Laboratory for Glycobiology and Glycotechnology, Department of Industrial Chemistry, School of Engineering, Tokai University, 1117 Kitakaname, Hiratsuka City, Kanagawa 2591292, Japan
Received on August 2, 1999; accepted on August 20, 1999.
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Abstract |
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Key words: cDNA cloning/EST database/glycosylation/lipid-linked oligosaccharide/mannosyltransferase
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Introduction |
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We have studied the effect of alterations of the biosynthesis of lipid-linked oligosaccharides on cell growth, for example, the reversible G1 arrest of the human Burkitt lymphoma cell line, Raji, by tunicamycin (Nishikawa et al., 1980) and the isolation of a mouse FM3A mutant G258 cell line that has temperature-sensitive defects in both cell growth and the synthesis of lipid-linked oligosaccharides (Nishikawa, 1984
). The biochemical lesion of the G258 mutant may have resulted from a defect in GDP-mannose dependent mannosylation of Man3GlcNAc2-P-P-Dol (mannosyltransferase IV) (Nishikawa, 1991
). We have reported on a human gene that complements the defect of the synthesis of lipid-linked oligosaccharide of the G258 mutant (Kataoka et al., 1997
, 1999). The latter study prompted us to attempt the isolation of other GDP-mannose dependent mannosyltransferase genes. The present study reports the identification of the human gene, designated Hmat-1, that must encode mannosyltransferase I (MT-I) which catalyzes the following reaction:
GlcNAcß14GlcNAc-P-P-Dol + GDP-Man
Manß14GlcNAcß1
4GlcNAc-P-P-Dol + GDP
MT-I catalyzes the first mannosylation step in the synthesis of lipid-linked oligosaccharides and is the only mannosyltransferase which forms the ß14 mannose linkage in the lipid-linked oligosaccharides (Couto et al., 1984
). The MT-I gene has already been isolated from yeast (Albright and Robbins, 1990
), nematode (Wilson et al., 1994
), and slime mold (Lee et al., 1997
). In this study, we have cloned and characterized the cDNA containing the Hmat-1 gene. This is the first report on the cloning of a mammalian mannosyltransferase gene involved in the early assembly of lipid-linked oligosaccharides.
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Results |
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Amino acid sequence deduced from the Hmat-1 gene
Based on the nucleotide sequence of the 1.8 kb Hmat-1 cDNA, we deduced the amino acid sequence (Figure 4). The predicted protein consists of 464 amino acid residues equivalent to a 52.5 kDa molecular weight. It has no potential N-glycosylation site (-Asn-X-Ser/Thr-), although the S.cerevisiae MT-I protein has four such sites (Albright and Robbins, 1990). Moreover, it has a hydrophobic region at the amino terminus (Figure 5, amino acids 620) followed by short hydrophilic and hydrophobic regions similar to the S.cerevisiae MT-I protein.
The amino acid sequence of the protein that the Hmat-1 gene encodes was compared with those of both the S.cerevisiae MT-I (Albright and Robbins, 1990) and the C.elegans T26A5.4 gene product (Wilson et al., 1994
) which was predicted to be MT-I (Figure 5). The Hmat-1 gene product showed 36% identity with both MT-I proteins.
The Hmat-1 gene can complement the yeast alg1-1 mutation
To ascertain whether the 1.8 kb Hmat-1 cDNA encodes the human MT-I, we subcloned it into the S.cerevisiae episomal expression vector, YEp352GAP, which contains URA3 gene and GAP promoter sequence (Yoko-o et al., 1993). Then, with the resultant recombinant plasmid, we transformed the S.cerevisiae PRY56 strain, which has the alg1-1 mutation (Huffaker and Robbins, 1982
). The PRY56 strain did not grow on the plate of YPD medium at 36°C nor on the plates of SD medium lacking uracil [SD(Ura-) medium] at either 26°C or 36°C (Figure 6BD). The mock-transformant grew on the SD(Ura-) plate at 26°C (Figure 6B), but did not grow on either the YPD or SD(Ura-) plates at 36°C (Figure 6C,D). Only the transformant with the Hmat-1 cDNA grew under the nonpermissive condition for the alg1-1 mutant (i.e., incubation at 36°C for 3 days, Figure 6C,D). To prove the presence of the introduced Hmat-1 cDNA, PCR with A1-10FX and A1-8RS primers was performed using as templates DNA from the PRY56 strain, the mock-transformant and the transformant with the Hmat-1 cDNA. The 1.8 kb PCR product was detected only in the transformant with the Hmat-1 cDNA (data not shown). These observations indicate that the expressed Hmat-1 gene product could complement the temperature-sensitivity for cell growth due to the S.cerevisiae alg1-1 mutation.
