The bHLH Protein Nulp1 is Essential for Femur Development Via Acting as a Cofactor in Wnt Signaling in Drosophila.
The bHLH Protein Nulp1 is Essential for Femur Development Via Acting as a Cofactor in Wnt Signaling in Drosophila.
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bHLH 蛋白 Nulp1 通过作为果蝇 Wnt 信号转导的辅助因子对股骨发育至关重要
DOI：
10.2174/1566524018666180212145714
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发表时间：
2018-03-09
影响因子：
2.5
通讯作者：
Wu X
中科院分区：
文献类型：
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作者：
Zeng Q;Wan Y;Zhu P;Zhao M;Jiang F;Chen J;Tang M;Zhu X;Li Y;Zha H;Wang Y;Hu M;Mo X;Zhang Y;Chen Y;Chen Y;Ye X;Bodmer R;Ocorr K;Jiang Z;Zhuang J;Yuan W;Wu X
背景资料：碱性螺旋-环-螺旋（basic helix-loop-helix，bHLH）蛋白家族是一类与细胞增殖、组织分化和其他重要发育过程相关的转录因子。核定位蛋白1（Nuclear localized protein-1，Nulp 1）可能是一种新的bHLH转录因子，参与细胞功能的调控。然而，它在体内发育中的作用仍然未知。方法：通过CRISPR/Cas9靶向其C末端的未知功能结构域（DUF 654）产生Nulp 1（dNulp 1）突变体。通过qRT-PCR分析Wg靶基因的表达。我们使用Top-Flash荧光素酶报告基因测定来响应Wg信号传导。结果如下：在这里，我们表明，由CRISPR/Cas9靶向其C末端的未知功能结构域（DUF 654）产生的果蝇Nulp 1（dNulp 1）突变体是部分纯合致死的，并且罕见的逃生者具有弯曲的股骨，这与人类Stuve-Weidemann综合征中先天性骨发育不良的主要表现相似。果蝇表型可以通过dNulp 1的过表达来挽救，这表明dNulp 1对果蝇股骨发育和存活是必不可少的。此外，dNulp 1过表达抑制了由典型Wnt级联反应抑制剂sgg/GSK 3 β过表达引起的缺口翼表型。此外，qRT-PCR分析表明，dNulp 1基因敲除后，Wg信号通路正调控的7个靶基因表达下调，而2个负调控的Wg靶基因表达上调。最后，dNulp 1过表达显著激活了Top-Flash Wnt信号报告基因。结论：我们的结论是，bHLH蛋白dNulp 1是必不可少的股骨发育和生存在果蝇作为一个积极的辅因子Wnt/Wingless信号。
Background: The basic helix-loop-helix (bHLH) protein families are a large class of transcription factors, which are associated with cell proliferation, tissue differentiation, and other important development processes. We reported that the Nuclear localized protein-1 (Nulp1) might act as a novel bHLH transcriptional factor to mediate cellular functions. However, its role in development in vivo remains unknown. Methods: Nulp1 (dNulp1) mutants are generated by CRISPR/Cas9 targeting the Domain of Unknown Function (DUF654) in its C terminal. Expression of Wg target genes are analyzed by qRT-PCR. We use the Top-Flash luciferase reporter assay to response to Wg signaling. Results: Here we show that Drosophila Nulp1 (dNulp1) mutants, generated by CRISPR/Cas9 targeting the Domain of Unknown Function (DUF654) in its C terminal, are partially homozygous lethal and the rare escapers have bent femurs, which are similar to the major manifestation of congenital bent-bone dysplasia in human Stuve-Weidemann syndrome. The fly phenotype can be rescued by dNulp1 over-expression, indicating that dNulp1 is essential for fly femur development and survival. Moreover, dNulp1 overexpression suppresses the notch wing phenotype caused by the overexpression of sgg/GSK3β, an inhibitor of the canonical Wnt cascade. Furthermore, qRT-PCR analyses show that seven target genes positively regulated by Wg signaling pathway are down-regulated in response to dNulp1 knockout, while two negatively regulated Wg targets are up-regulated in dNulp1 mutants. Finally, dNulp1 overexpression significantly activates the Top-Flash Wnt signaling reporter. Conclusion: We conclude that bHLH protein dNulp1 is essential for femur development and survival in Drosophila by acting as a positive cofactor in Wnt/Wingless signaling.