Chang DR et al. (2013), Lung epithelial branching program antagonizes a...

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Proc Natl Acad Sci U S A 2013 Nov 05;11045:18042-51. doi: 10.1073/pnas.1311760110.

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Lung epithelial branching program antagonizes alveolar differentiation.

Chang DR , Martinez Alanis D , Miller RK , Ji H , Akiyama H , McCrea PD , Chen J .

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Mammalian organs, including the lung and kidney, often adopt a branched structure to achieve high efficiency and capacity of their physiological functions. Formation of a functional lung requires two developmental processes: branching morphogenesis, which builds a tree-like tubular network, and alveolar differentiation, which generates specialized epithelial cells for gas exchange. Much progress has been made to understand each of the two processes individually; however, it is not clear whether the two processes are coordinated and how they are deployed at the correct time and location. Here we show that an epithelial branching morphogenesis program antagonizes alveolar differentiation in the mouse lung. We find a negative correlation between branching morphogenesis and alveolar differentiation temporally, spatially, and evolutionarily. Gain-of-function experiments show that hyperactive small GTPase Kras expands the branching program and also suppresses molecular and cellular differentiation of alveolar cells. Loss-of-function experiments show that SRY-box containing gene 9 (Sox9) functions downstream of Fibroblast growth factor (Fgf)/Kras to promote branching and also suppresses premature initiation of alveolar differentiation. We thus propose that lung epithelial progenitors continuously balance between branching morphogenesis and alveolar differentiation, and such a balance is mediated by dual-function regulators, including Kras and Sox9. The resulting temporal delay of differentiation by the branching program may provide new insights to lung immaturity in preterm neonates and the increase in organ complexity during evolution.

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Species referenced: Xenopus laevis
Genes referenced: aqp1 gnas kras nkx2-1 npat sftpb sftpc sox13 sox9

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Fig. 1. Temporal (A), spatial (B), and evolutionary (C) negative correlations between branching morphogenesis and alveolar differentiation. (A) Time-course microarray expression profiling of FACS-purified distal lung epithelial cells. Gene expression values (log2) in E14 samples are used as a baseline for comparison. Horizontal histograms for E15ï¿½E19 samples are the frequency distributions of average fold-changes for all genes; the units are shown as the number of genes (# genes). Alveolar markers are up-regulated (red), and branching (green) and cell cycle (blue) related genes are down-regulated over time. Down-regulated genes appear to have a smaller fold-change than up-regulated genes, possibly because of perdurance of transcripts. (B) Confocal images of immunostained E18 lung sections in areas where the airway lumen can be continuously traced from the proximal conducting airways (black/white long dashed lines) to the distal nonbranch tip (magenta) and branch tip (green) regions, as illustrated in the schematics. Branch tips are outlined with dashed lines. The boxed regions are enlarged as insets outlined in corresponding colors, showing a spatial negative correlation between branching and alveolar differentiation. AQP1 labels alveolar type I cells, the vasculature and the mesothelium, which are separated by dashed lines (Insets). There is a very low level of SOX9 expression in nonbranch tip epithelium, possibly because of protein perdurance. (Scale bars, 20 Î¼m.) (C) Whole-mount in situ hybridization and immunostaining of Xenopus embryos of indicated stages (St) showing the absence of Sox9 expression and branching in the Xenopus lung. The lungs are indicated with dashed lines if stained or arrowheads if unstained. Open arrowheads indicate Sox9 expression in the pharyngeal arches. The two Xenopus Sox9 homologs (Sox9a and Sox9b) are 95% identical on the nucleotide level. OPT images of immunostained embryos (Left, Bottom) are shown as maximal intensity projection (Left) and sectional view (Right). (Scale bar, 200 Î¼m.)

Fig. S4. Xenopus lungs lack the branching program and Sox9 expression, and initiates alveolar differentiation immediately after lung specification. Whole mount in situ hybridization of Xenopus embryos at indicated stages (St). The lungs are indicated with dashed lines if stained or arrowheads if unstained. Open arrowheads indicate Sox9 expression in the pharyngeal arches. The onset of Sftpb expression is slightly later than that of Sftpc expression (Fig. 1C). (Scale bar, 200 Î¼m.)

References [+] :

Agrons, From the archives of the AFIP: Lung disease in premature neonates: radiologic-pathologic correlation. 2005, Pubmed