Organization of Golgi Glycosyltransferases in Membranes: Complexity via Complexes

Young, William W.

doi:10.1007/s00232-004-0656-5

Cited by 46 publications

(30 citation statements)

References 93 publications

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“…The GAUT1 aa sequence surrounding this proposed cleavage site is consistent with the consensus motif ([R/K]-[X]n-[R/K], n = 0, 2, 4, or 6) recognized by subtilisin-like proprotein convertases in the secretory pathway (36). Proteolytic cleavage at stem regions by secretory pathway proteases has been documented with many other GTs (29,37). Whereas the resulting soluble, truncated GTs are typically secreted out of the cell, the N-glycosylation enzyme GlcNAcT-I was reported cleaved but retained in the Golgi via inclusion in HMW oligomers mediated Proteomics analyses were conducted on multiple independent sets of immunoprecipitations performed using anti-GAUT1-IgG (IP-GAUT1), anti-GAUT7-IgG (IP-GAUT7), and preimmune IgG (IP-control) (see details in SI Results and Discussion).…”

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confidence: 70%

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Galacturonosyltransferase (GAUT)1 and GAUT7 are the core of a plant cell wall pectin biosynthetic homogalacturonan:galacturonosyltransferase complex

Atmodjo

Sakuragi

Zhu

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

193

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Plant cell wall pectic polysaccharides are arguably the most complex carbohydrates in nature. Progress in understanding pectin synthesis has been slow due to its complex structure and difficulties in purifying and expressing the low-abundance, Golgi membranebound pectin biosynthetic enzymes. Arabidopsis galacturonosyltransferase (GAUT) 1 is an α-1,4-galacturonosyltransferase (GalAT) that synthesizes homogalacturonan (HG), the most abundant pectic polysaccharide. We now show that GAUT1 functions in a protein complex with the homologous GAUT7. Surprisingly, although both GAUT1 and GAUT7 are type II membrane proteins with single Nterminal transmembrane-spanning domains, the N-terminal region of GAUT1, including the transmembrane domain, is cleaved in vivo. This raises the question of how the processed GAUT1 is retained in the Golgi, the site of HG biosynthesis. We show that the anchoring of GAUT1 in the Golgi requires association with GAUT7 to form the GAUT1:GAUT7 complex. Proteomics analyses also identified 12 additional proteins that immunoprecipitate with the GAUT1:GAUT7 complex. This study provides conclusive evidence that the GAUT1: GAUT7 complex is the catalytic core of an HG:GalAT complex and that cell wall matrix polysaccharide biosynthesis occurs via protein complexes. The processing of GAUT1 to remove its N-terminal transmembrane domain and its anchoring in the Golgi by association with GAUT7 provides an example of how specific catalytic domains of plant cell wall biosynthetic glycosyltransferases could be assembled into protein complexes to enable the synthesis of the complex and developmentally and environmentally plastic plant cell wall. disulfide bond | biomass | primary cell wall | secondary cell wall

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supporting

confidence: 70%

“…Some GT complexes are known to contain disulfide bonds (29). We tested whether GAUT1 and GAUT7 associated via disulfide bonds by separating proteins from the Arabidopsis SP fraction on reducing versus nonreducing SDS/ PAGE followed by Western blotting (Fig.…”

mentioning

confidence: 99%

Galacturonosyltransferase (GAUT)1 and GAUT7 are the core of a plant cell wall pectin biosynthetic homogalacturonan:galacturonosyltransferase complex

Atmodjo

Sakuragi

Zhu

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

193

200

View full text Add to dashboard Cite

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“…It is now believed that cargo remains in one cisterna and that this Golgi compartment traffics in the anterograde direction, whereas the Golgi enzymes traffic backward by COPI vesicles. This model is based on the experimental observations that cargo indeed remains in a specific cisterna and that COPI vesicles are enriched with Golgi enzymes (70,71 ).…”

Section: Golgi Trafficmentioning

confidence: 99%

“…The fact that Golgi proteins have shorter transmembrane domains than do plasma membrane proteins suggests that cisternae of a specific compartment can accommodate glycosyltransferases with a transmembrane domain of matching length. However, it has been shown that some soluble forms of glycosyltransferases, which have lost their transmembrane domain, are retained in the Golgi probably as a result of being associated in complexes (70,71 ). It is likely that more independent signals act together to mediate efficient Golgi localization.…”

Section: Golgi Trafficmentioning

confidence: 99%

Mechanisms in Protein O-Glycan Biosynthesis and Clinical and Molecular Aspects of Protein O-Glycan Biosynthesis Defects: A Review

et al. 2006

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Background: Genetic diseases that affect the biosynthesis of protein O-glycans are a rapidly growing group of disorders. Because this group of disorders does not have a collective name, it is difficult to get an overview of O-glycosylation in relation to human health and disease. Many patients with an unsolved defect in Nglycosylation are found to have an abnormal O-glycosylation as well. It is becoming increasingly evident that the primary defect of these disorders is not necessarily localized in one of the glycan-specific transferases, but can likewise be found in the biosynthesis of nucleotide sugars, their transport to the endoplasmic reticulum (ER)/Golgi, and in Golgi trafficking. Already, disorders in O-glycan biosynthesis form a substantial group of genetic diseases. In view of the number of genes involved in O-glycosylation processes and the increasing scientific interest in congenital disorders of glycosylation, it is expected that the number of identified diseases in this group will grow rapidly over the coming years. Content: We first discuss the biosynthesis of protein O-glycans from their building blocks to their secretion from the Golgi. Subsequently, we review 24 different genetic disorders in O-glycosylation and 10 different genetic disorders that affect both N-and O-glycosylation. The key clinical, metabolic, chemical, diagnostic, and genetic features are described. Additionally, we describe methods that can be used in clinical laboratory screening for protein O-glycosylation biosynthesis defects and their