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Discussion |
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MT-I is involved in the first mannosylation step of lipid-linked oligosaccharide synthesis. In yeast, the defect in the function of this enzyme (i.e., alg1-1 mutation) directly resulted in underglycosylation of glycoproteins and cell death (Huffaker and Robbins, 1982). Furthermore, expression of the MT-I gene is regulated at the transcriptional level in the cell division cycle (Lennon et al., 1995
) or morphogenesis (Lee et al., 1997
). These studies prove that MT-I is one of the key enzymes in the assembly of lipid-linked oligosaccharides.
We have isolated the 1.8 kb Hmat-1 cDNA and determined its nucleotide sequence (Figure 4). It contained both a methionine codon whose surrounding sequence (Figure 4, nucleotides 1019) matched a Kozak initiation codon consensus sequence (Kozak, 1984) and a termination codon. These results suggest that it contains the full-length coding region of the Hmat-1 gene. The predicted Hmat-1 protein was very similar to the S.cerevisiae ALG1 protein and was suggestive of a type II transmembrane protein. However, in this protein, there was no sequence which was homologous to the potential dolichol recognition sequence in the S.cerevisiae ALG1 protein (Albright et al., 1989
). Alternatively, the Hmat-1 protein had eight highly conserved sequences shared with ALG1 protein and T26A5.4 gene product (Figure 5, 39VLGDXXXSPR48, 130QXPPXXP136, 154IDWHNXXY161, 283STSXTXDEXXXILXXA298, 319ITGKGP324, 347WLXXEDYP354, 361DXGXXLHXSXSGXDLPMKXXDXFGXXXPXXA391, and 399ELVXXXXNG407). Moreover, these conserved sequences also exist in MT-I from Dictyostelium discoideum (Lee et al., 1997
) and Schizosaccharomyces pombe (GenBank accession number Z99753) (data not shown). Some of these conserved regions may function in the topological localization to the rough ER membrane, in the recognition of GDP-Man and/or GlcNAc2-P-P-Dol, and in the catalytic domain of this enzyme. Particularly, the highly hydrophobic N-terminal region containing the 39VLGDXXXSPR48 sequence may play an important role in functional expression of this enzyme, because the truncated Hmat-1 cDNA lacking this region (Figure 4, nucleotides 1186) could not complement the S.cerevisiae alg1-1 mutation (data not shown).
The fact that the 1.8 kb Hmat-1 cDNA complemented the S.cerevisiae alg1-1 mutation strongly suggests that the Hmat-1 gene encodes human MT-I and that the expressed human MT-I functions normally in yeast. This leads to the possibility that other human glycosyltransferases involved in the early assembly of lipid-linked oligosaccharides should be functionally conserved between yeast and human. This possibility is supported by the successful complementation of the S.cerevisiae alg7 mutation with human GlcNAc-1-P transferase (Eckert et al., 1998). If all the glycosyltransferases involved in the biosynthesis of lipid-linked oligosaccharides were functionally conserved between yeast and human, the S.cerevisiae N-glycosylation mutants such as alg2 (Huffaker and Robbins, 1983
), alg3 (Huffaker and Robbins, 1983
), alg6 (Reiss et al., 1996
), alg8 (Stagljar et al., 1994
), alg9 (Burda et al., 1996
), and alg10 (Burda and Aebi, 1998
) might be useful for the isolation of their human counterpart genes by the expression cloning method.
