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Unraveling the transcriptional regulation of TWIST1 in limb development

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Item Type:Article
Title:Unraveling the transcriptional regulation of TWIST1 in limb development
Creators Name:Hirsch, N. and Eshel, R. and Bar Yaacov, R. and Shahar, T. and Shmulevich, F. and Dahan, I. and Levaot, N. and Kaplan, T. and Lupiáñez, D.G. and Birnbaum, R..
Abstract:The transcription factor TWIST1 plays a vital role in mesoderm development, particularly in limb and craniofacial formation. Accordingly, haploinsufficiency of TWIST1 can cause limb and craniofacial malformations as part of Saethre-Chotzen syndrome. However, the molecular basis of TWIST1 transcriptional regulation during development has yet to be elucidated. Here, we characterized active enhancers in the TWIST1-HDAC9 locus that drive transcription in the developing limb and branchial arches. Using available p300 and H3K27ac ChIP-seq data, we identified 12 enhancer candidates, located both within and outside the coding sequences of the neighboring gene, Histone deacetyase 9 (HDAC9). Using zebrafish and mouse enhancer assays, we showed that eight of these candidates have limb/fin and branchial arch enhancer activity that resemble Twist1 expression. Using 4C-seq, we showed that the Twist1 promoter region interacts with three enhancers (eTw-5, 6, 7) in the limb bud and branchial arch of mouse embryos at day 11.5. Furthermore, we found that two transcription factors, LMX1B and TFAP2, bind these enhancers and modulate their enhancer activity. Finally, using CRISPR/Cas9 genome editing, we showed that homozygous deletion of eTw5-7 enhancers reduced Twist1 expression in the limb bud and caused pre-axial polydactyly, a phenotype observed in Twist1+/- mice. Taken together, our findings reveal that each enhancer has a discrete activity pattern, and together comprise a spatiotemporal regulatory network of Twist1 transcription in the developing limbs/fins and branchial arches. Our study suggests that mutations in TWIST1 enhancers could lead to reduced TWIST1 expression, resulting in phenotypic outcome as seen with TWIST1 coding mutations.
Keywords:Branchial Region, Congenital Limb Deformities, Developmental Gene Expression Regulation, Extremities, Genetic Enhancer Elements, Histone Deacetylases, Homeobox Genes, Homeodomain Proteins, Inbred C57BL Mice, Limb Buds, Organogenesis, Repressor Proteins, Transcription Factor AP-2, Transcription Factors, Twist-Related Protein 1, Zebrafish Proteins, Animals, Mice, Zebrafish
Source:PLoS Genetics
ISSN:1553-7404
Publisher:Public Library of Science (U.S.A.)
Volume:14
Number:10
Page Range:e1007738
Date:29 October 2018
Official Publication:https://doi.org/10.1371/journal.pgen.1007738
PubMed:View item in PubMed

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