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Knock-down of ZBED6 in insulin-producing cells promotes N-cadherin junctions between beta-cells and neural crest stem cells in vitro
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
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2016 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, 19006Article in journal (Refereed) Published
Abstract [en]

The role of the novel transcription factor ZBED6 for the adhesion/clustering of insulin-producing mouse MIN6 and βTC6 cells was investigated. Zbed6-silencing in the insulin producing cells resulted in increased three-dimensional cell-cell clustering and decreased adhesion to mouse laminin and human laminin 511. This was paralleled by a weaker focal adhesion kinase phosphorylation at laminin binding sites. Zbed6-silenced cells expressed less E-cadherin and more N-cadherin at cell-to-cell junctions. A strong ZBED6-binding site close to the N-cadherin gene transcription start site was observed. Three-dimensional clustering in Zbed6-silenced cells was prevented by an N-cadherin neutralizing antibody and by N-cadherin knockdown. Co-culture of neural crest stem cells (NCSCs) with Zbed6-silenced cells, but not with control cells, stimulated the outgrowth of NCSC processes. The cell-to-cell junctions between NCSCs and βTC6 cells stained more intensely for N-cadherin when Zbed6-silenced cells were co-cultured with NCSCs. We conclude that ZBED6 decreases the ratio between N- and E-cadherin. A lower N- to E-cadherin ratio may hamper the formation of three-dimensional beta-cell clusters and cell-to-cell junctions with NCSC, and instead promote efficient attachment to a laminin support and monolayer growth. Thus, by controlling beta-cell adhesion and cell-to-cell junctions, ZBED6 might play an important role in beta-cell differentiation, proliferation and survival.

Place, publisher, year, edition, pages
2016. Vol. 6, 19006
National Category
Cell and Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-223614DOI: 10.1038/srep19006ISI: 000368128900001PubMedID: 26750727OAI: oai:DiVA.org:uu-223614DiVA: diva2:713355
Funder
Swedish Diabetes AssociationSwedish Child Diabetes FoundationNovo NordiskSwedish Research Council, 20716Stiftelsen Olle Engkvist Byggmästare
Available from: 2014-04-22 Created: 2014-04-22 Last updated: 2017-12-05Bibliographically approved
In thesis
1. Study of the Proliferation, Function and Death of Insulin-Producing Beta-Cells in vitro: Role of the Transcription Factor ZBED6
Open this publication in new window or tab >>Study of the Proliferation, Function and Death of Insulin-Producing Beta-Cells in vitro: Role of the Transcription Factor ZBED6
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

A thorough understanding of beta-cell proliferation, function, death and regeneration under normal condition as well as in the progression of diabetes is crucial to the conquest of this disease. The work presented in this thesis aimed to investigate the expression and role of a novel transcription factor, Zinc finger BED domain-containing protein 6 (ZBED6), in beta-cells.

ZBED6 was present in mouse βTC-6 cells and human islets as a double nuclear band at 115/120 kDa and as a single cytoplasmic band at 95-100 kDa, which lacked N-terminal nuclear localization signals. Lentiviral shRNA-mediated stable silencing of ZBED6 in βTC-6 cells resulted in altered morphology, decreased proliferation, a partial S/G2 cell cycle arrest, increased expression of beta-cell specific genes, and higher rates of apoptosis. ChIP sequencing of human islets showed that ZBED6 binding was preferentially to genes that control transcription, macromolecule biosynthesis and apoptosis. We proposed that ZBED6 supported proliferation and survival of beta-cells, possibly at the expense of specialized beta-cell function, i.e. insulin production.

To further investigate the role of ZBED6 in beta-cells, ChIP sequencing and whole transcriptome analysis were performed using MIN6 cells. More than 4000 putative target genes of ZBED6 were identified, including Pdx1, MafA and Nkx6.1. ZBED6-silencing resulted in differential expression of more than 700 genes, which was paralleled by an increase in the content and release of insulin in response to a high glucose concentration. Altered morphology/growth patterns as indicated by increased cell clustering were observed in ZBED6 silenced cells. We found also that ZBED6 decreased the ratio between N- and E-cadherin. A lower N- to E-cadherin ratio may hamper the formation of three-dimensional beta-cell clusters and cell-to-cell junctions with neural crest stem cells, and instead promote efficient attachment to a laminin support and monolayer growth. Thus, by controlling beta-cell adhesion and cell-to-cell junctions, ZBED6 might play an important role in beta-cell differentiation, proliferation and survival.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. 53 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1002
Keyword
ZBED6, Pancreatic beta-cell, Proliferation, Insulin secretion, Apoptosis, Adhesion
National Category
Medical and Health Sciences Cell and Molecular Biology
Research subject
Medical Science
Identifiers
urn:nbn:se:uu:diva-223616 (URN)978-91-554-8959-5 (ISBN)
Public defence
2014-06-10, A1:107a, BMC, Husargatan 3, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2014-05-20 Created: 2014-04-22 Last updated: 2014-06-30

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Wang, XuanXie, BeichenWallerman, OlaVasylovska, SvitlanaAndersson, LeifKozlova, ElenaWelsh, Nils

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