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protein-O-mannosyltransferase 1 (POMT1)

(last modified June 17, 2008)

Contents

·        Summary

·        The protein-O-mannosyltransferase 1 gene

·        The protein-O-mannosyltransferase 1 mRNA

o      protein-O-mannosyltransferase 1 expression

·        The protein-O-mannosyltransferase 1 protein

·        Protein-O-mannosyltransferase 1 and disease:

§       Walker-Warburg syndrome

§       Muscular dystrophy, limb-girdle, type 2k

§       Muscle-eye-brain disease

 

o      sequence variations (mutations and polymorphisms)

·        Animal models

·        Miscellaneous

o      Primers

o      The occuring SNPs in POMT1

o      GeneTree

o      HomoloGene

o      Ensembl: different markers

 

Summary

Protein-O-mannosyltransferase 1 was first described by Jurado et al. (1999) as a 3.1 to 3.2 kb mRNA in all tissues tested, with slightly stronger expression in skeletal muscle and heart. Jurado et al. (1999) also reported this by using homology with Drosophila rt, Jurado et al. (1999) to identify an EST for POMT1. The gene for protein-O-mannosyltransferase 1 maps to 9q34.1, spans 20 kb of genomic DNA and contains 20 exons. Protein-O-mannosyltransferase 1 is a 725 amino acid protein and has a calculated molecular mass of about 82.5 kD. POMT1 contains 7 to 12 putative transmembrane regions and a C-terminal ER membrane retention signal. It is primary found in the endoplasmic reticulum. POMT1 shares sequence similarity with protein O-mannosyltransferases of S. cerevisiae. In yeast, these enzymes are located in the endoplasmic reticulum (ER) and are required for cell integrity and cell wall rigidity. POMT1 also shows similarity to the Drosophila 'rotated abdomen' (rt) gene, which when mutated causes defects in myogenesis and muscle structure. There are 3 named isoforms Q9Y6A1-1, Q9Y6A1-2 and Q9Y6A1-3.

The protein-O-mannosyltransferase 1 gene

Links to other databases:
NCBI Entrez Gene Entrez Gene POMT1 OMIM

Protein-O-mannosyltransferase 1 (Gene symbol POMT1, aliases RP11-334J6.2, FLJ37239, LGMD2K, RT) was first described by Jurado et al. (1999) as a 82,5 kD protein with high homology to the yeast mannosyl-transferases (Pmts). In the article they predicted that given the strong conservation of protein motifs between POMT1 and the yeast Pmts, POMT1 may function as a mannosyl-transferase involved in O-mannosylation of proteins, being the first of such a class found in mammals. They were also able to clone the full-length POMT1 cDNA by 5-prime cDNA walking performed by vector/insert PCR, and by anchor PCR of fetal brain RNA. The gene maps to chromosome 9q34.1. The POMT1 locus is flanked by markers D9S260 and D9S7293 on 9q34 (Beltran-Valero de Bernabe et al. (2002)).

The POMT1 geneID = 10585. The gene spans about 20 kB of genomic DNA and contains 20 exons. The initiator ATG is located in exon 2. There are a variable number of 17- to 19-nucleotide tandem repeats within intron 13. Jurado et al. (1999) identified 8 different allelic variants carrying 36 to 56 repeats within normal chromosomes. The 56-repeat allele was the most frequent. Intron 2 also contains a (CA)n microsatellite.

The POMT1 gene is flanked 5’ by the unidentified KIAA0515 and SNORD62A (small nucleolar RNA) gene and 3´ by the UCK1 gene and the hypothetical LOC642515.

 

The following link gives you detailed exon information: Ensembl.

