title: Human and viral Golgi anti-apoptotic proteins (GAAPs) oligomerize via different mechanisms and monomeric GAAP inhibits apoptosis and modulates calcium.
abstract: Golgi anti-apoptotic proteins (GAAPs) are hydrophobic proteins resident in membranes of the Golgi complex. They protect cells from a range of apoptotic stimuli, reduce the Ca(2+) content of intracellular stores, and regulate Ca(2+) fluxes. GAAP was discovered in camelpox virus, but it is highly conserved throughout evolution and encoded by all eukaryote genomes examined. GAAPs are part of the transmembrane Bax inhibitor-containing motif (TMBIM) family that also includes other anti-apoptotic and Ca(2+)-modulating membrane proteins. Most TMBIM members show multiple bands when analyzed by SDS-PAGE, suggesting that they may be oligomeric. However, the molecular mechanisms of oligomerization, the native state of GAAPs in living cells and the functional significance of oligomerization have not been addressed. TMBIM members are thought to have evolved from an ancestral GAAP. Two different GAAPs, human (h) and viral (v)GAAP were therefore selected as models to examine oligomerization of TMBIM family members. We show that both hGAAP and vGAAP in their native states form oligomers and that oligomerization is pH-dependent. Surprisingly, hGAAP and vGAAP do not share the same oligomerization mechanism. Oligomerization of hGAAP is independent of cysteines, but oligomerization of vGAAP depends on cysteines 9 and 60. A mutant vGAAP that is unable to oligomerize revealed that monomeric vGAAP retains both its anti-apoptotic function and its effect on intracellular Ca(2+) stores. In conclusion, GAAP can oligomerize in a pH-regulated manner, and monomeric GAAP is functional.
authors.affiliation: Department of Pathology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QP, United Kingdom.
authors.forename: Nuno
authors.initials: N
authors.lastname: Saraiva
authors.forename: David L
authors.initials: DL
authors.lastname: Prole
authors.forename: Guia
authors.initials: G
authors.lastname: Carrara
authors.forename: Carlos
authors.initials: C
authors.lastname: Maluquer de Motes
authors.forename: Benjamin F
authors.initials: BF
authors.lastname: Johnson
authors.forename: Bernadette
authors.initials: B
authors.lastname: Byrne
authors.forename: Colin W
authors.initials: CW
authors.lastname: Taylor
authors.forename: Geoffrey L
authors.initials: GL
authors.lastname: Smith
mesh_terms: D019943:Amino Acid Substitution; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D018155:Orthopoxvirus / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D051034:Inhibitor of Apoptosis Proteins; D008565:Membrane Proteins; C518281:TMBIM4 protein, human; D014764:Viral Proteins; D002118:Calcium
keywords: Apoptosis; Bax Inhibitor 1; Calcium; Golgi; Golgi Anti-apoptotic Protein; Oligomerization; Poxvirus; Protein Complexes; TMBIM; Viral Protein