The importance of Asn-linked glycosylation in human has been demonstrated in various ways. Particularly, in the inherited group of diseases, carbohydrate deficient glycoprotein syndrome (CDGS), the Asn-linked oligosaccharides were severely impaired either in number per molecule of glycoprotein (underglycosylation, CDGS type I) or in structure (CDGS type II). It has been demonstrated that CDGS type II is due to the deficiency of N-acetylglucosaminyltransferase II (Jaeken et al., 1994; Tan et al., 1996
). On the other hand, it has been demonstrated that about 80% of patients with CDGS type I have a deficiency of phosphomannomutase (CDGS type Ia) (Van Schaftingen and Jaeken, 1995
). A deficiency of the enzyme phosphomannose isomerase has also been demonstrated to be responsible for patients showing a CDGS type I phenotype (CDGS type Ib) (de Koning et al., 1998
; Niehues et al., 1998
). Genes for the human glycosyltransferases involved in the synthesis of lipid-linked oligosaccharides are potential mutation sites for other subtypes of CDGS type I. Recently, it have been reported that a deficiency of glucosyltransferase I involved in the synthesis of lipid-linked oligosaccharides caused underglycosylation in a CDGS patient (CDGS type Ic or V) (Körner et al., 1998
; Burda et al., 1998
). Therefore, genetic analysis of CDGS patients for defects in mannosyltransferase genes involved in the synthesis of lipid-linked oligosaccharides may be clinically useful.
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Materials and methods |
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Design of gene-specific PCR primers
Based on the coincident nucleotide sequences among seven homologous human EST clones detected in the homology search described above (Figure 1), we designed two PCR primers, A1-1F and A1-2R (Table I, Greiner Japan). PCR primers, A1-10F and A1-8RS (Table I, Greiner Japan) were designed from the 5' terminus of AA675921 (GenBank accession number) and 3' terminus of HR1-4, respectively (Figure 3). These primers also contained restriction enzyme recognition sequences at their 5' ends, which were used for subcloning into the plasmid vector (Table I, underlined). A1-10FX is the same as A1-10F, except a XhoI site, instead of a BamHI site in A1-10F.
Screening of cDNA library by PCR
Using two primers, A1-1F and A1-2R, we screened a gt10 human fetal brain cDNA library (Clontech) by PCR. The PCR reaction mixture was prepared as follows. Phage solution containing about 1010 clones was first heated at 90°C for 10 min. The final PCR reaction mixture contained 10 mM Tris pH 8.0, 50 mM KCl, 1.5 mM MgCl2, 200 µM each dNTPs, 1 µM of both A1-1F primer and A1-2R primer, and 2.5 U Taq DNA polymerase (Roche Diagnostics) in a total volume of 100 µl. After overlaying 100 µl mineral oil (Sigma), the first round of PCR was performed by 30 cycles as follows: 94°C 15 sec (45 sec at initial cycle), 60°C 30 sec, 72°C 2 min (10 min at the last cycle) using PC-700 thermal cycler (Astek). The second round of PCR was carried out using 5 µl of the first PCR reaction mixture under the same conditions. After the PCR, 10 µl of the PCR reaction mixture was analyzed on 1% agarose gel electrophoresis using
DNA (Takara) digested with Sty I (Roche Diagnostics) as size marker.
Rescreening of cDNA library
Using the 430 bp PCR product (Figure 2, lane B) obtained from the above PCR as a probe, the same cDNA library was rescreened by plaque hybridization. Digoxigenin (DIG)-labeled probe was prepared by Digoxigenin (DIG) DNA Labeling Kit (Roche Diagnostics).
The cDNA library was diluted with SM solution (50 mM Tris HCl pH 7.5, 0.1 M NaCl, 8 mM MgSO4, 0.01% gelatin). The Escherichia coli C600hfl strain (hflA150[chr::Tn10] lacY1 leuB6 mcrA- supE44 thi-1 thr-1 tonA21) was infected with the diluted cDNA library at 37°C for 20 min, mixed with LB liquid medium containing 0.7% agar, plated onto LB solid medium containing 1.5% agar, and incubated at 37°C for 8 h in order to form about 50,000 plaques per plate. Plaques were transferred onto Colony Plaque Screen nylon membranes (NEN) and DNAs on the membranes were denaturated by 0.5 N NaOH/1.5M NaCl solution for 1 min, neutralized by 0.5 M Tris HCl pH 7.5/1.5M NaCl solution for 1 min, and washed in 2 x SSC solution (0.3 M NaCl, 0.034 M sodium citrate) for 5 min. The membranes were dried for plaque hybridization.
