Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.4.99.6 (sialyltransferase)
 1,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Three key regulatory enzymes in ganglioside biosynthesis, sialyltransferase I (ST1), sialyltransferase II (ST2), and N-acetylgalactosaminyltransferase I (GalNAcT), have been expressed as fusion proteins with green, yellow, or red fluorescent protein (GFP, YFP, or RFP) in F-11A cells. F-11A cells are a substrain of murine neuroblastoma F-11 cells that contain only low endogenous ST2 and GalNAcT activity. The subcellular localization of the fusion proteins has been determined by fluorescence microscopy, and the ganglioside composition of these cells was analyzed by high-performance thin-layer chromatography (HPTLC). ST2-GFP (85 kDa) shows a distinct Golgi localization, whereas ST1-YFP (85 kDa) and GalNAcT-RFP (115 kDa) are broadly distributed in ER and Golgi. Untransfected F-11A cells contain mainly GM3, whereas stable transfection with ST2 or GalNAcT results in the predominant expression of b-series complex gangliosides (BCGs). This result indicates that the expression of ST2 enhances the activity of endogenous GalNAcT and vice versa. The specificity of this reaction has been verified by in vitro activity assays with detergent-solubilized enzymes, suggesting the formation of an enzyme complex between ST2 and GalNAcT but not with ST1. Complex formation has also been verified by co-immunoprecipitation of ST2-GFP upon transient transfection with GalNAcT-HA-RFP and by GFP-to-RFP FRET signals that are confined to the Golgi. FRET analysis also suggests that ST2-GFP binds tightly to pyrene-labeled GM3 but not to ST1. We hypothesize that an ST2-GM3 complex is associated with GalNAcT, resulting in the enhanced conversion of GM3 to GD3 and BCGs in the Golgi. Taken together, our results support the concept that ganglioside biosynthesis is tightly regulated by the formation of glycosyltransferase complexes in the ER and/or Golgi.
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PMID:Regulation of ganglioside biosynthesis by enzyme complex formation of glycosyltransferases. 1223 91

Golgi equivalents (GEs) process materials in the fungal secretory pathway. Despite the importance of localized secretion in fungal tip growth, GE behaviour in living hyphae has not been documented. The distribution was monitored of an Aspergillus nidulans putative GE-associated protein, CopA, tagged with GFP (CopA-GFP). This co-localized with a Golgi body/GE marker established in other systems, alpha-2,6-sialyltransferase, tagged with red fluorescent protein (ST-RFP). CopA-GFP and ST-RFP distributions responded similarly to brefeldin A, which impairs Golgi/GE trafficking. We used a CopA-GFP, hypA1 strain to study GE distribution and behaviour in growing A. nidulans hyphae. This strain has a wild-type phenotype at 28 degrees C, can be manipulated by changing growth temperature or by use of cytoskeleton inhibitors, and its GE behaviour is consistent with that in a wild-type-morphology strain. A. nidulans GEs were more abundant at hyphal tips than subapically, and showed saltatory motility in all directions. Anterograde GE movements predominated. These were positively correlated with, but at least 10-fold faster than, hyphal growth rate, under all growth and experimental conditions investigated. The actin inhibitor latrunculin B reduced both anterograde GE movement and hyphal growth rate, whereas the microtubule (MT) depolymerizer benomyl increased anterograde GE movement and decreased hyphal growth rate. The MT stabilizer taxol increased A. nidulans GE movement but not hyphal growth rate. A. nidulans GE motility appears to have a complex dependence on both actin and MTs. We present a model for apical delivery of growth materials in which A. nidulans GEs play a role in long-distance transport.
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PMID:Rapid tip-directed movement of Golgi equivalents in growing Aspergillus nidulans hyphae suggests a mechanism for delivery of growth-related materials. 1845 Oct 63