Cloning and expression of a novel type (III) of human γ‐glutamyltransferase truncated mRNA

Leh, Hervé; Courtay, C; Gérardin, Philippe; Wellman, Maria; Siest, Gérard; Visvikis, Athanase

doi:10.1016/0014-5793(96)00965-9

Cited by 6 publications

(2 citation statements)

References 20 publications

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“…However, we note that the identical intron sequences are also found in the human lung-specific group II GGT cDNAs where they reside not in the coding domain but in the 5Ј-untranslated region (11). Similar, but not identical, sequences are also present in a human intronic GGT promoter (21).…”

Section: Discussionmentioning

confidence: 86%

γ-Glutamyltransferase and Its Isoform Mediate an Endoplasmic Reticulum Stress Response

Joyce-Brady

Jean

Hughey

2001

Journal of Biological Chemistry

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Although use of multiple alternative first exons generates unique noncoding 5-ends for ␥-glutamyltransferase (GGT) cDNAs in several species, we show here that alternative splicing events also alter coding exons in mouse GGT to produce at least four protein isoforms. GGT⌬1 introduces CAG four bases upstream of the primary ATG codon and encodes an active GGT heterodimeric ectoenzyme identical to constitutive GGT cDNA but translational efficiency is reduced 2-fold. GGT⌬2-5 deletes the last eight nucleotides of exon 2 through most of exon 5 in-frame, selectively eliminating residues 96 -231 from the amphipathic N-terminal subunit, including four N-glycan consensus sites, while leaving the C-terminal hydrophilic subunit intact. GGT⌬7 introduces 22 bases from intron 7 causing a frameshift and a premature stop codon so a truncated polypeptide is encoded terminating with 14 novel residues but retaining the first 339 residues of the native GGT protein. GGT⌬8 -9 deletes the terminal four nucleotides of exon 8 plus all of exon 9 and inserts 24 bases from intron 9 in-frame so the C-terminal subunit of the encoded polypeptide loses residues 401-444 but gains eight internal hydrophobic residues. In contrast to the product of GGT⌬1, those derived from GGT⌬2-5, ⌬7, ⌬8 -9 all lack transferase activity and persist as singlechain glycoproteins retained largely in the endoplasmic reticulum as determined by immunofluorescence microscopy and constitutive endoglycosidase H sensitivity in metabolically labeled cells. The developmental-stage plus tissue-specific regulation of the alternative splicing events at GGT⌬7 and GGT⌬8 -9 implies unique roles for these GGT protein isoforms. The ability of the GGT⌬1 and GGT⌬7 to mediate the induction of C/EBP homologous protein-10, CHOP-10, and immunoglobulin heavy chain binding protein, BiP, implicates a specific role for these two GGT protein isoforms in the endoplasmic reticulum stress response.The entire intron/exon structure of the mouse ␥-glutamyltransferase gene (EC 2.3.2.2, GGT) 1 has been defined. This single copy gene is regulated by multiple alternative promoters that are coupled with alternative splicing mechanisms to generate several GGT cDNAs each with a unique 5Ј-noncoding region. But all encode the same protein, because coding exons appear to be spliced only in a constitutive fashion (1). The protein product is synthesized as a single-chain N-glycosylated propeptide, processed into an N-terminal amphipathic subunit and a smaller C-terminal subunit, and expressed on the cell surface where it functions as a key enzyme in glutathione metabolism (2). Several human GGT cDNAs also exhibit unique 5Ј-ends and encode a protein that is processed in a similar fashion and shares 79% amino acid identity with that of the mouse (3). In addition, an alternatively processed human GGT cDNA has been described that contains an insertion of 22 bases within the coding domain. The extra nucleotide bases introduce a frameshift and a premature stop codon so that the predicted polypeptide would be a truncat...

