Ablation of mouse phosphomannose isomerase (Mpi) causes mannose 6-phosphate accumulation, toxicity, and embryonic lethality.
MPI encodes phosphomannose isomerase, which interconverts fructose 6-phosphate and mannose 6-phosphate (Man-6-P), used for glycoconjugate biosynthesis. MPI mutations in humans impair protein glycosylation causing congenital disorder of glycosylation Ib (CDG-Ib), but oral mannose supplements normalize glycosylation. To establish a mannose-responsive mouse model for CDG-Ib, we ablated Mpi and provided dams with mannose to rescue the anticipated defective glycosylation. Surprisingly, although glycosylation was normal, Mpi(-/-) embryos died around E11.5. Mannose supplementation even hastened their death, suggesting that man-nose was toxic. Mpi(-/-) embryos showed growth retardation and placental hyperplasia. More than 90% of Mpi(-/-) embryos failed to form yolk sac vasculature, and 35% failed chorioallantoic fusion. We generated primary embryonic fibroblasts to investigate the mechanisms leading to embryonic lethality and found that mannose caused a concentration- and time-dependent accumulation of Man 6-P in Mpi(-/-) fibroblasts. In parallel, ATP decreased by more than 70% after 24 h compared with Mpi(+/+) controls. In cell lysates, Man-6-P inhibited hexokinase (70%), phosphoglucose isomerase (65%), and glucose-6-phosphate dehydrogenase (85%), but not phosphofructokinase. Incubating intact Mpi(-/-) fibroblasts with 2-[(3)H]deoxyglucose confirmed mannose-dependent hexokinase inhibition. Our results in vitro suggest that mannose toxicity in Mpi(-/-) embryos is caused by Man-6-P accumulation, which inhibits glucose metabolism and depletes intracellular ATP. This was confirmed in E10.5 Mpi(-/-) embryos where Man-6-P increased more than 10 times, and ATP decreased by 50% compared with Mpi(+/+) littermates. Because Mpi ablation is embryonic lethal, a murine CDG-Ib model will require hypomorphic Mpi alleles.
DeRossi
Charles
C
Glycobiology and Carbohydrate Chemistry Program, Burnham Institute for Medical Research, 10901 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Bode
Lars
L
Eklund
Erik A
EA
Zhang
Fangrong
F
Davis
Joseph A
JA
Westphal
Vibeke
V
Wang
Ling
L
Borowsky
Alexander D
AD
Freeze
Hudson H
HH
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002239:Carbohydrate Metabolism, Inborn Errors; D002478:Cells, Cultured; D020964:Embryo Loss; D004622:Embryo, Mammalian / Q000033:anatomy & histology / Q000473:pathology / Q000502:physiology*; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D018390:Gene Targeting; D005838:Genotype; D005865:Gestational Age; D006593:Hexokinase / Q000378:metabolism; D006801:Humans; D008297:Male; D008358:Mannose / Q000008:administration & dosage / Q000378:metabolism* / Q000633:toxicity; D008359:Mannose-6-Phosphate Isomerase / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D008360:Mannosephosphates / Q000378:metabolism / Q000633:toxicity; D051379:Mice; D018345:Mice, Knockout; D011134:Polysaccharides / Q000096:biosynthesis; D011247:Pregnancy
D008360:Mannosephosphates; D011134:Polysaccharides; C027693:mannose-6-phosphate; D000255:Adenosine Triphosphate; D006593:Hexokinase; D008359:Mannose-6-Phosphate Isomerase; D008358:Mannose