Consequences of disrupting the gene that encodes alpha-glucosidase II in the N-linked oligosaccharide biosynthesis pathway of Dictyostelium discoideum.
We have identified and disrupted the gene coding for alpha-glucosidase II in Dictyostelium discoideum. This enzyme is responsible for removing two alpha 1,3-linked glucose residues from N-linked oligosaccharides on newly synthesized glycoproteins. Mutagenesis by restriction enzyme-mediated integration (REMI) generated a clone, DG1033, which grows well but forms abnormal fruiting bodies with short, thick stalks. The strain lacks alpha-glucosidase II activity and makes incompletely processed N-linked oligosaccharides that are abnormally large and have fewer sulfate and phosphate esters. The morphological, enzymatic, and oligosaccharide profile phenotypes of the disruption mutant are all recapitulated by a targeted disruption of the normal gene. Furthermore, all of these defects are corrected in cells transformed with a normal, full-length copy of the gene. The phenotypic characteristics of DG1033 as well as chromosomal mapping of the disrupted gene indicate that it is the site of the previously characterized modA mutation. The Dictyostelium gene is highly homologous to alpha-glucosidase II genes in the human and the pig, C. elegans, and yeast. Although various cell lines have been reported to be defective in alpha-glucosidase II activity, disruption of the Dictyostelium gene gives the first example of a clear developmental phenotype associated with loss of this enzyme.
Freeze
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HH
Burnham Institute, La Jolla, CA 92037, USA.
Lammertz
M
M
Iranfar
N
N
Fuller
D
D
Panneerselvam
K
K
Loomis
W F
WF
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004023:Dictyostelium / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D017125:Genes, Protozoan; D006801:Humans; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis* / Q000235:genetics; D016415:Sequence Alignment; D017384:Sequence Deletion; D000520:alpha-Glucosidases / Q000235:genetics* / Q000378:metabolism
D009844:Oligosaccharides; C034251:4-nitrophenyl-alpha-glucosidase; D000520:alpha-Glucosidases