DPAGT1-CDG

What is DPAGT1?

DPAGT1 is the name of a gene that encodes an enzyme with the unwieldy name UDP-N-acetylglucosamine—dolichyl-phosphate N-acetylglucosaminephosphotransferase. 

DPAGT1 and the ALG13/ALG14 duo create a foundational dolichol-linked N-acetylglucosamine (abbreviated GlcNAc) disaccharide that serves as the substrate for the first mannosyltransferase ALG1 in the assembly line of ER-membrane anchored glucosyltransferase. That description understates DPAGT1’s role as the glucosyltransferase that catalyzes the first committed step of N-linked protein glycosylation. The business end of DPAGT1 is located in cytoplasm, which is also true for the next two glucosyltransferases down the line, ALG13 and ALG14.

Orthologs of DPAGT1 are conserved in mice, zebrafish, flies, worms, budding yeast and fission yeast.

What does DPAGT1 do?

DPAGT1 takes UDP-N-acetylglucosamine and dolichyl phosphate and turns them into UMP and dolichol phosphate N-acetylglucosamine-1-phosphate (also known as GlcNAc-P-dolichol). As a glucosyltransferase, DPAGT1 adds the first N-acetylglucosamine building block to the lipid carrier dolichol, setting the stage for all further monosaccharide additions. DPAGT1 is in fact the rate-limiting step in the formation of lipid-linked oligosaccharides (LLOs).

DPAGT1 regulates and is regulated by the Wnt/beta-catenin signaling pathway, which is involved in development and cancer (Varelas et al., 2014). DPAGT1 is also the target of the natural product and widely used chemical probe tunicamycin (Yoo et al., 2018).

Clinical presentation of DPAGT1-CDG

DPAGT1-CDG is formerly known as CDG-1j and after PMM2-CDG is one of the more common observed CDGs – but still rare in terms of patient numbers.

Wu et al., 2003 is the first published clinical report of DPAGT1-CDG. Diagnostic criteria for DPAGT1-CDG include pronounced hypotonia, intractable seizures, intellectual disability, microcephaly, exotropia (misalignment of the eyes) and premature death. The most recent clinical report by Ng et al., 2019 brought the total number of diagnosed DPAGT1-CDG patients to 39. They also described several adult patients with a mild presentation of DPAGT1-CDG.

Unexpectedly, mutations in the DPAGT1 gene were shown to be cause limb-girdle congenital myasthenic syndrome with tubular aggregates (Belaya et al., 2012). Congenital myasthenic syndrome is a neuromuscular disorder characterized by muscle weakness and fatigue. This result was confirmed by multiple groups since. The neuromuscular synapse where neurons makes connections to muscles is particularly sensitive to loss of DPAGT1 because synaptic glycoproteins mediate cell-to-cell contact and adhesion. 

Variable clinical presentation from patient to patient is the norm in CDGs. Kane et al., 2016 provide an interesting explanation which applies to DPAGT1-CDG: “Here we have provided evidence of mitotic intragenic recombination in compound heterozygous CDGs. This suggests that recombination between compound heterozygous alleles of glycosylation genes causes reversion to a wild-type allele and that survival or growth advantage of cells with the wild-type allele ameliorates the phenotype of the CDGs. A model of CDG survival attributed to mitotic intragenic recombination within somatic cells is further supported by residual enzymatic activity in CDG individual cells, and the small number of reported homozygous mutations among the CDGs.”

In other words, since most CDG patients inherit two different loss-of-function mutations from each parent, the process call mitotic recombination occasionally results in correction of one of the inherited mutations, and the cells in which this correction occurs have a growth advantage over sister cells in which mitotic recombination doesn’t occur.

DPAGT1-CDG’s unexpected role in cancer

Human salivary carcinoma cells express DPAGT1 at higher levels than usual according to Nita-Lazar et al., 2009. Sengupta et al., 2010 showed that the Wnt/beta-catenin signaling pathway regulates DPAGT1 expression, which may explain the role DPAGT1 in driving oral cancer.