Plaque hybridization was carried out in the mixture containing 5 x SSC (0.75 M NaCl, 0.085M sodium citrate), 0.1% (w/v) N-lauroylsarcosine, 0.02% (w/v) SDS, 1% blocking reagent (Roche Diagnostics) at 68°C for 20 h. After washing with Solution I (2 x SSC, 0.1% SDS) at room temperature for 15 min twice and then solution II (0.2 x SSC, 0.1% SDS) at 65°C for 30 min twice, positive plaques were detected, using DIG Luminescent Detection Kit (Roche Diagnostics) and Hyper Film-ECL (Amersham).
Each positive plaque was recovered in 100 µl of SM solution and subjected to PCR. Two PCR primers, GT10-F and GT10-R (Table I, Greiner Japan), which were designed based on the flanking sequences of the gt10 cloning site, were used for amplification of the cDNA inserts.
Cloning and nucleotide sequencing of cDNAs
The original PCR product (about 430 bp) and the cDNA insert from HR1-3 were digested with BamHI and HindIII (Roche Diagnostics) and ligated into a plasmid vector, pUC118 or pUC119 (Vieira and Messing, 1987) previously digested with the same restriction enzymes. The cDNA insert from HR1-4 was digested with BamHI and EcoRI (Roche Diagnostics) and ligated into a plasmid vector, pUC118 or pUC119 previously digested with the same restriction enzymes. All the ligation reactions were carried out with DNA Ligation Kit Ver.2 (Takara). The ligation mixtures were used to transform E.coli JM109 strain (recA1 endA1 gyrA96 thi hsdR17 supE44 relA1
(lac-proAB)/F'[traD36 proAB+ lacIq lacZ
M15]) by the calcium chloride method (Cohen et al., 1972
). Selection of the resultant transformants was performed by the standard method (Sambrook et al., 1989
).
Cloning of the 1.8 kb Hmat-1 cDNA was performed as follows. Marathon-Ready cDNA (from human brain, Clontech) was subjected to PCR with the primers, A1-10F and A1-8RS. PCR was carried out under the same condition described above except that the total volume of the reaction mixture at the first round of PCR was 50 µl. The PCR product (about 1.8 kb) was digested with BamHI and SalI (Roche Diagnostics), ligated to pUC118 or pUC119 previously digested with the same restriction enzymes. The ligation reaction, transformation of JM109 strain, and selection of the resultant transformants were performed as described above.
Single-stranded DNAs were prepared by infecting the resultant transformants with the helper phage M13K07 (Vieira and Messing, 1987) and sequenced by the dideoxy chain termination method (Sanger et al., 1977
). If necessary, construction of deletion subclones for sequencing was carried out with Deletion Kit for Kilo-Sequence (Takara). The electrophoresis for sequencing with 8% polyacrylamide gel was performed at 15 mA for 4.5 to 9 h. Sequencing reaction and detection of DNA ladders were performed by Sequencing High -Cycle- Kit (Toyobo) and Hyper Film-ECL (Amersham).
Complementation test
The S.cerevisiae PRY56 strain (MATa alg1-1 ura3-52) (Huffaker and Robbins, 1982), was maintained on nutrient YPD medium at 26°C (permissive temperature for PRY56). The yeast episomal expression vector, YEp352GAP (Yoko-o et al., 1993
), which contains URA3 gene and GAP promoter, was used for the expression of the Hmat-1 gene in PRY56 strain.
The 1.8 kb Hmat-1 cDNA subcloned into the pUC vector was reamplified by PCR with the primers, A1-10FX and A1-8RS. PCR was carried out under the same condition described above. The PCR product was digested with XhoI and SalI (Roche Diagnostics) and ligated into YEp352GAP previously digested with the same restriction enzymes. The ligation reaction, transformation of JM109 strain, and selection of the resultant transformants were performed as described above. The recombinant plasmid DNA was prepared by the standard method (Sambrook et al., 1989) and the PRY56 strain was transformed by the lithium acetate method (Ito et al., 1983
). The transformants were first selected as uracil-auxotrophs on synthetic SD medium lacking uracil [SD(Ura-) medium] at 26°C, and then the growth of the transformants was examined on YPD and SD(Ura-) media at 36°C (nonpermissive temperature for PRY56).
