Identification of GFAT1-L, a novel splice variant of human glutamine: fructose-6-phosphate amidotransferase (GFAT1) that is expressed abundantly in skeletal muscle

Niimi, Manabu; Ogawara, Takeshi; Yamashita, Tatsuya; Yamamoto, Yuhei; Ueyama, Atsunori; Kambe, Toshimi; Okamoto, Takashi; Ban, Takashi; Tamanoi, Hidenori; Ozaki, Kouichi; Fujiwara, Tsutomu; Fukui, Hirokazu; Takáhashi, Eiichi; Kyushiki, Hiroyuki; Tanigami, Akira

doi:10.1007/s100380170022

Cited by 33 publications

(18 citation statements)

References 20 publications

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“…The GFPT1 enzyme is expressed in both nerve and muscle tissue [28,29]. Endplate morphology analysis in patient LGM14.3 showed unspecific abnormalities and the ultrastructural data we have available so far do not clarify if the origin of the neurotransmission defect is primarily presynaptic or postsynaptic, or indeed a combination of both pre-and postsynaptic abnormalities.…”

Section: Discussionmentioning

confidence: 50%

Congenital myasthenic syndrome with tubular aggregates caused by GFPT1 mutations

et al. 2011

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Congenital myasthenic syndrome (CMS) is a clinically and genetically heterogeneous group of inherited disorders of the neuromuscular junction. A difficult to diagnose subgroup of CMS is characterised by proximal muscle weakness and fatigue while ocular and facial involvement is only minimal. DOK7 mutations have been identified as causing the disorder in about half of the cases. More recently, using classical positional cloning, we have identified mutations in a previously unrecognised CMS gene, GFPT1, in a series of DOK7-negative cases. However, detailed description of clinical features of GFPT1 patients has not been reported yet. Here we describe the clinical picture of 24 limb-girdle CMS (LG-CMS) patients and pathological findings of 18 of them, all carrying GFPT1 mutations. Additional patients with CMS, but without tubular aggregates, and patients with non-fatigable weakness with tubular aggregates were also screened. In most patients with GFPT1 mutations, onset of the disease occurs in the first decade of life with characteristic limb-girdle weakness and fatigue. A common feature was beneficial and sustained response to acetylcholinesterase inhibitor treatment. Most of the patients who had a muscle biopsy showed tubular aggregates in myofibers. Analysis of endplate morphology in one of the patients revealed unspecific abnormalities. Our study delineates the phenotype of CMS associated with GFPT1 mutations and expands the understanding of neuromuscular junction disorders. As tubular aggregates in context of a neuromuscular transmission defect appear to be highly indicative, we suggest calling this condition congenital myasthenic syndrome with tubular aggregates (CMS-TA).

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Section: Discussionmentioning

confidence: 50%

Congenital myasthenic syndrome with tubular aggregates caused by GFPT1 mutations

et al. 2011

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“…Huynh et al (10) reported the characterization of rat liver GFAT, and Milewski et al (8) reported the biochemical properties of the Candida enzyme, which exhibits similarities to the mammalian forms. Recent descriptions of the GFAT1 isoform, GFAT1Alt, demonstrated that GFAT1 and GFAT1Alt differ in their sensitivity to inhibition by UDP-GlcNAc (11,12). These studies agree that feedback regulation by the pathway end product, UDP-GlcNAc, is a common feature of all mammalian GFATs studied to date, unlike the bacterial forms (13,14).…”

mentioning

confidence: 56%

Kinetic Characterization of Human Glutamine-fructose-6-phosphate Amidotransferase I

