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Baf60c is essential for function of BAF chromatin remodelling complexes in heart development

Abstract

Tissue-specific transcription factors regulate several important aspects of embryonic development. They must function in the context of DNA assembled into the higher-order structure of chromatin. Enzymatic complexes such as the Swi/Snf-like BAF complexes remodel chromatin to allow the transcriptional machinery access to gene regulatory elements1,2. Here we show that Smarcd3, encoding Baf60c, a subunit of the BAF complexes, is expressed specifically in the heart and somites in the early mouse embryo. Smarcd3 silencing by RNA interference in mouse embryos derived from embryonic stem cells causes defects in heart morphogenesis that reflect impaired expansion of the anterior/secondary heart field, and also results in abnormal cardiac and skeletal muscle differentiation. An intermediate reduction in Smarcd3 expression leads to defects in outflow tract remodelling reminiscent of human congenital heart defects. Baf60c overexpressed in cell culture can mediate interactions between cardiac transcription factors and the BAF complex ATPase Brg1, thereby potentiating the activation of target genes. These results reveal tissue-specific and dose-dependent roles for Baf60c in recruiting BAF chromatin remodelling complexes to heart-specific enhancers, providing a novel mechanism to ensure transcriptional regulation during organogenesis.

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Figure 1: Baf60c is required for heart formation.
Figure 2: Cardiac gene expression in wild-type (WT) and Smarcd3 knockdown (siSmarcd3) embryos.
Figure 3: Transgenic rescue of heart defects in siSmarcd3 embryos.
Figure 4: Baf60c potentiates transcription by promoting interactions between transcription factors and the BAF complex.
Figure 5: Dosage-sensitive role of Baf60c in outflow tract septation.

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Acknowledgements

We thank J. Sharpe for providing a prototype OPT system, S. McMaster for tetraploid aggregations, D. Holmyard for scanning electron microscopy, K. Harpal and K. Koshiba-Takeuchi for histology, H. Hamada and C. Meno for the unpublished Pitx2 reporter, and A. Nagafuchi, M. Nemer, R. Schwartz, D. Srivastava and T. Takeuchi for reporter and expression constructs. This research was funded by the Canadian Institutes of Health Research (CIHR; to B.G.B., J.R. and J.L.W.), the Heart and Stroke Foundation of Ontario (to B.G.B.), the March of Dimes Birth Defects Foundation (to B.G.B.), NCIC (to J.L.W.), the Canada Foundation for Innovation (to R.M.H.), an Emmy–Noether fellowship of the Deutsche Forschungsgemeinschaft (to H.L.), a Human Frontiers Science Program long-term fellowship (to J.K.T.), an OGSST fellowship (to J.R.W.), and the Koeln Fortune Program, University of Cologne (to I.v.B.).

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Correspondence to Janet Rossant or Benoit G. Bruneau.

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Supplementary information

Supplementary Figure 1

Smarcd3 gene expression at E8.5 and E10.5, showing expression in myocardium, high levels of expression in the outflow and inflow tracts, and expression in the myotome and limb bud mesenchyme. (JPG 142 kb)

Supplementary Figure 2

Expression of Smarcd1-3 in mouse development at E7.5, E8.75, and E9.25, showing lack of expression of Smarcd1 or Smarcd2 in the heart. (JPG 140 kb)

Supplementary Figure 3

Smarcd3 siRNA sequences , quantitation of Smarcd3 levels in transgenic ES cells, and protein levels of Baf60c and other Baf proteins. (JPG 176 kb)

Supplementary Figure 4

Normal Tbx5, Isl1, Fgf10, Gata4, and Nkx2-5 gene expression and abnormal Tbx20 and Irx3 gene expression in Smarcd3 knockdown embryos. (JPG 254 kb)

Supplementary Figure 5

Defective skeletal muscle differentiation in Smarcd3 knockdown embryos. (JPG 116 kb)

Supplementary Table 1

Cardiac function of Smarcd3 intermediate knockdown (siSMarcd3-A#2) and wild-type mouse embryos at E11.5. (DOC 22 kb)

Supplementary Movie

Rendered OPT of a wild-type (left) and Smarcd3 knockdown (right) embryos at E9.5. The heart is labeled red, and the rest of the embryo is translucent white. (MOV 5228 kb)

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Lickert, H., Takeuchi, J., von Both, I. et al. Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature 432, 107–112 (2004). https://doi.org/10.1038/nature03071

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