The protein-O-mannosyltransferase 1 mRNA

 

Links to other databases:     RefSeq: NM_007171                  RefSeq: NM_001077365                  RefSeq: NM_001077366                     UniGene: Hs.522449

 

Northern blot analysis revealed a diffuse band of 3.1 to 3.2 kb in all tissues tested, with slightly stronger expression in skeletal muscle and heart. RNA dot blot analysis revealed ubiquitous expression, with maximum levels in testis and high levels in fetal brain and pituitary Jurado et al. (1999). By this method, expression in skeletal muscle and heart was not significantly higher than expression in other tissues. RT-PCR revealed several mRNA splice variants. Southern blot analysis indicated Pomt1 expression in all mammalian DNAs tested, as well as weak but specific signals in bird, reptile, and amphibian DNAs; no signal was detected in fish or plant DNAs Jurado et al. (1999). And determined that the POMT1 gene contains 20 exons and spans about 20 kb. The initiator ATG(see POMT1 - coding DNA reference sequence) is located in exon 2. There are a variable number of 17- to 19-nucleotide tandem repeats within intron 13. Jurado et al. (1999) identified 8 different allelic variants carrying 36 to 56 repeats within normal chromosomes. The 56-repeat allele was the most frequent. Intron 2 also contains a (CA)n microsatellite.

 

POMT1 showed several mRNA variants by reverse transcriptase (RT)-PCR analysis using primers from exons 1 to 5 Jurado et al. (1999). Analysing human and embryonic tissue mRNA through sequence analysis revealed the different mRNA species resulted from alternative splicing, producing the loss of exon 2, exon 3, or exon 4 or the different combinations of those exons in the mature transcripts. The alternative splicing in the 5’ region interrupts the open reading frame in cases and/or deletes the initiator ATG in exon 2.

An a additional splicing variant is created by the use of different donor splice sites for intron 8. This mRNA variant predicts the inclusion of 22 amino acids Jurado et al. (1999) between exons 8 and 9.

 

The protein-O-mannosyltransferase 1 protein

 

Links to other databases: RefSeq: NP_009102.3         RefSeq: NP_001070833.1            RefSeq: NP_001070834.1

POMT1 contains 7 to 12 putative transmembrane regions and a C-terminal ER membrane retention signal. POMT1 shares 40% identity with rt, and it averages 54% similarity with the yeast Pmts.

 

This protein consists of different domains:

http://www.ebi.ac.uk/interpro/IEntry?ac=IPR003608

The MIR domain may have a ligand transferase function. This domain has a closed beta-barrel structure with a hairpin triplet, and has an internal pseudo-threefold symmetry. The MIR motifs that make up the MIR domain consist of ~50 residues and are often found in multiple copies.

 

http://www.ebi.ac.uk/interpro/IEntry?ac=IPR003342

Glycosyl transferase, family 39 The biosynthesis of disaccharides, oligosaccharides and polysaccharides involves the action of hundreds of different glycosyltransferases. These enzymes catalyse the transfer of sugar moieties from activated donor molecules to specific acceptor molecules, forming glycosidic bonds.

 

http://www.ebi.ac.uk/interpro/IEntry?ac=IPR002255

 

There are three isoforms of POMT1, a, b and c, caused by alternative splicing. Additional isoforms seem to exist. Protein-O-mannosyltransferase 1 isoform a consists of 747 amino acids, isoform b of 725 amino acids and isoform c of 671 amino acids.

A tabel with the structural features of POMT1 can be found by using the following link: Expasy. Also information is given about the differences between the three isoforms and the mutations associated with disease.

protein-O-mannosyltransferase 1 (POMT1) and disease

Links to other databases:   OMIM: *607423

·        Walker-Warburg syndrome

·        Muscular dystrophy, limb-girdle, type 2k

·        Muscle-eye-brain disease

Jurado et al. (1999) first reported the identification of an EST for POMT1 out of an isolate from a human gene homologous to Drosophila melanogaster rotated abdomen. The POMT1 locus has been assigned to human chromosome 9q34.1 by somatic cell hybrids, radiation hybrids, and linkage analysis. On the basis of the rt phenotype, POMT1 could be a candidate for uncharacterized genetic disorders of the muscular system, such as some forms of congenital muscular dystrophy or congenital myopathy.

