Abstract
Mutations of the CUL4B ubiquitin ligase gene are causally linked to syndromic X-linked mental retardation (XLMR). However, the pathogenic role of CUL4B mutations in neuronal and developmental defects is not understood. We have generated mice with targeted disruption of Cul4b, and observed embryonic lethality with pronounced growth inhibition and increased apoptosis in extra-embryonic tissues. Cul4b, but not its paralog Cul4a, is expressed at high levels in extra-embryonic tissues post implantation. Silencing of CUL4B expression in an extra-embryonic cell line resulted in the robust accumulation of the CUL4 substrate p21Cip1/WAF and G2/M cell cycle arrest, which could be partially rescued by silencing of p21Cip1/WAF. Epiblast-specific deletion of Cul4b prevented embryonic lethality and gave rise to viable Cul4b null mice. Therefore, while dispensable in the embryo proper, Cul4b performs an essential developmental role in the extra-embryonic tissues. Our study offers a strategy to generate viable Cul4b-deficient mice to model the potential neuronal and behavioral deficiencies of human CUL4B XLMR patients.
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Acknowledgements
We thank Sundeep Kalantry (University of Michigan, USA) for the XEN cells, Wei Gu (Columbia Universtity, USA) for help on generation of the germline-transmitted Cul4bf/+ mice and Willie Mark (Sloan-Kettering Cancer Center, USA) for the CAG-Cre transgenic mice, and Jennifer Lee and Brent Bany for critical reading of the manuscript. This work is supported by National Institutes of Health Grants CA098210 and CA118085 to PZ, ES016597 to LM, and GM58726 and CA08748 to EL. PZ is supported in part by the Irma T Hirschl Career Scientist Award and the Tri-Institutional Stem Cell Initiative. EL is also supported in part by NYSTEM (CO2433).
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
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Supplementary information, Table S1 (download PDF )
Genotyping analysis of offspring from breeding between EIIA-Cre+/+ (♂) and Cul4bf/+ (♀) mice and mosaic mice (PDF 31 kb)
Supplementary information, Figure S1 (download PDF )
Extraembryonic tissues of heterozygous Cul4b+/− embryos exhibited imprinted paternal X chromosome inactivation. (PDF 86 kb)
Supplementary information, Figure S2 (download PDF )
Normal morphology and size of the conceptuses. (PDF 83 kb)
Supplementary information, Figure S3 (download PDF )
Evaluation of early stage apoptosis by annexin V-FITC/PI staining. (PDF 121 kb)
Supplementary information, Figure S4 (download PDF )
Silencing of Cul4a expression has in minimal or no effect on CUL4 substrates in the extraembryonic XEN cells. (PDF 60 kb)
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Liu, L., Yin, Y., Li, Y. et al. Essential role of the CUL4B ubiquitin ligase in extra-embryonic tissue development during mouse embryogenesis. Cell Res 22, 1258–1269 (2012). https://doi.org/10.1038/cr.2012.48
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DOI: https://doi.org/10.1038/cr.2012.48
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