CKD is a significant health concern with an underlying genetic component. Multiple genome-wide association studies (GWASs) strongly associated CKD with the shroom family member 3 (SHROOM3) gene, which encodes an actin-associated protein important in epithelial morphogenesis. However, the role of SHROOM3 in kidney development and function is virtually unknown. Studies in zebrafish and rat showed that alterations in Shroom3 can result in glomerular dysfunction. Furthermore, human SHROOM3 variants can induce impaired kidney function in animal models. Here, we examined the temporal and spatial expression of Shroom3 in the mammalian kidney. We detected Shroom3 expression in the condensing mesenchyme, Bowman's capsule, and developing and mature podocytes in mice. Shroom3 null (Shroom3Gt/Gt ) mice showed marked glomerular abnormalities, including cystic and collapsing/degenerating glomeruli, and marked disruptions in podocyte arrangement and morphology. These podocyte-specific abnormalities are associated with altered Rho-kinase/myosin II signaling and loss of apically distributed actin. Additionally, Shroom3 heterozygous (Shroom3 Gt/+ ) mice showed developmental irregularities that manifested as adult-onset glomerulosclerosis and proteinuria. Taken together, our results establish the significance of Shroom3 in mammalian kidney development and progression of kidney disease. Specifically, Shroom3 maintains normal podocyte architecture in mice via modulation of the actomyosin network, which is essential for podocyte function. Furthermore, our findings strongly support the GWASs that suggest a role for SHROOM3 in human kidney disease.

Purpose Pathological blood vessel growth in the eye is implicated in several diseases that result in vision loss, including age-related macular degeneration and diabetic retinopathy. The limits of current disease therapies have created the need to identify and characterize new antiangiogenic drugs. Here, we identify the secreted chemorepellent semaphorin-3fa (Sema3fa) as an endogenous anti-angiogenic in the eye. Methods We generated a CRISPR/Cas9 sema3fa zebrafish mutant line, sema3fa ca304/304 . We assessed the retinal and choroidal vasculature in both larval and adult wild-type and sema3fa mutant zebrafish. Results We find sema3fa mRNA is expressed by the ciliary marginal zone, neural retina, and retinal pigment epithelium of zebrafish larvae as choroidal vascularization emerges and the hyaloid/retinal vasculature is remodeled. The hyaloid vessels of sema3fa mutants develop appropriately but fail to remodel during the larval period, with adult mutants exhibiting a denser network of capillaries in the retinal periphery than seen in wild-type. The choroid vasculature is also defective in that it develops precociously, and aberrant, leaky sprouts are present in the normally avascular outer retina of both sema3fa ca304/304 larvae and adult fish. Conclusions Sema3fa is a key endogenous signal for maintaining an avascular retina and preventing pathologic vascularization. Furthermore, we provide a new experimentally accessible model for studying choroid neovascularization (CNV) resulting from primary changes in the retinal environment that lead to downstream vessel infiltration.

During development, neuroepithelial progenitors acquire apico-basal polarity and adhere to one another via apically located tight and adherens junction complexes. This polarized neuroepithelium must continue to integrate cells arising through cell divisions and intercalation, and allow for cell movements, at the same time as undergoing morphogenesis. Cell proliferation, migration and intercalation all occur in the morphing embryonic eye. To understand how eye development might depend on dynamic epithelial adhesion, we investigated the function of a known regulator of junctional plasticity, Tumour necrosis factor receptor-associated factor 4 (Traf4). traf4a mRNA is expressed in the developing eye vesicle over the period of optic cup morphogenesis, and Traf4a loss leads to disrupted evagination and elongation of the eye vesicles, and aberrant organization and apico-basal polarity of the eye epithelium. We propose a model whereby Traf4a regulates apical junction plasticity in nascent eye epithelium, allowing for its polarization and morphogenesis. Symbols and Abbreviations: AB: apico-basal; aPKC: atypical protein kinase-C; CRISPR: clustered regularly-interspaced short palindromic repeats; GFP: green fluorescent protein; hpf: hours post-fertilization; MO: antisense morpholino oligonucleotide; pHH3: phospho histone H3; ss: somite stage; Traf4: Tumour necrosis factor receptor-associated factor 4; ZO-1: zona occludens-1.

Background During development a pool of precursors form a heart with atrial and ventricular chambers that exhibit distinct transcriptional and electrophysiological properties. Normal development of these chambers is essential for full term survival of the fetus, and deviations result in congenital heart defects. The large number of genes that may cause congenital heart defects when mutated, and the genetic variability and penetrance of the ensuing phenotypes, reveals a need to understand the molecular mechanisms that allow for the formation of chamber-specific cardiomyocyte differentiation. Methods We used in situ hybridization, immunohistochemistry and functional analyses to identify the consequences of the loss of the secreted semaphorin, Sema3fb, in the development of the zebrafish heart by using two sema3fb CRISPR mutant alleles. Results We find that in the developing zebrafish heart sema3fb mRNA is expressed by all cardiomyocytes, whereas mRNA for a known receptor Plexina3 (Plxna3) is expressed preferentially by ventricular cardiomyocytes. In sema3fb CRISPR zebrafish mutants, heart chamber development is impaired; the atria and ventricles of mutants are smaller in size than their wild type siblings, apparently because of differences in cell size and not cell numbers. Analysis of chamber differentiation indicates defects in chamber specific gene expression at the border between the ventricular and atrial chambers, with spillage of ventricular chamber genes into the atrium, and vice versa, and a failure to restrict specialized cardiomyocyte markers to the atrioventricular canal (AVC). The hypoplastic heart chambers are associated with decreased cardiac output and heart edema. Conclusions Based on our data we propose a model whereby cardiomyocytes secrete a Sema cue that, because of spatially restricted expression of the receptor, signals in a ventricular chamber-specific manner to establish a distinct border between atrial and ventricular chambers that is important to produce a fully functional heart.