Giantin, a novel conserved Golgi membrane protein containing a cytoplasmic domain of at least 350 kDa.

TLDR

The Golgi complex is composed of stacked cisternae linked by intercisternal cross‑bridges whose molecular identity remains unknown, and a large cytoplasmic domain is a feature shared with the calcium‑release channel of muscle. The authors propose that giantin, a conserved 400‑kDa integral membrane protein, may form these intercisternal cross‑bridges. They identified giantin as a 400‑kDa integral Golgi membrane protein conserved in mammals and birds, possessing a disulfide‑linked lumenal domain and a large cytoplasmic domain up to 350 kDa, supporting its role in cross‑bridge formation.

Abstract

The Golgi complex consists of a series of stacked cisternae in most eukaryotes. Morphological studies indicate the existence of intercisternal cross-bridge structures that may mediate stacking, but their identity is unknown. We have identified a 400-kDa protein, giantin, that is localized to the Golgi complex because its staining in double immunofluorescence experiments was coincident with that of galactosyltransferase, both in untreated cells and in cells treated with agents that disrupt Golgi structure. A monoclonal antibody against giantin yielded Golgi staining in one avian and all mammalian cell types tested, indicating that giantin is a conserved protein. Giantin exhibited reduced mobility on nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was recovered in membrane fractions after differential centrifugation or sucrose flotation, and was not released from membranes by carbonate extraction. Thus, giantin appears to be an integral component of the Golgi membrane with a disulfide-linked lumenal domain. Strikingly, the majority of the polypeptide chain is cytoplasmically disposed, because large (up to 350 kDa) proteolytic fragments of giantin could be released from intact Golgi vesicles. This feature, a large contiguous cytoplasmic domain, is present in the calcium-release channel of muscle that cross-bridges the sarcoplasmic reticulum and transverse tubule membranes. Therefore, giantin's localization, conservation, and physical properties suggest that it may participate in forming the intercisternal cross-bridges of the Golgi complex.