Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II

Claudia I Robledo-Ortiz, Arturo Flores-Carreón, Arturo Hernández-Cervantes, Aurelio Álvarez-Vargas, Keunsook K Lee, Diana F Díaz-Jiménez, Carol A Munro, Carmen Cano-Canchola, Héctor M Mora-Montes

Research output: Contribution to journalArticle

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Abstract

The N-linked glycosylation is a ubiquitous protein modification in eukaryotic cells. During the N-linked glycan synthesis, the precursor Glc(3)Man(9)GlcNAc(2) is processed by endoplasmic reticulum (ER) glucosidases I, II and α1,2-mannosidase, before transporting to the Golgi complex for further structure modifications. In fungi of medical relevance, as Candida albicans and Aspergillus, it is well known that ER glycosidases are important for cell fitness, cell wall organization, virulence, and interaction with the immune system. Despite this, little is known about these enzymes in Sporothrix schenckii, the causative agent of human sporotrichosis. This limited knowledge is due in part to the lack of a genome sequence of this organism. In this work we used degenerate primers and inverse PCR approaches to isolate the open reading frame of S. schenckii ROT2, the encoding gene for α subunit of ER glucosidase II. This S. schenckii gene complemented a Saccharomyces cerevisiae rot2Δ mutant; however, when expressed in a C. albicans rot2Δ mutant, S. schenckii Rot2 partially increased the levels of α-glucosidase activity, but failed to restore the N-linked glycosylation defect associated to the mutation. To our knowledge, this is the first report where a gene involved in protein N-linked glycosylation is isolated from S. schenckii.

Original languageEnglish
Pages (from-to)910-918
Number of pages9
JournalFungal Biology
Volume116
Issue number8
DOIs
Publication statusPublished - Aug 2012

Fingerprint

Sporothrix
Sporothrix schenckii
glucosidases
Glycosylation
Endoplasmic Reticulum
endoplasmic reticulum
Candida albicans
glycosylation
gene
Mannosidases
Sporotrichosis
Glucosidases
Genes
protein
genes
Glycoside Hydrolases
Golgi Apparatus
Eukaryotic Cells
immune system
Aspergillus

Keywords

  • amino acid sequence
  • endoplasmic reticulum
  • fungal proteins
  • glycosylation
  • humans
  • molecular sequence data
  • sequence alignment
  • sporothrix
  • alpha-glucosidases

Cite this

Robledo-Ortiz, C. I., Flores-Carreón, A., Hernández-Cervantes, A., Álvarez-Vargas, A., Lee, K. K., Díaz-Jiménez, D. F., ... Mora-Montes, H. M. (2012). Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II. Fungal Biology, 116(8), 910-918. https://doi.org/10.1016/j.funbio.2012.06.002

Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II. / Robledo-Ortiz, Claudia I; Flores-Carreón, Arturo; Hernández-Cervantes, Arturo; Álvarez-Vargas, Aurelio; Lee, Keunsook K; Díaz-Jiménez, Diana F; Munro, Carol A; Cano-Canchola, Carmen; Mora-Montes, Héctor M.

In: Fungal Biology, Vol. 116, No. 8, 08.2012, p. 910-918.

Research output: Contribution to journalArticle

Robledo-Ortiz, CI, Flores-Carreón, A, Hernández-Cervantes, A, Álvarez-Vargas, A, Lee, KK, Díaz-Jiménez, DF, Munro, CA, Cano-Canchola, C & Mora-Montes, HM 2012, 'Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II', Fungal Biology, vol. 116, no. 8, pp. 910-918. https://doi.org/10.1016/j.funbio.2012.06.002
Robledo-Ortiz CI, Flores-Carreón A, Hernández-Cervantes A, Álvarez-Vargas A, Lee KK, Díaz-Jiménez DF et al. Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II. Fungal Biology. 2012 Aug;116(8):910-918. https://doi.org/10.1016/j.funbio.2012.06.002
Robledo-Ortiz, Claudia I ; Flores-Carreón, Arturo ; Hernández-Cervantes, Arturo ; Álvarez-Vargas, Aurelio ; Lee, Keunsook K ; Díaz-Jiménez, Diana F ; Munro, Carol A ; Cano-Canchola, Carmen ; Mora-Montes, Héctor M. / Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II. In: Fungal Biology. 2012 ; Vol. 116, No. 8. pp. 910-918.
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AU - Flores-Carreón, Arturo

AU - Hernández-Cervantes, Arturo

AU - Álvarez-Vargas, Aurelio

AU - Lee, Keunsook K

AU - Díaz-Jiménez, Diana F

AU - Munro, Carol A

AU - Cano-Canchola, Carmen

AU - Mora-Montes, Héctor M

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N2 - The N-linked glycosylation is a ubiquitous protein modification in eukaryotic cells. During the N-linked glycan synthesis, the precursor Glc(3)Man(9)GlcNAc(2) is processed by endoplasmic reticulum (ER) glucosidases I, II and α1,2-mannosidase, before transporting to the Golgi complex for further structure modifications. In fungi of medical relevance, as Candida albicans and Aspergillus, it is well known that ER glycosidases are important for cell fitness, cell wall organization, virulence, and interaction with the immune system. Despite this, little is known about these enzymes in Sporothrix schenckii, the causative agent of human sporotrichosis. This limited knowledge is due in part to the lack of a genome sequence of this organism. In this work we used degenerate primers and inverse PCR approaches to isolate the open reading frame of S. schenckii ROT2, the encoding gene for α subunit of ER glucosidase II. This S. schenckii gene complemented a Saccharomyces cerevisiae rot2Δ mutant; however, when expressed in a C. albicans rot2Δ mutant, S. schenckii Rot2 partially increased the levels of α-glucosidase activity, but failed to restore the N-linked glycosylation defect associated to the mutation. To our knowledge, this is the first report where a gene involved in protein N-linked glycosylation is isolated from S. schenckii.

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