title: Oligomerization of the Sec7 domain Arf guanine nucleotide exchange factor GBF1 is dispensable for Golgi localization and function but regulates degradation.
abstract: Members of the large Sec7 domain-containing Arf guanine nucleotide exchange factor (GEF) family have been shown to dimerize through their NH2-terminal dimerization and cyclophilin binding (DCB) and homology upstream of Sec7 (HUS) domains. However, the importance of dimerization in GEF localization and function has not been assessed. We generated a GBF1 mutant (91/130) in which two residues required for oligomerization (K91 and E130 within the DCB domain) were replaced with A and assessed the effects of these mutations on GBF1 localization and cellular functions. We show that 91/130 is compromised in oligomerization but that it targets to the Golgi in a manner indistinguishable from wild-type GBF1 and that it rapidly exchanges between the cytosolic and membrane-bound pools. The 91/130 mutant appears active as it integrates within the functional network at the Golgi, supports Arf activation and COPI recruitment, and sustains Golgi homeostasis and cargo secretion when provided as a sole copy of functional GBF1 in cells. In addition, like wild-type GBF1, the 91/130 mutant supports poliovirus RNA replication, a process requiring GBF1 but believed to be independent of GBF1 catalytic activity. However, oligomerization appears to stabilize GBF1 in cells, and the 91/130 mutant is degraded faster than the wild-type GBF1. Our data support a model in which oligomerization is not a key regulator of GBF1 activity but impacts its function by regulating the cellular levels of GBF1.
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: Jay M
authors.initials: JM
authors.lastname: Bhatt
authors.affiliation: Department of Veterinary Medicine, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, Maryland; and.
authors.forename: Ekaterina G
authors.initials: EG
authors.lastname: Viktorova
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: Theodore
authors.initials: T
authors.lastname: Busby
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: Paulina
authors.initials: P
authors.lastname: Wyrozumska
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia.
authors.forename: Laura E
authors.initials: LE
authors.lastname: Newman
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: Helen
authors.initials: H
authors.lastname: Lin
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: Eunjoo
authors.initials: E
authors.lastname: Lee
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama;
authors.forename: John
authors.initials: J
authors.lastname: Wright
authors.affiliation: Department of Veterinary Medicine, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, Maryland; and.
authors.forename: George A
authors.initials: GA
authors.lastname: Belov
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Kahn
authors.affiliation: Department of Cell, Developmental and Integrative Biology, University of Alabama at Birmingham, Birmingham, Alabama; esztul@uab.edu.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Sztul
mesh_terms: D045744:Cell Line, Tumor; D020755:Coat Protein Complex I / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D059748:Proteolysis
chemical_list: D020755:Coat Protein Complex I; C120445:GBF1 protein, human; D020662:Guanine Nucleotide Exchange Factors
keywords: ADP-ribosylation factors; GBF1; Golgi; Sec7 guanine nucleotide exchange factor; membrane trafficking; oligomerization