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Acknowledgments |
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Abbreviations |
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Footnotes |
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References |
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Abeijon,C. and Hirschberg,C.B. (1992) Topography of glycosylation reactions in the endoplasmic reticulum. Trends Biochem. Sci., 17, 3236.[ISI][Medline]
Albright,C.F. and Robbins,P.W. (1990) The sequence and transcript heterogeneity of the yeast gene ALG1, an essential mannosyltransferase involved in N-glycosylation. J. Biol. Chem., 265, 70427049.
Albright,C.F., Orlean,P. and Robbins,P.W. (1989) A 13-amino acid peptide in three yeast glycosyltransferases may be involved in dolichol recognition. Proc. Natl. Acad. Sci. USA, 86, 73667369.[Abstract]
Altschul,S.F., Gish,W., Miller,W., Myers,E.W. and Lipman,D.J. (1990) Basic local alignment search tool. J. Mol. Biol., 215, 403410.[ISI][Medline]
Arnold,E. and Tanner,W. (1982) An obligatory role of protein glycosylation in the life cycle of yeast cells. FEBS Lett., 148, 4953.[ISI][Medline]
Barnes,G., Hansen,W.J., Holcomb,C.L. and Rine,J. (1984) Asparagine-linked glycosylation in Saccharomyces cerevisiae: genetic analysis of an early step. Mol. Cell. Biol., 4, 23812388.[ISI][Medline]
Burda,P. and Aebi,M. (1998) The ALG10 locus of Saccharomyces cerevisiae encodes the -1,2 glucosyltransferase of the endoplasmic reticulum: the terminal glucose of the lipid-linked oligosaccharide is required for efficient N-linked glycosylation. Glycobiology, 8, 455462.
Burda,P., Borsig,L., de Rijk-van Andel,J., Wevers,R., Jaeken,J., Carchon,H., Berger,E.G. and Aebi,M. (1998) A novel carbohydrate-deficient glycoprotein syndrome characterized by a deficiency in glucosylation of the dolichol-linked oligosaccharide. J. Clin. Invest., 102, 647652.
Burda,P., te Heesen,S., Brachat,A., Wach,A., Düsterhöft,A. and Aebi,M. (1996) Stepwise assembly of the lipid-linked oligosaccharide in the endoplasmic reticulum of Saccharomyces cerevisiae: identification of the ALG9 gene encoding a putative mannosyltransferase. Proc. Natl. Acad. Sci. USA, 93, 71607165.
Cohen,S.N., Chang,A.C.Y. and Hsu,L. (1972) Nonchromosomal antibiotic resistance in bacteria: Genetic transformation of Escherichia coli by R-factor DNA. Proc. Natl. Acad. Sci. USA, 69, 21102114.[Abstract]
Couto,J.R., Huffaker,T.C. and Robbins,P.W. (1984) Cloning and expression in Escherichia coli of a yeast mannosyltransferase from the asparagine-linked glycosylation pathway. J. Biol. Chem., 259, 378382.
de Koning,T.J., Dorland,L., van Diggelen,O.P., Boonman,A.M., de Jong,G.L., van Noort,,W.L., De Schryver,J., Duran,M., van den Berg,I.E., Gerwig,G.J., Berger,R. and Poll-The,B.T. (1998) A novel disorder of N-glycosylation due to phosphomannose isomerase deficiency. Biochem. Biophys. Res. Commun., 245, 3842.[ISI][Medline]
Eckert,V., Blank,M., Mazhari-Tabrizi,R., Mumberg,D., Funk,M. and Schwarz,R.T. (1998) Cloning and functional expression of the human GlcNAc-1-P transferase, the enzyme for the committed step of the dolichol-cycle by heterologous complementation in Saccharomyces cerevisiae. Glycobiology, 8, 7785.
Herscovics,A. and Orlean,P. (1993) Glycoprotein biosynthesis in yeast. FASEB J., 7, 540550.