Villanova et al. (2000) reported 4 Italian patients from 3 families affected with an autosomal recessive form of congenital muscular dystrophy. The phenotype was similar in all cases, and was characterized by hypotonia at birth, joint contractures associated with severe psychomotor retardation, inability to walk, striking enlargement of the calf and quadriceps muscles, absent speech, and mental retardation. Cranial MRI showed enlargement of the cisterna magna and cerebellar hypoplasia, without evidence of neuronal migration defects. Muscle biopsy showed changes consistent with muscular dystrophy and also showed a mild to moderate reduction of laminin alpha-2 (LAMA2) and overexpression of laminin alpha-5 (LAMA5). Linkage analysis in the family with 2 affected members excluded linkage to LAMA2 on chromosome 6q22, MEB on 1p34, and FCMD on 9q31.

Beltran-Valero de Bernabe et al. (2002) identified several families in which Walker-Warburg syndrome, a severe recessive congenital muscular dystrophy associated with defects in neuronal migration that produce complex brain and eye abnormalities, was caused by mutation in the POMT1 gene.

Dincer et al. (2003) reported 7 patients from 6 consanguineous Turkish families with autosomal recessive muscular dystrophy and mental retardation. An eighth British patient, who was not from a consanguineous family, had a similar phenotype. All patients acquired early motor milestones, excluding a congenital muscular dystrophy. Age at onset ranged from 1 to 6 years, with difficulty in walking and climbing stairs. Other features included slow progression, proximal muscle weakness, mild muscle hypertrophy, increased serum creatine kinase, microcephaly, and mental retardation (IQ range 50 to 76). Brain imaging was normal in all cases, with no structural abnormalities or white matter changes. Skeletal muscle biopsy showed dystrophic changes, including mild fibrosis with many regenerating and few necrotic fibers, increased fiber size variability, and multiple central nuclei. Immunohistochemical staining showed severe hypoglycosylation of alpha-dystroglycan.

Kim et al. (2004) identified a homozygous 3-bp deletion (1260delCCT) in the POMT1 gene of a Japanese boy with Walker-Warburg syndrome. Resulting in the deletion of a highly conserved leucine at codon 421. The mutation was not identified in 100 Japanese controls. Immunohistochemical studies of skeletal muscle showed hypoglycosylation of alpha-dystroglycan and defective laminin binding.

Balci et al. (2005) identified a homozygous 598G-C transversion in exon 7 of the POMT1 gene, resulting in an ala200-to-pro (A200P) substitution in a highly conserved residue in loop 4 of a cytoplasmic domain of the protein. All patients were born of consanguineous parents. The mutation was not identified in 212 control chromosomes. And noted that A200P was the first reported POMT1 mutation within the cytoplasmic domain and that the phenotype associated with this mutation is significantly milder than Walker-Warburg syndrome, which is caused by other POMT1 mutations. Most significantly, none of the patients with the A200P mutation had structural brain abnormalities on imaging that would signify a cortical migration defect. Haplotype analysis indicated that A200P is a common founder mutation.

van Reeuwijk et al. (2006) identified compound heterozygosity for 2 mutations in the POMT1 gene: a 193G-A transition, resulting in a gly65-to-arg (G65R) substitution, and a 1746G-C transversion, resulting in a trp582-to-cys (W582C) substitution. The G65R substitution occurs within the protein mannosyltransferase (PMT) domain but is not highly conserved, whereas the W582C substitution affects a highly conserved residue in the endoplasmic reticulum domain. van Reeuwijk et al. (2006) suggested that the relatively milder phenotype observed in these patients was due to some residual POMT1 activity. The findings extended the phenotypic spectrum of Walker-Warburg syndrome.

Bouchet et al. (2007) identified mutations in the POMT1 gene in 13 (32%) of 41 families in which at least 1 fetus had severe type II lissencephaly. The minimum diagnostic criteria included hydrocephalus, agyria, thickened leptomeninges filled with neuroglial ectopia, disorganized cortical ribbon, and cerebellar dysplasia. Mutations in the POMGNT1 and POMT2 genes were identified in 6 (15%) and 3 (7%) families, respectively. Overall, mutations were identified in 22 of 41 families included in the study. Definitive pathogenic mutations were not identified in the FKRP, FKTN, or LARGE genes. 

