus while resident Golgi proteins remain in the organelle. However, the pathways of membrane traffic within the Golgi are still uncertain. Most of the available data can be accommodated by the cisternal maturation model, which postulates that Golgi cisternae form de novo, carry secretory cargoes forward and ultimately disappear. The entry face of the Golgi receives material that has been exported from transitional endoplasmic reticulum sites, and the exit face of the Golgi is intimately connected with endocytic compartments. These conserved features are enhanced by cell-type-specific elaborations such as tubular connections between mammalian Golgi cisternae. Key mechanistic questions remain about the formation and maturation of Golgi cisternae, the recycling of resident Golgi proteins, the origins of Golgi compartmental identity, the establishment of Golgi architecture, and the roles of Golgi structural elements in membrane traffic.
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                Author and article information

                Journal
                Int J Mol Sci
                Int J Mol Sci
                ijms
                International Journal of Molecular Sciences
                MDPI
                1422-0067
                03 January 2021
                January 2021
                : 22
                : 1
                : 423
                Affiliations
                [1 ]Department of Cell Biology, Faculty of Biology, University of Seville, 41012 Seville, Spain; ssabido@ 123456us.es (S.S.-B.); anamaripl87@ 123456hotmail.com (A.M.P.-L.); jmanzano@ 123456us.es (J.M.-L.); srodriguez13@ 123456us.es (S.R.-G.); auxi@ 123456us.es (A.A.-R.); acgomez@ 123456us.es (A.C.-G.); serglom@ 123456us.es (S.L.)
                [2 ]Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41012 Seville, Spain
                [3 ]Centro Andaluz de Biología Molecular y Medicina Regenerativa-CABIMER, Universidad de Sevilla-CSIC-Universidad Pablo de Olavide, 41012 Sevilla, Spain; ralf.wellinger@ 123456cabimer.es
                [4 ]Department of Genetics, University of Seville, 41012 Seville, Spain
                Author notes
                [* ]Correspondence: mmuniz@ 123456us.es
                [†]

                These authors contributed equally.

                Author information
                https://orcid.org/0000-0002-5753-2505
                https://orcid.org/0000-0001-5952-3568
                https://orcid.org/0000-0002-4421-6618
                https://orcid.org/0000-0001-8011-6991
                Article
                ijms-22-00423
                10.3390/ijms22010423
                7794930
                33401608
                2ef6ab9e-b919-4e40-bc17-1f5f0c4e5239
                © 2021 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 30 October 2020
                : 30 December 2020
                Categories
                Article

                Molecular biology
                p24 complex,copi,golgi,arfgefs,arf1
                Molecular biology
                p24 complex, copi, golgi, arfgefs, arf1

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