Hirschberg,C.B. and Snider,M.D. (1987) Topography of glycosylation in the rough endoplasmic reticulum and Golgi apparatus. Annu. Rev. Biochem., 56, 6387.[ISI][Medline]
Huffaker,T.C. and Robbins,P.W. (1982) Temperature-sensitive yeast mutants deficient in asparagine-linked glycosylation. J. Biol. Chem., 257, 32033210.
Huffaker,T.C. and Robbins,P.W. (1983) Yeast mutants deficient in protein glycosylation. Proc. Natl. Acad. Sci. USA, 80, 74667470.[Abstract]
Ito,H., Fukuda,Y., Murata,K. and Kimura,A. (1983) Transformation of intact yeast cells treated with alkali cations. J. Bacteriol., 153, 163168.[ISI][Medline]
Jackson,B.J., Kukuruzinska,M.A. and Robbins,P.W. (1993) Biosynthesis of asparagine-linked oligosaccharides in Saccharomyces cerevisiae: the alg2 mutation. Glycobiology, 3, 357364.[Abstract]
Jaeken,J., Schachter,H., Carchon,H., De Cock,P., Coddeville,B. and Spik,G. (1994) Carbohydrate deficient glycoprotein syndrome type II: a deficiency in Golgi localised N-acetylglucosaminyltransferase II. Arch. Dis. Child., 71, 123127.[Abstract]
Kataoka,K., Takahashi,T. and Nishikawa,Y. (1997) Sequence analysis of a human marker DNA isolated from a human genomic DNA-transformant of a mouse lipid-linked oligosaccharide synthesis mutant, G258, Glycoconjugate J., 14 (Suppl. 1), S35.
Kataoka,K., Takahashi,T., Ayusawa,D. and Nishikawa,Y. (1999) Characterization of a human genomic DNA fragment which rescues defective lipid-linked oligosaccharide synthesis in a mutant G258 cell line isolated from the FM3A mouse mammary carcinoma cell line. Somat. Cell Mol. Genet., 24, 235243.[ISI]
Körner,C., Knauer,R., Holzbach,U., Hanefeld,F., Lehle,L. and von Figura,K. (1998) Carbohydrate-deficient glycoprotein syndrome type V: deficiency of dolichyl-P-Glc:Man9GlcNAc2-PP-dolichyl glucosyltransferase. Proc. Natl. Acad. Sci. USA, 95, 1320013205.
Kozak,M. (1984) Compilation and analysis of sequences upstream from the translational start site in eukaryotic mRNAs. Nucleic Acids Res., 12, 857872.[Abstract]
Kukuruzinska,M.A., Bergh,M.L.E. and Jackson,B.J. (1987) Protein glycosylation in yeast. Annu. Rev. Biochem., 56, 915944.[ISI][Medline]
Lee,S.-K., Li,G., Yu,S.-L., Alexander,H. and Alexander,S. (1997) The Dictyostelium discoideum ß-1,4-mannosyltransferase gene, mntA, has two periods of developmental expression. Gene, 204, 251258.[ISI][Medline]
Lennon,K., Pretel,R., Kesselheim,R., te Heesen,S. and Kukuruzinska,M.A. (1995) Proliferation-dependent differential regulation of dolichol pathway genes in Saccharomyces cerevisiae. Glycobiology, 5, 633642.[Abstract]
Niehues,R., Hasilik,M., Alton,G., Körner,C., Schiebe-Sukumar,M., Koch,H.G., Zimmer,K.P., Wu,R., Harms,E., Reiter,K., von Figura,K., Freeze,H.H., Harms,H.K. and Marquardt,T. (1998) Carbohydrate-deficient glycoprotein syndrome type Ib. Phosphomannose isomerase deficiency and mannose therapy. J. Clin. Invest., 101, 14141420.
Nishikawa,Y. (1984) Isolation of a temperature-sensitive FM3A mutant deficient in asparagine-linked glycosylation by selecting for resistance to tritiated mannose suicide. J. Cell Physiol., 119, 260266.[ISI][Medline]
Nishikawa,Y. (1991) An FM3A mutant, G258, with a mutation that affects both cell growth and oligosaccharide-lipid synthesis. Biochim. Biophys. Acta, 1091, 135140.
