We study kidney organogenesis using the zebrafish to explore conserved molecular mechanisms underlying kidney cell differentiation, morphogenesis, and regeneration. We also study organ pathologies that results from mutations in genes required for cilia biogenesis and function. These include kidney cystic disease, retinal degeneration and left-right asymmetry defects.
The zebrafish pronephros as a model of the vertebrate kidney. As part of the Boston large-scale forward genetic screens on zebrafish, we defined zebrafish pronephric kidney development and cell structure, generated assays for nephron function, and characterized fifteen different mutant loci that caused kidney cyst formation and altered epithelial membrane protein targeting. Current work examines novel kidney mutants using Next-gen sequencing to rapidly map causative mutations.
Cell signaling in kidney development and patterning. Functionally distinct kidney cell types often exist right next to each other however the developmental mechanisms driving this patterning was unknown. We showed that "salt and pepper" patterns of cell types in the zebrafish kidney are generated by Notch signaling. We went on to show that altering kidney nephron patterning by modifying Notch signaling altered kidney function. We have also defined roles for the transcription factors pax2 and osr1 in nephron segment patterning and tubule vs. endothelial cell fate. Our current work focuses on using genetically encoded fluorescent biosensors to probe cell signaling during glomerular development and capillary development.
Kidney morphogenesis and epithelial cell dynamics. The kidney convoluted proximal tubule is specialized for interaction with the kidney vasculature and segment boundary positions are tightly regulated however little was known about morphogenetic processes that control nephron development. Using live, in vivo imaging in transparent zebrafish embryos, we showed that nephron morphogenesis is driven by collective cell migration that generates proximal convolutions and defines the final position of nephron segment boundaries. Also, we showed that cell migration was signaled by the onset of nephron filtration and luminal fluid flow, linking nephron morphogenesis with nephron function. We have also used live imaging to describe tubule interconnection in the developing mouse kidney and we have shown that cell migration plays a primary role in responses to kidney injury.
Primary Ciliary Dyskinesia (PCD) is a genetic disorder affecting 1 in 15,000 births that causes cilia paralysis with left-right organ asymmetry defects, bronchiectasis, sinusitis, and infertility. Using an unbiased forward genetic approach and targeted reverse genetic approaches in zebrafish, we cloned novel cilia paralysis genes, (schmalhans / ccdc103, C21orf59, and CCDC65) and went on to show that human patient families with PCD carried mutations in orthologous human genes, identifying novel causes of PCD. A key element of this work was developing an in vivo assay to test the pathogenicity of candidate human disease alleles by mRNA injection and rescue of the zebrafish phenotype. This work established the fish as an in vivo assay system for distinguishing pathogenic human gene mutations from benign sequence polymorphisms. We have also used fish genetics to reveal new cilia mediators of tubulin post-translational modification and new insights into physiological regulation of ciliogenesis.
Kidney regeneration from adult tissue stem cells. The zebrafish has a remarkable capacity to generate new nephrons from stem cells following renal injury. To restore kidney function, filtering nephrons must connect to a tubule network to pass fluid. How cells rearrange and form new connections is not known. We are using the regenerating zebrafish kidney as a model to discover how tubule interconnections are made and to uncover signals that drive cell rearrangments required to "plumb" the kidney. Knowledge of these signals will be an important part of the molecular toolbox for growing new organs.