Numerous signals drive the proliferative expansion of the distal endoderm and the underlying mesenchyme during lung branching morphogenesis, but little is known about how these signals are integrated. Here, we show by analysis of conditional double mutants that the two T-box transcription factor genes Tbx2 and Tbx3 act together in the lung mesenchyme to maintain branching morphogenesis. Expression of both genes depends on epithelially derived Shh signaling, with additional modulation by Bmp, Wnt, and Tgfβ signaling. Genetic rescue experiments reveal that Tbx2 and Tbx3 function downstream of Shh to maintain pro-proliferative mesenchymal Wnt signaling, in part by direct repression of the Wnt antagonists Frzb and Shisa3. In combination with our previous finding that Tbx2 and Tbx3 repress the cell-cycle inhibitors Cdkn1a and Cdkn1b, we conclude that Tbx2 and Tbx3 maintain proliferation of the lung mesenchyme by way of at least two molecular mechanisms: regulating cell-cycle regulation and integrating the activity of multiple signaling pathways.

The magnetic compass of birds seems to be based on light-dependent radical-pair processes in the eyes. Cryptochromes are currently the only candidate proteins known in vertebrates that may serve as the primary radical-pair-based magnetoreceptor molecules. Previous immunohistochemical studies have suggested that cryptochrome 1a (Cry1a) is localised in the photoreceptor outer segments of the ultraviolet/violet (UV/V) cones, and it has been claimed that differences in Cry1a antibody staining intensities show that Cry1a is activated by light and that this should make Cry1a the most likely magnetoreceptive candidate molecule. Here, we present an independent study of Cry1a distribution within retinae of several bird species, ranging from non-migratory domestic chicken and rock pigeon to night-migratory passerines, using both the previously used antibody and two newly generated antibodies, one against the same epitope as the originally used antibody and one against a different epitope of Cry1a. We confirm the UV/V cone outer segment localisation of Cry1a in all the tested bird species. In some stainings, we found Cry1a immunoreactivity as a distinct punctate pattern throughout the whole length of the UV/V cone outer segments. These dots with a diameter of around 170 nm might suggest that many Cry1a molecules accumulate in distinct spots in the UV/V cone outer segments. However, we did not see any notable difference in Cry1a immunoreactivity between lightand dark-adapted retinae. We find no evidence whatsoever that a C-terminal antibody against Cry1a labels only a light-activated form of the Cry1a protein.

The differentiated cell types of the epithelial and mesenchymal tissue compartments of the mature ureter of the mouse arise in a precise temporal and spatial sequence from uncommitted precursor cells of the distal ureteric bud epithelium and its surrounding mesenchyme. Previous genetic efforts identified a member of the Hedgehog (HH) family of secreted proteins, Sonic hedgehog (SHH) as a crucial epithelial signal for growth and differentiation of the ureteric mesenchyme. Here, we used conditional loss- and gain-of-function experiments of the unique HH signal transducer Smoothened (SMO) to further characterize the cellular functions and unravel the effector genes of HH signaling in ureter development. We showed that HH signaling is not only required for proliferation and SMC differentiation of cells of the inner mesenchymal region but also for survival of cells of the outer mesenchymal region, and for epithelial proliferation and differentiation. We identified the Forkhead transcription factor gene Foxf1 as a target of HH signaling in the ureteric mesenchyme. Expression of a repressor version of FOXF1 in this tissue completely recapitulated the mesenchymal and epithelial proliferation and differentiation defects associated with loss of HH signaling while re-expression of a wildtype version of FOXF1 in the inner mesenchymal layer restored these cellular programs when HH signaling was inhibited. We further showed that expression of Bmp4 in the ureteric mesenchyme depends on HH signaling and Foxf1, and that exogenous BMP4 rescued cell proliferation and epithelial differentiation in ureters with abrogated HH signaling or FOXF1 function. We conclude that SHH uses a FOXF1-BMP4 module to coordinate the cellular programs for ureter elongation and differentiation, and suggest that deregulation of this signaling axis occurs in human congenital anomalies of the kidney and urinary tract (CAKUT).