Nishikawa,Y., Yamamoto,Y., Kaji,K. and Mitsui,H. (1980) Reversible G1 arrest of a human Burkitt lymphoma cell line (Raji) induced by tunicamycin. Biochem. Biophys. Res. Commun., 97, 12961303.[ISI][Medline]
Rajput,B., Ma,J., Muniappa,N., Schantz,L., Naylor,S.L., Lalley,P.A. and Vijay,I.K. (1992) Mouse UDP-GlcNAc: dolichyl-phosphate N-acetylglucosaminephosphotransferase. Molecular cloning of the cDNA, generation of anti-peptide antibodies and chromosomal localization. Biochem. J., 285, 985992.[ISI][Medline]
Reiss,G., te Heesen,S., Zimmerman,J., Robbins,P.W. and Aebi,M. (1996) Isolation of the ALG6 locus of Saccharomyces cerevisiae required for glucosylation in the N-linked glycosylation pathway. Glycobiology, 6, 493498.[Abstract]
Rine,J., Hansen,W., Hardeman,E. and Davis,R.W. (1983) Targeted selection of recombinant clones through gene dosage effects. Proc. Natl. Acad. Sci. USA, 80, 67506754.[Abstract]
Sambrook,J., Fritsch,E.F. and Maniatis,T. (1989) Molecular Cloning: A Laboratory Manual. Second edition. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
Sanger,F., Nicklen,S. and Coulson,A.R. (1977) DNA sequencing with chain-terminating inhibitors. Proc. Natl. Acad. Sci. USA, 74, 54635467.[Abstract]
Scocca,J.R. and Krag,S.S. (1990) Sequence of a cDNA that specifies the uridine diphosphate N-acetyl-D-glucosamine:dolichol phosphate N-acetylglucosamine-1-phosphate transferase from Chinese hamster ovary cells. J. Biol. Chem., 265, 2062120626.
Snider,M.D. and Robbins,P.W. (1982) Transmembrane organization of protein glycosylation. Mature oligosaccharide-lipid is located on the luminal side of microsomes from Chinese hamster ovary cells. J. Biol. Chem., 257, 67966801.
Stagljar,I., te Heesen,S. and Aebi,M. (1994) New phenotype of mutants deficient in glucosylation of the lipid-linked oligosaccharide: cloning of the ALG8 locus. Proc. Natl. Acad. Sci. USA, 91, 59775981.[Abstract]
Takahashi,T., Mori,H. and Nishikawa,Y. (1997) Cloning of putative human mannosyltransferase genes which are involved in lipid-linked oligosaccharide synthesis. Glycocojugate J., 14 (Suppl. 1), S35.
Tan,J., Dunn,J., Jaeken,J. and Schachter,H. (1996) Mutations in the MGAT2 gene controlling complex N-glycan synthesis cause carbohydrate-deficient glycoprotein syndrome type II, an autosomal recessive disease with defective brain development. Am. J. Hum. Genet., 59, 810817.[ISI][Medline]
Van Schaftingen,E. and Jaeken,J. (1995) Phosphomannomutase deficiency is a cause of carbohydrate-deficient glycoprotein syndrome type I. FEBS Lett., 377, 318320.[ISI][Medline]
Vieira,J. and Messing,J. (1987) Production of single-stranded plasmid DNA. Methods Enzymol., 153, 311.[ISI][Medline]
Wilson,R., Ainscough,R., Anderson,K., Baynes,C., Berks,M., Bonfield,J., Burton,J., Connell,M., Copsey,T., Cooper,J., et al. (1994) 2.2 Mb of contiguous nucleotide sequence from chromosome III of C. elegans. Nature, 368, 3238.[ISI][Medline]
Yoko-o,T., Matsui,Y., Yagisawa,H., Nojima,H., Uno,I. and Toh-e,A. (1993) The putative phosphoinositide-specific phospholipase C gene, PLC1, of the yeast Saccharomyces cerevisiae is important for cell growth. Proc. Natl. Acad. Sci. USA, 90, 18041808.[Abstract]
Zhu,X. and Lehrman,M.A. (1990) Cloning, sequence, and expression of a cDNA encoding hamster UDP-GlcNAc:dolichol phosphate N-acetylglucosamine-1-phosphate transferase. J. Biol. Chem., 265, 1425014255.