title: Wt1a, Foxc1a, and the Notch mediator Rbpj physically interact and regulate the formation of podocytes in zebrafish.
abstract: Podocytes help form the glomerular blood filtration barrier in the kidney and their injury or loss leads to renal disease. The Wilms' tumor suppressor-1 (Wt1) and the FoxC1/2 transcription factors, as well as Notch signaling, have been implicated as important regulators of podocyte fate. It is not known whether these factors work in parallel or sequentially on different gene targets, or as higher-order transcriptional complexes on common genes. Here, we use the zebrafish to demonstrate that embryos treated with morpholinos against wt1a, foxc1a, or the Notch transcriptional mediator rbpj develop fewer podocytes, as determined by wt1b, hey1 and nephrin expression, while embryos deficient in any two of these factors completely lack podocytes. From GST-pull-downs and co-immunoprecipitation experiments we show that Wt1a, Foxc1a, and Rbpj can physically interact with each other, whereas only Rbpj binds to the Notch intracellular domain (NICD). In transactivation assays, combinations of Wt1, FoxC1/2, and NICD synergistically induce the Hey1 promoter, and have additive or repressive effects on the Podocalyxin promoter, depending on dosage. Taken together, these data suggest that Wt1, FoxC1/2, and Notch signaling converge on common target genes where they physically interact to regulate a podocyte-specific gene program. These findings further our understanding of the transcriptional circuitry responsible for podocyte formation and differentiation during kidney development.
authors.affiliation: Center for Regenerative Medicine and Department of Medicine, Massachusetts General Hospital, Harvard Medical School and Harvard Stem Cell Institute, Boston, MA 02114, USA.
authors.forename: Lori L
authors.initials: LL
authors.lastname: O'Brien
authors.forename: Michael
authors.initials: M
authors.lastname: Grimaldi
authors.forename: Zachary
authors.initials: Z
authors.lastname: Kostun
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Wingert
authors.forename: Rori
authors.initials: R
authors.lastname: Selleck
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Davidson
mesh_terms: D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D016373:DNA, Antisense / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D051858:Forkhead Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D008954:Models, Biological; D050199:Podocytes / Q000166:cytology* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D054730:Protein Interaction Domains and Motifs; D051880:Receptors, Notch / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D025721:WT1 Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D016373:DNA, Antisense; D051858:Forkhead Transcription Factors; D051819:Immunoglobulin J Recombination Signal Sequence-Binding Protein; D051880:Receptors, Notch; D012097:Repressor Proteins; D025721:WT1 Proteins; D029961:Zebrafish Proteins; C426857:Foxc1a protein, zebrafish; C405694:Hairy, HRT1 protein