Broschat¹,

Gorka²,

Page³

et al. 2002

Journal of Biological Chemistry

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Glutamine-fructose-6-phosphate amidotransferase (GFAT) catalyzes the first committed step in the pathway for biosynthesis of hexosamines in mammals. A member of the N-terminal nucleophile class of amidotransferases, GFAT transfers the amino group from the L-glutamine amide to D-fructose 6-phosphate, producing glutamic acid and glucosamine 6-phosphate. The kinetic constants reported previously for mammalian GFAT implicate a relatively low affinity for the acceptor substrate, fructose 6-phosphate (Fru-6-P, K m 0.2-1 mM). Utilizing a new sensitive assay that measures the production of glucosamine 6-phosphate (GlcN-6-P), purified recombinant human GFAT1 (hGFAT1) exhibited a K m for Fru-6-P of 7 M, and was highly sensitive to product inhibition by GlcN-6-P. In a second assay method that measures the stimulation of glutaminase activity, a K d of 2 M was measured for Fru-6-P binding to hGFAT1. Further, we report that the product, GlcN-6-P, is a potent competitive inhibitor for the Fru-6-P site, with a K i measured of 6 M. Unlike other members of the amidotransferase family, where glutamate production is loosely coupled to amide transfer, we have demonstrated that hGFAT1 production of glutamate and GlcN-6-P are strictly coupled in the absence of inhibitors. Similar to other amidotransferases, competitive inhibitors that bind at the synthase site may inhibit the synthase activity without inhibiting the glutaminase activity at the hydrolase domain. GlcN-6-P, for example, inhibited the transfer reaction while fully activating the glutaminase activity at the hydrolase domain. Inhibition of hGFAT1 by the end product of the pathway, UDPGlcNAc, was competitive with a K i of 4 M. These data suggest that hGFAT1 is fully active at physiological levels of Fru-6-P and may be regulated by its product GlcN-6-P in addition to the pathway end product, UDP-GlcNAc.

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“…Because GFAT is expressed in all cells of the body (8,32), the effects of increased HBP flux may be relevant to other cell types and may be responsible for hyperglycemiaassociated cellular dysfunctions in other tissues. In fact, previous reports have associated GFAT overexpression to activation of proatherosclerotic pathways in messangial and vascular smooth muscle cells (22,39).…”

Section: Discussionmentioning

confidence: 99%

Hexosamine biosynthesis pathway flux promotes endoplasmic reticulum stress, lipid accumulation, and inflammatory gene expression in hepatic cells

Sage¹,

Walter²,

Shi³

et al. 2010

American Journal of Physiology-Endocrinology and Metabolism

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There is increasing evidence that endoplasmic reticulum (ER) stress contributes to the development of atherosclerosis in diabetes mellitus. The purpose of this study was to determine the effects of increased hexosamine biosynthesis pathway (HBP) flux on ER stress levels and the complications of ER stress associated with diabetes and atherosclerosis in hepatic cells. Glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme of the HBP, was overexpressed in HepG2 cells by use of an adenoviral expression system. The ER stress response and downstream effects, including activation of lipid and inflammatory pathways, were determined using real-time PCR, immunoblot analysis, and cell staining techniques. GFAT overexpression resulted in increased expression of ER stress markers, including Grp78, Grp94, calreticulin, and GADD153, relative to cells infected with an empty adenoviral vector. In addition, GFAT overexpression promoted lipid, but not cholesterol, accumulation in hepatic cells as well as inflammatory pathway activation. Treatment with 6-diazo-5-oxo-norleucine, a GFAT antagonist, blocked the effects of GFAT overexpression. Consistent with our in vitro data, hyperglycemic mice presented with elevated markers of hepatic ER stress, glucosamine and lipid accumulation. Together, these data suggest that HBP flux-induced ER stress plays a role in the development of hepatic steatosis and atherosclerosis under conditions of hyperglycemia.

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Identification of GFAT1-L, a novel splice variant of human glutamine: fructose-6-phosphate amidotransferase (GFAT1) that is expressed abundantly in skeletal muscle

Cited by 33 publications

References 20 publications

Congenital myasthenic syndrome with tubular aggregates caused by GFPT1 mutations

Congenital myasthenic syndrome with tubular aggregates caused by GFPT1 mutations

Kinetic Characterization of Human Glutamine-fructose-6-phosphate Amidotransferase I

Hexosamine biosynthesis pathway flux promotes endoplasmic reticulum stress, lipid accumulation, and inflammatory gene expression in hepatic cells

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