title: A zebrafish model of PMM2-CDG reveals altered neurogenesis and a substrate-accumulation mechanism for N-linked glycosylation deficiency.
abstract: Congenital disorder of glycosylation (PMM2-CDG) results from mutations in pmm2, which encodes the phosphomannomutase (Pmm) that converts mannose-6-phosphate (M6P) to mannose-1-phosphate (M1P). Patients have wide-spectrum clinical abnormalities associated with impaired protein N-glycosylation. Although it has been widely proposed that Pmm2 deficiency depletes M1P, a precursor of GDP-mannose, and consequently suppresses lipid-linked oligosaccharide (LLO) levels needed for N-glycosylation, these deficiencies have not been demonstrated in patients or any animal model. Here we report a morpholino-based PMM2-CDG model in zebrafish. Morphant embryos had developmental abnormalities consistent with PMM2-CDG patients, including craniofacial defects and impaired motility associated with altered motor neurogenesis within the spinal cord. Significantly, global N-linked glycosylation and LLO levels were reduced in pmm2 morphants. Although M1P and GDP-mannose were below reliable detection/quantification limits, Pmm2 depletion unexpectedly caused accumulation of M6P, shown earlier to promote LLO cleavage in vitro. In pmm2 morphants, the free glycan by-products of LLO cleavage increased nearly twofold. Suppression of the M6P-synthesizing enzyme mannose phosphate isomerase within the pmm2 background normalized M6P levels and certain aspects of the craniofacial phenotype and abrogated pmm2-dependent LLO cleavage. In summary, we report the first zebrafish model of PMM2-CDG and uncover novel cellular insights not possible with other systems, including an M6P accumulation mechanism for underglycosylation.
authors.affiliation: Complex Carbohydrate Research Center, University of Georgia, Athens, GA 30602, USA.
authors.forename: Abigail
authors.initials: A
authors.lastname: Cline
authors.forename: Ningguo
authors.initials: N
authors.lastname: Gao
authors.forename: Heather
authors.initials: H
authors.lastname: Flanagan-Steet
authors.forename: Vandana
authors.initials: V
authors.lastname: Sharma
authors.forename: Sabrina
authors.initials: S
authors.lastname: Rosa
authors.forename: Roberto
authors.initials: R
authors.lastname: Sonon
authors.forename: Parastoo
authors.initials: P
authors.lastname: Azadi
authors.forename: Kirsten C
authors.initials: KC
authors.lastname: Sadler
authors.forename: Hudson H
authors.initials: HH
authors.lastname: Freeze
authors.forename: Mark A
authors.initials: MA
authors.lastname: Lehrman
authors.forename: Richard
authors.initials: R
authors.lastname: Steet
mesh_terms: D000818:Animals; D002356:Cartilage / Q000187:drug effects / Q000196:embryology / Q000473:pathology; D048430:Cell Shape / Q000187:drug effects; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D018981:Congenital Disorders of Glycosylation / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D019465:Craniofacial Abnormalities / Q000196:embryology / Q000473:pathology; D004195:Disease Models, Animal; D004625:Embryo, Nonmammalian / Q000002:abnormalities / Q000187:drug effects / Q000201:enzymology; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006031:Glycosylation / Q000187:drug effects; D008070:Lipopolysaccharides / Q000378:metabolism; D008359:Mannose-6-Phosphate Isomerase / Q000378:metabolism; D008360:Mannosephosphates / Q000378:metabolism; D060172:Morpholinos / Q000494:pharmacology; D009046:Motor Neurons / Q000187:drug effects / Q000473:pathology; D009068:Movement / Q000187:drug effects; D055495:Neurogenesis / Q000187:drug effects; D017875:Phosphotransferases (Phosphomutases) / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D012886:Skull / Q000002:abnormalities / Q000187:drug effects / Q000196:embryology; D013116:Spinal Cord / Q000187:drug effects / Q000196:embryology / Q000473:pathology; D013379:Substrate Specificity / Q000187:drug effects; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
chemical_list: D008070:Lipopolysaccharides; D008360:Mannosephosphates; D060172:Morpholinos; D029961:Zebrafish Proteins; C023023:lipid-linked oligosaccharides; C027693:mannose-6-phosphate; D008359:Mannose-6-Phosphate Isomerase; D017875:Phosphotransferases (Phosphomutases); C052799:phosphomannomutase