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Research article
Ranjana Arya
Jawaharlal Nehru University
Corresponding Author
ORCiD: https://orcid.org/0000-0001-5924-9351
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UDP-N-acetyl glucosamine-2 epimerase/ N-acetyl mannosamine kinase (GNE) catalyzes key enzymatic reactions in the biosynthesis of sialic acid. Mutation in GNE gene causes GNE-Myopathy characterized by adult onset muscle weakness and degeneration. GNE is involved in different cellular processes like adhesion, apoptosis, ER stress and autophagy. Lack of appropriate model system limits drug and treatment options for GNE Myopathy as GNE knock out was found to be embryonically lethal. In the present study, we have generated L6 rat skeletal muscle cell-based model system for GNE Myopathy where GNE gene is knocked out at exon 3 using AAV mediated SEPT homology recombination. The cell line was heterozygous for GNE gene with one wild type and one truncated allele as confirmed by sequencing. The phenotype showed reduced GNE epimerase activity with little reduction in sialic acid content. In addition, the GNE knock out cell line revealed altered cytoskeletal organization with disrupted actin filament. The signaling cascade regulating actin dynamics such as Rho A, Cofilin, FAK and Src were altered leading to reduced cell migration in GNE heterozygous cells. Our study indicates possible role of GNE in regulating actin dynamics and cell migration of skeletal muscle cell. The skeletal muscle cell based system offers great potential in understanding pathomechanism and target identification for GNE Myopathy.
Keywords
Sialylation, L6 myoblast, SEPT homology recombination, GNE myopathy,actin dynamics
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A new preprint design is coming!
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Ranjana Arya
Jawaharlal Nehru University
Corresponding Author
ORCiD: https://orcid.org/0000-0001-5924-9351
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 30 Jun, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
License:
This work is licensed under a CC BY 4.0 License. Read Full License
UDP-N-acetyl glucosamine-2 epimerase/ N-acetyl mannosamine kinase (GNE) catalyzes key enzymatic reactions in the biosynthesis of sialic acid. Mutation in GNE gene causes GNE-Myopathy characterized by adult onset muscle weakness and degeneration. GNE is involved in different cellular processes like adhesion, apoptosis, ER stress and autophagy. Lack of appropriate model system limits drug and treatment options for GNE Myopathy as GNE knock out was found to be embryonically lethal. In the present study, we have generated L6 rat skeletal muscle cell-based model system for GNE Myopathy where GNE gene is knocked out at exon 3 using AAV mediated SEPT homology recombination. The cell line was heterozygous for GNE gene with one wild type and one truncated allele as confirmed by sequencing. The phenotype showed reduced GNE epimerase activity with little reduction in sialic acid content. In addition, the GNE knock out cell line revealed altered cytoskeletal organization with disrupted actin filament. The signaling cascade regulating actin dynamics such as Rho A, Cofilin, FAK and Src were altered leading to reduced cell migration in GNE heterozygous cells. Our study indicates possible role of GNE in regulating actin dynamics and cell migration of skeletal muscle cell. The skeletal muscle cell based system offers great potential in understanding pathomechanism and target identification for GNE Myopathy.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
This is a list of supplementary files associated with this preprint. Click to download.
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