Next steps toward the clinic

Yeast DPAG1-CDG models have been described for use in functional studies but no one has taken the next step and used them for drug screens. A worm model of DPAGT1-CDG has been de-risked by Kanaki et al., 2019. Fly and zebrafish DPAGT1-CDG disease models are low hanging fruit. It’s not clear if any attempts at making at DPAGT1 knockout mouse have been successful but chances are a null mutant would be inviable. So a DPAGT1 patient mutation knock-in mouse or some kind of conditional DPAGT1 knockout mouse will be necessary to test the promising therapeutic candidates.

As is almost always the case in CDGs, DPAGT1-CDG patient fibroblasts have already been validated. From there drug repurposing screens offer the fastest and cheapest path to the first approved treatment for DPAGT1-CDG. The goal is to identify a bridge therapy that buys time for gene therapy and gene editing technologies to mature. In the case of DPAGT1 splice mutations, personalized antisense oligonucleotides (ASOs) offer hope.

References

Belaya K, Finlayson S, Slater CR, Cossins J, Liu WW, Maxwell S, McGowan SJ, Maslau S, Twigg SR, Walls TJ, Pascual Pascual SI, Palace J, Beeson D. (2012). Mutations in DPAGT1 cause a limb-girdle congenital myasthenic syndrome with tubular aggregates. American Journal of Human Genetics. 91: 193-201.

Kanaki N, Matsuda A, Dejima K, Murata D, Nomura KH, Ohkura T, Gengyo-Ando K, Yoshina S, Mitani S, Nomura K. (2019). UDP-N-acetylglucosamine-dolichyl-phosphate N-acetylglucosaminephosphotransferase is indispensable for oogenesis, oocyte-to-embryo transition, and larval development of the nematode Caenorhabditis elegans. Glycobiology. 29: 163-178.

Kane MS, Davids M, Adams C, Wolfe LA, Cheung HW, Gropman A, Huang Y; NISC Comparative Sequencing Program, Ng BG, Freeze HH, Adams DR, Gahl WA, Boerkoel CF. (2016). Mitotic Intragenic Recombination: A Mechanism of Survival for Several Congenital Disorders of Glycosylation. American Journal of Human Genetics. 98: 339-346.

Ng BG, Underhill HR, Palm L, Bengtson P, Rozet JM, Gerber S, Munnich A, Zanlonghi X, Stevens CA, Kircher M, Nickerson DA, Buckingham KJ, Josephson KD, Shendure J, Bamshad MJ; University of Washington Center for Mendelian Genomics, Freeze HH, Eklund EA. (2019). DPAGT1 Deficiency with Encephalopathy (DPAGT1-CDG): Clinical and Genetic Description of 11 New Patients. JIMD Reports. 44: 85-92.

Nita-Lazar M, Noonan V, Rebustini I, Walker J, Menko AS, Kukuruzinska MA. (2009). Overexpression of DPAGT1 leads to aberrant N-glycosylation of E-cadherin and cellular discohesion in oral cancer. Cancer Research. 69: 5673-5680.

Sengupta PK, Bouchie MP, Kukuruzinska MA. (2010). N-glycosylation gene DPAGT1 is a target of the Wnt/beta-catenin signaling pathway. Journal of Biological Chemistry. 285: 31164-73.

Varelas X, Bouchie MP, Kukuruzinska MA. (2014). Protein N-glycosylation in oral cancer: dysregulated cellular networks among DPAGT1, E-cadherin adhesion and canonical Wnt signaling. Glycobiology. 24: 579-91.

Wu X, Rush JS, Karaoglu D, Krasnewich D, Lubinsky MS, Waechter CJ, Gilmore R, Freeze HH. (2003). Deficiency of UDP-GlcNAc:Dolichol Phosphate N-Acetylglucosamine-1 Phosphate Transferase (DPAGT1) causes a novel congenital disorder of Glycosylation Type Ij. Human Mutations. 22: 144-150.

Yoo J, Mashalidis EH, Kuk ACY, Yamamoto K, Kaeser B, Ichikawa S, Lee SY. (2018). GlcNAc-1-P-transferase-tunicamycin complex structure reveals basis for inhibition of N-glycosylation. Nature Structural Molecular Biology. 25: 217-224.

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