Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos.
Mucin-type O-glycosylation is a ubiquitous posttranslational modification in which N-Acetylgalactosamine (GalNAc) is added to the hydroxyl group of select serine or threonine residues of a protein by the family of UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferases (GalNAc-Ts; EC 2.4.1.41). Pr...
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Public Library of Science (PLoS)
2017-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC5407767?pdf=render |
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| author | Amber L Famiglietti Zheng Wei Thomas M Beres Adina L Milac Duy T Tran Divya Patel Robert C Angerer Lynne M Angerer Lawrence A Tabak |
| author_facet | Amber L Famiglietti Zheng Wei Thomas M Beres Adina L Milac Duy T Tran Divya Patel Robert C Angerer Lynne M Angerer Lawrence A Tabak |
| author_sort | Amber L Famiglietti |
| collection | DOAJ |
| description | Mucin-type O-glycosylation is a ubiquitous posttranslational modification in which N-Acetylgalactosamine (GalNAc) is added to the hydroxyl group of select serine or threonine residues of a protein by the family of UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferases (GalNAc-Ts; EC 2.4.1.41). Previous studies demonstrate that O-glycosylation plays essential roles in protein function, cell-cell interactions, cell polarity and differentiation in developing mouse and Drosophila embryos. Although this type of protein modification is highly conserved among higher eukaryotes, little is known about this family of enzymes in echinoderms, basal deuterostome relatives of the chordates. To investigate the potential role of GalNAc-Ts in echinoderms, we have begun the characterization of this enzyme family in the purple sea urchin, S. purpuratus. We have fully or partially cloned a total of 13 genes (SpGalnts) encoding putative sea urchin SpGalNAc-Ts, and have confirmed enzymatic activity of five recombinant proteins. Amino acid alignments revealed high sequence similarity among sea urchin and mammalian glycosyltransferases, suggesting the presence of putative orthologues. Structural models underscored these similarities and helped reconcile some of the substrate preferences observed. Temporal and spatial expression of SpGalnt transcripts, was studied by whole-mount in situ hybridization. We found that many of these genes are transcribed early in developing embryos, often with restricted expression to the endomesodermal region. Multicolor fluorescent in situ hybridization (FISH) demonstrated that transcripts encoding SpGalnt7-2 co-localized with both Endo16 (a gene expressed in the endoderm), and Gcm (a gene expressed in secondary mesenchyme cells) at the early blastula stage, 20 hours post fertilization (hpf). At late blastula stage (28 hpf), SpGalnt7-2 message co-expresses with Gcm, suggesting that it may play a role in secondary mesenchyme development. We also discovered that morpholino-mediated knockdown of SpGalnt13 transcripts, results in a deficiency of embryonic skeleton and neurons, suggesting that mucin-type O-glycans play essential roles during embryonic development in S. purpuratus. |
| first_indexed | 2024-12-10T16:45:50Z |
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| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-10T16:45:50Z |
| publishDate | 2017-01-01 |
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| spelling | doaj.art-009a4e20b2db4cb4a328fa9cb7d6b44d2022-12-22T01:41:03ZengPublic Library of Science (PLoS)PLoS ONE1932-62032017-01-01124e017647910.1371/journal.pone.0176479Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos.Amber L FamigliettiZheng WeiThomas M BeresAdina L MilacDuy T TranDivya PatelRobert C AngererLynne M AngererLawrence A TabakMucin-type O-glycosylation is a ubiquitous posttranslational modification in which N-Acetylgalactosamine (GalNAc) is added to the hydroxyl group of select serine or threonine residues of a protein by the family of UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferases (GalNAc-Ts; EC 2.4.1.41). Previous studies demonstrate that O-glycosylation plays essential roles in protein function, cell-cell interactions, cell polarity and differentiation in developing mouse and Drosophila embryos. Although this type of protein modification is highly conserved among higher eukaryotes, little is known about this family of enzymes in echinoderms, basal deuterostome relatives of the chordates. To investigate the potential role of GalNAc-Ts in echinoderms, we have begun the characterization of this enzyme family in the purple sea urchin, S. purpuratus. We have fully or partially cloned a total of 13 genes (SpGalnts) encoding putative sea urchin SpGalNAc-Ts, and have confirmed enzymatic activity of five recombinant proteins. Amino acid alignments revealed high sequence similarity among sea urchin and mammalian glycosyltransferases, suggesting the presence of putative orthologues. Structural models underscored these similarities and helped reconcile some of the substrate preferences observed. Temporal and spatial expression of SpGalnt transcripts, was studied by whole-mount in situ hybridization. We found that many of these genes are transcribed early in developing embryos, often with restricted expression to the endomesodermal region. Multicolor fluorescent in situ hybridization (FISH) demonstrated that transcripts encoding SpGalnt7-2 co-localized with both Endo16 (a gene expressed in the endoderm), and Gcm (a gene expressed in secondary mesenchyme cells) at the early blastula stage, 20 hours post fertilization (hpf). At late blastula stage (28 hpf), SpGalnt7-2 message co-expresses with Gcm, suggesting that it may play a role in secondary mesenchyme development. We also discovered that morpholino-mediated knockdown of SpGalnt13 transcripts, results in a deficiency of embryonic skeleton and neurons, suggesting that mucin-type O-glycans play essential roles during embryonic development in S. purpuratus.http://europepmc.org/articles/PMC5407767?pdf=render |
| spellingShingle | Amber L Famiglietti Zheng Wei Thomas M Beres Adina L Milac Duy T Tran Divya Patel Robert C Angerer Lynne M Angerer Lawrence A Tabak Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. PLoS ONE |
| title | Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. |
| title_full | Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. |
| title_fullStr | Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. |
| title_full_unstemmed | Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. |
| title_short | Characterization and expression analysis of Galnts in developing Strongylocentrotus purpuratus embryos. |
| title_sort | characterization and expression analysis of galnts in developing strongylocentrotus purpuratus embryos |
| url | http://europepmc.org/articles/PMC5407767?pdf=render |
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