Roberds et al. [1994] first reported the identification of missense variations in both alleles of the gene in a family with late-onset severe chilhood autosomal recessive muscular dystrophy (SCARMD). Later, variations were described in other SCARMD families and in families with limb-girdle muscular dystrophy type (LGMD-2D).

Carrié et al. (1997) reported alpha-sarcoglycan variation screening in a set of 51 unrelated families of widespread geographical origin selected for (1) proximal muscular dystrophy, (2) normal dystrophin, and (3) alpha-sarcoglycan deficiency (i.e. absence or reduced staining ascertained by immunofluorescence and/or Westernblotting). In 20 of these families (39%), variations were found in the alpha-sarcoglycan gene, confirming the observation of Duggan that among sarcoglycanopathies, alpha-sarcolgycan variations are most frequent.

Carrié et al. (1997) reports 25 different variations. In patients, 46% of the chromosomes had variations in exon 3. 229C>T (Arg77Cys) was found on 32% of the chromosomes. mRNA-level and size were normal in all cases except for two where splice site variations resulted in the production of aberrant transcripts. This observation confirms that of Roberds et al. [1994], who suggested that the upto 80-90% reduction of alpha-sarcoglycan in DMD-patients and mdx-mice is likely a post-translational event. All missense variations, except 229C>T (Arg77Cys), resulted in a drastic decrease of alpha-sarcolgycan protein. The phenotype of 15 patients, described by Eymard et al.(1997), shows a large variability, including differences between affected sibs. Without exception, homozygous null variations are responsible for a severe clinical course.

The SGCA:c.229C>T change is the most frequent identified thus far, corresponding to 14-32% of the LGMD2D alleles found in different populations. In Brazil c.229C>T is found associated with at least three distinct haplotypes (Passos-Bueno [1995]). 229C lies in a CpG island and is considered a mutational hot spot causing recurrent mutations at this site.

Mutations associated with LGMD-2D are almost exclusively missense, spread across five exons resulting in amino acid substitutions localised in the extracellular domain of the protein.

protein o-mannosyltransferase 1 sequence variations
(mutations and polymorphisms)

POMT1-diagnosis

Animal models

Willer et al. (2004) found that during embryogenesis, the mouse Pomt1 gene is prominently expressed in the neural tube, the developing eye, and the mesenchyme. They noted that these sites of expression correlate with those in which the main tissue alterations are observed in patients with Walker-Warburg syndrome. Willer et al. (2004) inactivated a Pomt1 allele by gene targeting in mouse embryonic stem cells and produced chimeras transmitting the defect allele to offspring. Although heterozygous mice were viable and fertile, the total absence of homozygous Pomt1 -/- pups among the progeny of heterozygous intercrosses indicated that this genotype is embryonic lethal. Analysis of the mutant phenotype revealed that homozygous null mice suffered developmental arrest around embryonic day (E) 7.5 and died between E7.5 and E9.5. The Pomt1 -/- embryos presented defects in the formation of the Reichert membrane, the first basement membrane to form in the embryo. The Reichert membrane is a thick multilayered membrane between the parietal endoderm cells and the trophoblast cells of rodents; it is thought to function to allow free access of nutrients to the embryo while excluding maternal cells (Salamat et al., 1995). The failure of this membrane to form in the Pomt1 -/- embryos appeared to be the result of abnormal glycosylation and maturation of dystroglycan that may impair recruitment of laminin (see 150320), a structural component required for the formation of Reichert membrane in rodents. Willer et al. (2004) concluded that the targeted disruption of Pomt1 in mouse represents an example of an engineered deletion of a known glycosyltransferase involved in O-mannosyl glycan synthesis.

Miscellaneous

Jurado et al. (1999)

In this article you can find designed primers for POMT1.

The occuring SNPs in POMT1

GeneTree

HomoloGene

Using this link you can see a multiple alignment.

Ensembl: different markers

Different markers for the POMT1 gene are cited.

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