Characterization of the LARGE family of putative glycosyltransferases associated with dystroglycanopathies.
The Large(myd) mouse has a loss-of-function mutation in the putative glycosyltransferase gene Large. Mutations in the human homolog (LARGE) have been described in a form of congenital muscular dystrophy (MDC1D). Other genes (POMT1, POMGnT1, fukutin, and FKRP) that encode known or putative glycosylation enzymes are also causally associated with human congenital muscular dystrophies. All these diseases are associated with hypoglycosylation of the membrane protein alpha-dystroglycan (alpha-DG) and consequent loss of extracellular ligand binding. Hence, they are termed dystroglycanopathies. A paralogous gene for LARGE (LARGE2 or GYLTL1B) may also have a role in DG glycosylation. Using database interrogation and reverse-transcriptase polymerase chain reaction (RT-PCR), we identified vertebrate orthologs of each of these LARGE genes in many vertebrates, including human, mouse, dog, chicken, zebrafish, and pufferfish. However, within invertebrate genomes, we were able to identify only single homologs. We suggest that vertebrate LARGE orthologs be referred to as LARGE1. RT-PCR, dot-blot, and northern analysis indicated that LARGE2 has a more restricted tissue-expression profile than LARGE1. Using epitope-tagged proteins, we show that both LARGE1 and LARGE2 localize to the Golgi apparatus. The high similarity between the LARGE paralogs suggests that LARGE2 may also act on DG. Overexpression of LARGE2 in mouse C2C12 myoblasts results in increased glycosylation of alpha-DG accompanied by an increase in laminin binding. Thus, there may be functional redundancy between LARGE1 and LARGE2. Consistent with this idea, we show that alpha-DG is still fully glycosylated in kidney (a tissue that expresses a high level of LARGE2 mRNA) of Large(myd) mutant mice.
Grewal
Prabhjit K
PK
Institute of Genetics, Queen's Medical Centre, University of Nottingham, Nottingham NG7 2UH, UK.
McLaughlan
Jennifer M
JM
Moore
Christopher J
CJ
Browning
Claudia A
CA
Hewitt
Jane E
JE
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics*; D002460:Cell Line; D002645:Chickens; D004285:Dogs; D049030:Dystroglycans / Q000378:metabolism; D020440:Gene Duplication; D006031:Glycosylation; D016695:Glycosyltransferases / Q000096:biosynthesis / Q000235:genetics*; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D007797:Laminin / Q000378:metabolism; D008565:Membrane Proteins; D051379:Mice; D008969:Molecular Sequence Data; D009136:Muscular Dystrophies / Q000151:congenital / Q000235:genetics; D032446:Myoblasts / Q000166:cytology / Q000378:metabolism; D017351:N-Acetylglucosaminyltransferases / Q000096:biosynthesis / Q000235:genetics*; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics*; D011485:Protein Binding; D013045:Species Specificity; D023721:Tetraodontiformes; D015027:Zebrafish
D002352:Carrier Proteins; D007797:Laminin; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D049030:Dystroglycans; D016695:Glycosyltransferases; C500430:LARGE2 protein, human; C117674:LARGE1 protein, human; C502747:Large1 protein, mouse; C507407:Large2 protein, mouse; D017351:N-Acetylglucosaminyltransferases