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Section: Discussionmentioning

confidence: 86%

γ-Glutamyltransferase and Its Isoform Mediate an Endoplasmic Reticulum Stress Response

Joyce-Brady

Jean

Hughey

2001

Journal of Biological Chemistry

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“…Most of them are localized on chromosome 22 [11]. Several GGT mRNAs have been identified: (a) GGT I mRNAs encode for an enzyme which catalytic properties are identical to those of GGT from human tissues [12]; (b) truncated forms of GGT I mRNA ( GGT mRNA I tr ) that putatively encode for the light subunit [13]; types I and I tr mRNAs are transcripts of the same gene [14]; (c) GGT II mRNAs are transcribed from a separate gene and appear mainly in the liver [15]; and (d) GGT III mRNAs are truncated and are encoded by a yet different GGT gene [10–16].…”

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confidence: 99%

Structure of the 5′ sequences of the human γ‐glutamyltransferase gene

Visvikis

Pawlak

Accaoui

et al. 2001

European Journal of Biochemistry

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In humans, five distinct mRNAs code for gamma-glutamyltransferase (GGT). Their coding regions are identical and their 5' untranslated regions exhibit both common and type-specific sequences. To elucidate the mecanisms that generate these different mRNAs, we cloned and determined the structure of the 5' region of the human GGT gene. The common regions of the 5' UTR are encoded by five exons, localized within a 2.4-kb region of the genomic DNA. Three of them are separated only by intron-donor or intron-acceptor sites at their boundaries. Alternative splicing of these exons may determine the unique pattern of the different GGT mRNA 5' UTRs in a tissue-specific manner. In addition, we have isolated a genomic fragment containing the most distal 5' sequences of the major GGT mRNA in HepG2 cells. Primer extension analysis revealed one major transcription initiation site while 5' RACE indicated that one more distal initiation site could be present. In the putative promoter sequence neither classical TATA or CAAT boxes were found. However, sites for AP1, AP2, CREB, GRE and SP1 transcription factors were identified. Chimeric plasmids, containing this genomic region fused to the luciferase gene, were transiently expressed in three cell lines of different origin: HeLa cells, ovarian carcinoma A2780 cells and V79 lung fibroblasts. The significant promoter activities obtained indicate a transcription start within this region. However, differences in the level of expression were found between the different cell lines used. These data suggest that the human GGT gene employs regulatory sequences and alternative splicing, and gene expression may therefore be regulated in tissue specific and cell-type-specific manners.

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The human gamma-glutamyltransferase gene family

et al. 2008

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Assays for gamma-glutamyl transferase (GGT1, EC 2.3.2.2) activity in blood are widely used in a clinical setting to measure tissue damage. The well-characterized GGT1 is an extracellular enzyme that is anchored to the plasma membrane of cells. There, it hydrolyzes and transfers gamma-glutamyl moieties from glutathione and other gamma-glutamyl compounds to acceptors. As such, it has a critical function in the metabolism of glutathione and in the conversion of the leukotriene LTC4 to LTD4. GGT deficiency in man is rare and for the few patients reported to date, mutations in GGT1 have not been described. These patients do secrete glutathione in urine and fail to metabolize LTC4. Earlier pre-genome investigations had indicated that besides GGT1, the human genome contains additional related genes or sequences. These sequences were given multiple different names, leading to inconsistencies and confusion. Here we systematically evaluated all human sequences related to GGT1 using genomic and cDNA database searches and identified thirteen genes belonging to the extended GGT family, of which at least six appear to be active. In collaboration with the HUGO Gene Nomenclature Committee (HGNC) we have designated possible active genes with nucleotide or amino acid sequence similarity to GGT1, as GGT5 (formerly GGL, GGTLA1/GGT-rel), GGT6 (formerly rat ggt6 homologue) and GGT7 (formerly GGTL3, GGT4). Two loci have the potential to encode only the light chain portion of GGT and have now been designated GGTLC1 (formerly GGTL6, GGTLA4) and GGTLC2. Of the five full-length genes, three lack of significant nucleotide sequence homology but have significant (GGT5, GGT7) or very limited (GGT6) amino acid similarity to GGT1 and belong to separate families. GGT6 and GGT7 have not yet been described, raising the possibility that leukotriene synthesis, glutathione metabolism or gamma-glutamyl transfer is regulated by their, as of yet uncharacterized, enzymatic activities. In view of the widespread clinical use of assays that measure gamma-glutamyl transfer activity, this would appear to be of significant interest.

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Cloning and expression of a novel type (III) of human γ‐glutamyltransferase truncated mRNA

Cited by 6 publications

References 20 publications

γ-Glutamyltransferase and Its Isoform Mediate an Endoplasmic Reticulum Stress Response

γ-Glutamyltransferase and Its Isoform Mediate an Endoplasmic Reticulum Stress Response

Structure of the 5′ sequences of the human γ‐glutamyltransferase gene

The human gamma-glutamyltransferase gene family

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