title: Yes-associated protein (YAP) is a negative regulator of chondrogenesis in mesenchymal stem cells.
abstract: BACKGROUND
  The control of differentiation of mesenchymal stromal/stem cells (MSCs) is crucial for tissue engineering strategies employing MSCs. The purpose of this study was to investigate whether the transcriptional co-factor Yes-associated protein (YAP) regulates chondrogenic differentiation of MSCs.
  
  
  METHODS
  Expression of total YAP, its paralogue transcriptional co-activator with PDZ-binding motif (TAZ), and individual YAP transcript variants during in vitro chondrogenesis of human MSCs was determined by quantitative reverse transcription polymerase chain reaction (RT-PCR). YAP expression was confirmed by western blotting. To determine the effect of high YAP activity on chondrogenesis, C3H10T1/2 MSC-like cells were transduced with human (h)YAP and treated in micromass with bone morphogenetic protein-2 (BMP-2). Chondrogenic differentiation was assessed by alcian blue staining and expression of chondrocyte-lineage genes. BMP signalling was determined by detection of pSmad1,5,8 by western blotting and expression of BMP target genes by quantitative RT-PCR. Finally, YAP and pYAP were detected in mouse embryo hindlimbs by immunohistochemistry.
  
  
  RESULTS
  YAP, but not TAZ, was downregulated during in vitro chondrogenesis of human MSCs. One of the YAP transcript variants, however, was upregulated in high-density micromass culture. Overexpression of hYAP in murine C3H10T1/2 MSCs inhibited chondrogenic differentiation. High YAP activity in these cells decreased Smad1,5,8 phosphorylation and expression of the BMP target genes Inhibitor of DNA binding/differentiation (Id)1, Id2 and Id3 in response to BMP-2. In developing mouse limbs, Yap was nuclear in the perichondrium while mostly phosphorylated and cytosolic in cells of the cartilage anlage, suggesting downregulation of Yap co-transcriptional activity during physiological chondrogenesis in vivo.
  
  
  CONCLUSIONS
  Our findings indicate that YAP is a negative regulator of chondrogenic differentiation of MSCs. Downregulation of YAP is required for chondrogenesis through derepression of chondrogenic signalling. Therapeutic targeting of YAP to promote cartilage repair and prevent secondary osteoarthritis is an exciting prospect in rheumatology.
authors.affiliation: Musculoskeletal Research Programme, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, AB25 2ZD, UK. alexandrakarystinou@hotmail.com.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Karystinou
authors.affiliation: Musculoskeletal Research Programme, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, AB25 2ZD, UK. a.roelofs@abdn.ac.uk.
authors.forename: Anke J
authors.initials: AJ
authors.lastname: Roelofs
authors.affiliation: Musculoskeletal Research Programme, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, AB25 2ZD, UK. anna.neve@unifg.it.|Rheumatology Clinic, Department of Medical and Surgical Sciences, University of Foggia, Via Napoli 25, 71122, Foggia, Italy. anna.neve@unifg.it.
authors.forename: Anna
authors.initials: A
authors.lastname: Neve
authors.affiliation: Rheumatology Clinic, Department of Medical and Surgical Sciences, University of Foggia, Via Napoli 25, 71122, Foggia, Italy. francescopaolo.cantatore@unifg.it.
authors.forename: Francesco P
authors.initials: FP
authors.lastname: Cantatore
authors.affiliation: Musculoskeletal Research Programme, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, AB25 2ZD, UK. h.wackerhage@abdn.ac.uk.
authors.forename: Henning
authors.initials: H
authors.lastname: Wackerhage
authors.affiliation: Musculoskeletal Research Programme, Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen, AB25 2ZD, UK. c.debari@abdn.ac.uk.
authors.forename: Cosimo
authors.initials: C
authors.lastname: De Bari
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D010750:Phosphoproteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D010750:Phosphoproteins; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human