Sonic hedgehog signaling regulates Gli3 processing, mesenchymal proliferation, and differentiation during mouse lung organogenesis

Lack of Sonic hedgehog (Shh) signaling, mediated by the Gli proteins, leads to severe pulmonary hypoplasia. However, the precise role of Gli genes in lung development is not well established. We show Shh signaling prevents Gli3 proteolysis to generate its repressor forms (Gli3R) in the developing murine lung. In Shh(-/-) or cyclopamine-treated wild-type (WT) lung, we found that Gli3R level is elevated, and this upregulation appears to contribute to defects in proliferation and differentiation observed in the Shh(-/-) mesenchyme, where Gli3 is normally expressed. In agreement, we found Shh(-/-);Gli3(-/-) lungs exhibit enhanced growth potential. Vasculogenesis is also enhanced; in contrast, bronchial myogenesis remains absent in Shh(-/-);Gli3(-/-) compared with Shh(-/-) lungs. Genes upregulated in Shh(-/-);Gli3(-/-) relative to Shh(-/-) lung include Wnt2 and, surprisingly, Foxf1 whose expression has been reported to be Shh-dependent. Cyclins D1, D2, and D3 antibody labelings also reveal distinct expression patterns in the normal and mutant lungs. We found significant repression of Tbx2 and Tbx3, both linked to inhibition of cellular senescence, in Shh(-/-) and partial derepression in Shh(-/-); Gli3(-/-) lungs, while Tbx4 and Tbx5 expressions are less affected in the mutants. Our findings shed light on the role of Shh signaling on Gli3 processing in lung growth and differentiation by regulating several critical genes.     

Tbx Genes Specify Posterior Digit Identity through Shh and BMP Signaling

Despite extensive studies on the anterior-posterior (AP) axis formation of limb buds, mechanisms that specify digit identities along the AP axis remain obscure. Using the four-digit chick leg as a model, we report here that Tbx2 and Tbx3 specify the digit identities of digits IV and III, respectively. Misexpression of Tbx2 and Tbx3 induced posterior homeotic transformation of digit III to digit IV and digit II to digit III, respectively. Conversely, misexpression of their mutants VP16 Delta Tbx2 and VP16 Delta Tbx3 induced anterior transformation. In both cases, alterations in the expression of several markers (e.g., BMP2, Shh, and HoxD genes) were observed. In addition, Tbx2 and Tbx3 rescued Noggin-mediated inhibition of interdigital BMP signaling, signaling which is pivotal in establishing digit identities. Hence, we conclude that Tbx3 specifies digit III, and the combination of Tbx2 and Tbx3 specifies digit IV, acting together with the interdigital BMP signaling cascade.     

Tbx1基因功能下调影响斑马鱼Tbx20和Tbx2表达

目的采用吗啡啉修饰反义寡核苷酸显微注射方法下调斑马鱼Tbx1基因表达,研究斑马鱼Tbx1基因功能下调对其他两个T盒基因Tbx20和Tbx2表达的影响。方法采用吗啡啉修饰的反义寡核苷酸显微注射方法抑制斑马鱼Tbx1基因表达,分别将2.5、5、8、10 ng吗啡啉反义寡核苷酸在斑马鱼0-4细胞期注入胚胎,并构建Tbx20,骨形成蛋白2b（Bmp2b）和Tbx2反义RNA探针,进行整体原位杂交,观察Tbx1基因下调对Tbx20、Bmp2b及Tbx2表达的影响。结果Tbx1吗啡啉寡核苷酸显微注射组胚胎表现出鳃弓、耳囊、心血管系统和胸腺的发育异常。Tbx1基因下调导致Tbx20的表达出现改变,Tbx20在心脏的表达与对照组相比明显下调,神经元的表达范围明显缩小;Tbx1基因功能下调会导致Bmp2b在心脏和咽囊的表达减低,Bmp2b在后部咽囊的表达较前部咽囊减低得更为明显;Tbx1基因功能下调胚胎,Tbx2在鳃弓的表达模式发生改变,48 hpf,Tbx2在鳃弓的表达出现从后向前逐渐减低,鳃弓的表达范围较对照组明显缩小。结论Tbx1在发育过程中,会对其他T盒基因,如Tbx20和Tbx2具有激活或抑制的调控作用。Tbx1对Tbx20的作用可能是通过影响Bmp2b的途径,继发地影响Tbx20的表达。Tbx1基因功能下调,会改变Tbx2在鳃弓的表达模式。