Taspase1-dependent TFIIA cleavage coordinates head morphogenesis by limiting Cdkn2a locus transcription
Shugaku Takeda, Satoru Sasagawa, Toshinao Oyama, Adam C. Searleman, Todd D. Westergard, Emily H. Cheng, James J. Hsieh
Shugaku Takeda, Satoru Sasagawa, Toshinao Oyama, Adam C. Searleman, Todd D. Westergard, Emily H. Cheng, James J. Hsieh
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Taspase1-dependent TFIIA cleavage coordinates head morphogenesis by limiting Cdkn2a locus transcription

Abstract

Head morphogenesis requires complex signal relays to enable precisely coordinated proliferation, migration, and patterning. Here, we demonstrate that, during mouse head formation, taspase1-mediated (TASP1-mediated) cleavage of the general transcription factor TFIIA ensures proper coordination of rapid cell proliferation and morphogenesis by maintaining limited transcription of the negative cell cycle regulators p16Ink4a and p19Arf from the Cdkn2a locus. In mice, loss of TASP1 function led to catastrophic craniofacial malformations that were associated with inadequate cell proliferation. Compound deficiency of Cdkn2a, especially p16Ink4a deficiency, markedly reduced the craniofacial anomalies of TASP1-deficent mice. Furthermore, evaluation of mice expressing noncleavable TASP1 targets revealed that TFIIA is the principal TASP1 substrate that orchestrates craniofacial morphogenesis. ChIP analyses determined that noncleaved TFIIA accumulates at the p16Ink4a and p19Arf promoters to drive transcription of these negative regulators. In summary, our study elucidates a regulatory circuit comprising proteolysis, transcription, and proliferation that is pivotal for construction of the mammalian head.

Authors

Shugaku Takeda, Satoru Sasagawa, Toshinao Oyama, Adam C. Searleman, Todd D. Westergard, Emily H. Cheng, James J. Hsieh

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Figure 4

Craniofacial and body size anomalies of Tasp1–/– embryos are rescued by Cdkn2a deficiency.

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Craniofacial and body size anomalies of Tasp1–/– embryos are rescued by ...
(A) RNA harvested from E10.5 WT and Tasp1–/– heads was subjected to microarray. Gene Ontology analysis shows functional categories of differentially expressed genes in the order of the enrichment score. (B) GSEA of expression signals from WT and Tasp1–/– samples. Enrichment plot of the “CELL_CYCLE_KEGG” gene set, indicating enrichment of cell cycle–associated genes among significantly upregulated mRNAs in Tasp1–/– heads (P < 0.01, normalized enrichment scores = 1.57). Genes showing core enrichment (indicated by inclusion in the purple bar under the enrichment plot) are listed by relative expression (red, high; blue, low). (C) Increased p16Ink4a and p19Arf mRNA levels were detected in E10.5 Tasp1–/– embryonic heads by quantitative RT-PCR (n = 3 throughout). Error bars represent SD. *P < 0.05, **P < 0.01, unpaired 2-tailed Student’s t test. (D) Frequencies of the otocephalic phenotypes in E14.5 embryos of the indicated genotypes. Note that Cdkn2a deficiency significantly rescued Tasp1–/– craniofacial malformations. **P < 0.01, Fisher’s exact test. (E) Body sizes of E18.5 embryos of the indicated genotypes. The distance from the ears to the base of tail was measured using ImageJ analysis. Note that Cdkn2a deficiency significantly rescued the reduced body sizes of Tasp1–/– embryos (n = 4 throughout). Error bars represent SD. **P < 0.01, unpaired 2-tailed Student’s t test.