GMx33: A Novel Family of trans‐Golgi Proteins Identified by Proteomics

Abstract

The known functions of the Golgi complex include the sorting, packaging, post-translational modification, and transport of secretory proteins, membrane proteins, and lipids. Other functions still remain elusive to cell biologists. With the goal of identifying novel Golgi proteins, a proteomics project was undertaken to map the major proteins of the organelle using two-dimensional gels, to identify the unknowns using tandem mass spectrometry, and to screen for Golgi residents using GFP-fusion constructs. Multiple unknowns were identified, and the initial characterization of one of these proteins is reported here. GMx33a is a member of a conserved family of cytosolic Golgi-associated proteins with no known homology to any known functional domain or protein.Biochemical analyses show that GMx33a differentially partitions into all phases of multiple detergent extractions, and two-dimensional immunoblots reveal that there are multiple differentially modified forms of GMx33a associated with the Golgi, several of which are phosphorylated. Evidence suggests that these posttranslational modifications regulate its association with the Golgi. GMx33a was not found on Golgi budded vesicles, and immuno-electron microscopy co-localizes GMx33a to the trans-face on the same three cisternae as TGN38 in normal rat kidney cells. This work represents the preliminary characterization of a novel family of trans-Golgi-associated proteins.

GOLPH3 Mediated Golgi Stress Response in Modulating N2A Cell Death upon Oxygen-Glucose Deprivation and Reoxygenation Injury

“…b (II) The histogram of apoptosis proportion detected by flow cytometry before or after G3 ShRNA transfection is shown. Apoptosis was detected by annexin V-FITC/PI in the control, OGD/R 4-h, OGD/R 24-h, and corresponding OGD/R groups after G3 ShRNA transfection (*P<0.01 compared to Normal/NC and OGD/R 24-h; # P<0.01 in the OGD/R 4-h groups compared to the OGD/R 24-h groups; n=3, mean± SD) (RQ relative quantification, CON normal control, N.C negative control, ShRNA GOLPH3 ShRNA) maintenance, receptor sorting, protein glycosylation, and the cell signaling pathways that localize to the cytoplasmic face of the trans-Golgi [4][5][6][7][8][9][10][11]. Subcellular localization studies revealed the co-localization of GOLPH3 to the Golgi apparatus.…”

“…Mammalian GOLPH3, also known as GPP34/GM×33/MI-DAS, is an abundant 34-kDa phosphorylated protein initially identified through a proteomic analysis of detergent-insoluble proteins in an isolated Golgi fraction [4]. It represents an exciting new class of Golgi outer membrane protein involved in vesicular trafficking, Golgi architecture maintenance, receptor sorting, protein glycosylation, and cell signaling pathways that localize to the cytoplasmic face of the trans-Golgi [5][6][7][8][9][10][11][12].…”

Localization of Golgi-resident glycosyltransferases

“…Vps74p bound both the CT of glycosyltransferases as well as to the COPI coat, providing a mechanism by which these enzymes can be incorporated into retrograde transport vesicles [75]. Even though the Vps74p-binding motif is not present in the CTs of mammalian glycosyltransferases, the human orthologs of Vps74p (i.e., Gpp34/ GmX33a and Gpp34R/Gmx33b [79,80]) could still functionally substitute for Vps74p when they where expressed in yeast cells [75]. Although Vps74-like proteins are present in the majority of eukaryotes, which suggests a conserved mechanism, homologs have yet to be identified in higher plants and protozoa, an observation that supports the operation of multiple mechanisms in the retention of glycosyltransferases in the Golgi.…”

GOLPH3 Overexpression is Closely Correlated with Poor Prognosis in Human Non-Small Cell Lung Cancer and Mediates its Metastasis Through Upregulating MMP-2 and MMP-9

“…GOLPH3, also known as GPP34, GMX33, or MIDAS [4,5], is a phosphorylated protein located in the Golgi complex. Schmitz et al previously found that deletion of VPS74, which is a homologue of GOLPH3 in budding yeast, led to defective transition and secretion of glycosylated protein [14], and the authors proposed that GOLPH3 affects glycosyl transferases in the Golgi complex and is therefore involved in protein glycosylation.…”

“…Golgi phosphoprotein 3 (GOLPH3) is a highly conserved protein with a molecular weight of 34 kDa [4,5] that is predominantly located in the external membrane of the trans-face of the Golgi, and it was initially identified through a proteomics analysis of the Golgi apparatus [4]. In 2009, Scott et al first reported the overexpression of GOLPH3 in cancer [6].…”