Abstract
Hox genes orchestrate the segmental specification of the muscular circulatory system in invertebrates but it has not proven straightforward to decipher segmental parallels in the vertebrate heart. Recently, patients with HOXB gene cluster deletion were found to exhibit abnormalities including atrioventricular canal defects. Using CRISPR, we established a mutant with the orthologous hoxbb cluster deletion in zebrafish. The mutant exhibited heart failure and atrioventricular regurgitation at 5 days. Analyzing the four genes in the hoxbb cluster, isolated deletion of hoxb1b−/− recapitulated the cardiac abnormalities, supporting hoxb1b as the causal gene. Both in situ and in vitro data indicated that hoxb1b regulates gata5 to inhibit hand2 expression and ultimately is required to pattern the vertebrate atrioventricular boundary. Together, these data reveal a role for segmental specification in vertebrate cardiac development and highlight the utility of CRISPR techniques for efficiently exploring the function of large structural genomic lesions.
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Acknowledgements
We thank Dr. Bo Zhang (School of Life Sciences, Peking University) for providing the zCas9 plasmid, Dr. Jingwei Xiong (Institute of Molecular Medicine, Peking University) for providing the pUC19-scaffold plasmid for gRNA synthesis, Dr. Yi Zhou(Harvard Medical School) for the data discussion. We thank Jie Wang, Yumeng Rui, Xuebin Ye, Yihao Zhu, Xuhui Han, Wenhao Li, and Liang Jia for data analysis assistance.
Funding
This work was supported by the National Natural Science Foundation of China [No. 32170423 and 31501166 to Y.Z.], Shanghai Sailing Program of Science and Technology Commission of Shanghai Municipality [15YF1405000 to Y.Z.], Chenguang Program of the Shanghai Municipal Education Commission [14CG49 to Y.Z.], SciTech Funding by CSPFTZ Lingang Special Area Marine Biomedical Innovation Platform. Calum A. MacRae was funded by the NIH (OD017870).
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Conceptualization and design, Y.Z.; Investigation, P.H., Y.Z., B.W., D.J., Y.G., H.H., and X.M.; Formal analysis, Y.Z., P.H., B.W., D.J., Y.G., W.Z., D.Y.C. and C.A.M.; Writing, Y.Z., P.H., C.A.M. and B.W.; Resources, Y.Z. W. L. and C.A.M.; Funding acquisition, Y.Z. and C.A.M. All authors have read and approved the final manuscript.
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Hu, P., Wang, B., Jin, D. et al. Modeling of large-scale hoxbb cluster deletions in zebrafish uncovers a role for segmentation pathways in atrioventricular boundary specification. Cell. Mol. Life Sci. 80, 317 (2023). https://doi.org/10.1007/s00018-023-04933-2
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DOI: https://doi.org/10.1007/s00018-023-04933-2