Hoxa13 regulates expression of common Hox target genes involved in cartilage development to coordinate the expansion of the autopodal anlage
Hoxa13 regulates expression of common Hox target genes involved in cartilage development to coordinate the expansion of the autopodal anlage
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DOI：
10.1111/dgd.12601
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发表时间：
2019-03
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通讯作者：
Shiori Yamamoto;Yuji Uchida;Tomomi Ohtani;Erina Nozaki;Chunyang Yin;Yoshihiro Gotoh;Nayuta Yakushiji-Kaminatsui;T. Higashiyama;Takamasa Suzuki;T. Takemoto;Yoshiharu Shiraishi;A. Kuroiwa
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作者：
Shiori Yamamoto;Yuji Uchida;Tomomi Ohtani;Erina Nozaki;Chunyang Yin;Yoshihiro Gotoh;Nayuta Yakushiji-Kaminatsui;T. Higashiyama;Takamasa Suzuki;T. Takemoto;Yoshiharu Shiraishi;A. Kuroiwa
为了阐明Hox基因在肢体软骨发育中的作用，我们通过ChIP-Seq鉴定了HOXA 11和HOXA 13的靶基因。ChIP DNA片段含有进化上保守的序列和多个高度保守的HOX结合位点。HOXA 11 ChIP片段的相当大一部分与HOXA 13 ChIP片段重叠，表明两种因子具有共同的靶点。邻近Bmp 2或Tshz 2的靶区域的缺失降低了它们在直足动物中的表达，表明它们作为肢芽特异性增强子起作用。我们通过基因芯片分析鉴定了Hox下游基因在Hoxa 13敲除（KO）和Hoxd 11 - 13缺失双突变体（Hox 13 dKO）中表现出表达变化。与ChIP片段相邻的Hox下游基因被定义为Hox的直接靶标。我们分析了Hox靶基因的空间表达模式，该基因编码两种不同类别的转录因子，在直足发育和Hox 13 dKO肢芽。(a)Bcl 11 a，编码软骨分化的阻遏物，在E11.5 autopod中表达，并在Hox 13 dKO中显著降低。(b)转录因子Aff 3，Bnc 2，Nfib和Runx 1 t1在轭足软骨中表达，但由于Hox 13在E11.5时的抑制性或相对较弱的转录活性而不在足中表达。有趣的是，这些基因的表达后来在E12.5的自体足软骨中观察到。这些结果表明，Hox 13通过多种途径暂时暂停了autopodal原基中的软骨分化，直到建立产生五个手指所需的桨状结构。
To elucidate the role of Hox genes in limb cartilage development, we identified the target genes of HOXA11 and HOXA13 by ChIP‐Seq. The ChIP DNA fragment contained evolutionarily conserved sequences and multiple highly conserved HOX binding sites. A substantial portion of the HOXA11 ChIP fragment overlapped with the HOXA13 ChIP fragment indicating that both factors share common targets. Deletion of the target regions neighboring Bmp2 or Tshz2 reduced their expression in the autopod suggesting that they function as the limb bud‐specific enhancers. We identified the Hox downstream genes as exhibiting expression changes in the Hoxa13 knock out (KO) and Hoxd11‐13 deletion double mutant (Hox13 dKO) autopod by Genechip analysis. The Hox downstream genes neighboring the ChIP fragment were defined as the direct targets of Hox. We analyzed the spatial expression pattern of the Hox target genes that encode two different categories of transcription factors during autopod development and Hox13dKO limb bud. (a) Bcl11a, encoding a repressor of cartilage differentiation, was expressed in the E11.5 autopod and was substantially reduced in the Hox13dKO. (b) The transcription factors Aff3, Bnc2, Nfib and Runx1t1 were expressed in the zeugopodal cartilage but not in the autopod due to the repressive or relatively weak transcriptional activity of Hox13 at E11.5. Interestingly, the expression of these genes was later observed in the autopodal cartilage at E12.5. These results indicate that Hox13 transiently suspends the cartilage differentiation in the autopodal anlage via multiple pathways until establishing the paddle‐shaped structure required to generate five digits.