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“…In fact, there is compelling in vitro and in vivo evidence on the formation of oligomers by mammalian and yeast glycosyltransferases, respectively, from each major glycosylation category, namely, for proteoglycans, glycoproteins, and glycolipids (Schachter, 1986;Nilsson et al, 1994Nilsson et al, , 1996McCormick et al, 2000;Giraudo et al, 2001;Pinhal et al, 2001;Qian et al, 2001;Stolz and Munro, 2002;Young, 2004;Hassinen et al, 2011;Ferrari et al, 2012). Earlier studies mainly examined complex formation through in vitro coimmunoprecipitation (co-IP) assays following complete disruption of the cell, whereas in recent years, the advent of fluorescent protein technology and that of laser-based microscopy techniques utilizing bimolecular fluorescence complementation or Förster resonance energy transfer (FRET) have proven invaluable for the observation of proteinprotein interactions in vivo and in real time.…”

mentioning

confidence: 99%

Time-Resolved Fluorescence Imaging Reveals Differential Interactions ofN-Glycan Processing Enzymes across the Golgi Stack in Planta

et al. 2013

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N-Glycan processing is one of the most important cellular protein modifications in plants and as such is essential for plant development and defense mechanisms. The accuracy of Golgi-located processing steps is governed by the strict intra-Golgi localization of sequentially acting glycosidases and glycosyltransferases. Their differential distribution goes hand in hand with the compartmentalization of the Golgi stack into cis-, medial-, and trans-cisternae, which separate early from late processing steps. The mechanisms that direct differential enzyme concentration are still unknown, but the formation of multienzyme complexes is considered a feasible Golgi protein localization strategy. In this study, we used two-photon excitation-Förster resonance energy transfer-fluorescence lifetime imaging microscopy to determine the interaction of N-glycan processing enzymes with differential intra-Golgi locations. Following the coexpression of fluorescent protein-tagged amino-terminal Golgi-targeting sequences (cytoplasmic-transmembrane-stem [CTS] region) of enzyme pairs in leaves of tobacco (Nicotiana spp.), we observed that all tested cis-and medial-Golgi enzymes, namely Arabidopsis (Arabidopsis thaliana) Golgi a-mannosidase I, Nicotiana tabacum b1,2-N-acetylglucosaminyltransferase I, Arabidopsis Golgi a-mannosidase II (GMII), and Arabidopsis b1,2-xylosyltransferase, form homodimers and heterodimers, whereas among the late-acting enzymes Arabidopsis b1,3-galactosyltransferase1 (GALT1), Arabidopsis a1,4-fucosyltransferase, and Rattus norvegicus a2,6-sialyltransferase (a nonplant Golgi marker), only GALT1 and medial-Golgi GMII were found to form a heterodimer. Furthermore, the efficiency of energy transfer indicating the formation of interactions decreased considerably in a cis-to-trans fashion. The comparative fluorescence lifetime imaging of several full-length cis-and medial-Golgi enzymes and their respective catalytic domain-deleted CTS clones further suggested that the formation of protein-protein interactions can occur through their amino-terminal CTS region.The Golgi apparatus is a multifaceted, multitasking organelle that is pivotal to the life of the cell. Protein and lipid modifications, sorting of molecules, as well as the biosynthesis of cell wall polysaccharides all take place in the small stacks of flattened cisternae that make up the Golgi bodies, constituting the Golgi apparatus in plant cells. Among the various posttranslational modification reactions on proteins, the biosynthesis and processing of protein-bound N-linked oligosaccharides (N-glycans) is the most common. N-Glycans play a crucial role in protein folding, endoplasmic reticulum (ER) quality control (Liu and Howell, 2010), biotic (Saijo, 2010) and abiotic (Koiwa et al., 2003;Kang et al., 2008) stress responses, and are considered essential for the physicochemical properties and biological functions of glycoproteins. Consequently, the slightest alterations during N-glycan processing can drastically affect a protein's folding, stability, and biolog...

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Organization of Golgi Glycosyltransferases in Membranes: Complexity via Complexes

Cited by 46 publications

References 93 publications

Galacturonosyltransferase (GAUT)1 and GAUT7 are the core of a plant cell wall pectin biosynthetic homogalacturonan:galacturonosyltransferase complex

Galacturonosyltransferase (GAUT)1 and GAUT7 are the core of a plant cell wall pectin biosynthetic homogalacturonan:galacturonosyltransferase complex

Mechanisms in Protein O-Glycan Biosynthesis and Clinical and Molecular Aspects of Protein O-Glycan Biosynthesis Defects: A Review

Time-Resolved Fluorescence Imaging Reveals Differential Interactions ofN-Glycan Processing Enzymes across the Golgi Stack in Planta

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