Sunny Hartwig

The Pro-Apoptotic Protein Bim Is a MicroRNA Target in Kidney Progenitors

Understanding the mechanisms that regulate nephron progenitors during kidney development should aid development of therapies for renal failure. MicroRNAs, which modulate gene expression through post-transcriptional repression of specific target mRNAs, contribute to the differentiation of stem cells, but their role in nephrogenesis is incompletely understood. Here, we found that the loss of miRNAs in nephron progenitors results in a premature depletion of this population during kidney development. Increased apoptosis and expression of the pro-apoptotic protein Bim accompanied this depletion. Profiling of miRNA expression during nephrogenesis identified several highly expressed miRNAs (miR-10a, miR-106b, miR-17-5p) in nephron progenitors that are either known or predicted to target Bim. We propose that modulation of apoptosis by miRNAs may determine congenital nephron endowment. Furthermore, our data implicate the pro-apoptotic protein Bim as a miRNA target in nephron progenitors.

p38MAPK Acts in the BMP7-dependent Stimulatory Pathway during Epithelial Cell Morphogenesis and Is Regulated by Smad1

Bone morphogenetic protein (BMP)-7 exerts dose-dependent stimulatory and inhibitory effects during renal branching morphogenesis. Previously, we identified an inhibitory role for activin-like kinase receptors and Smad1 in BMP-dependent inhibition (Piscione, T. D., Phan, T., and Rosenblum, N. D. (2001) Am. J. Physiol. 280, F19–F33). Here we demonstrate a novel role for p38 mitogen-activated kinase (p38MAPK) in BMP7-dependent stimulatory signaling. Stimulatory doses (0.25 nm) of BMP7 increased p38MAPK activity and stimulated phosphorylation of endogenous activating transcription factor 2 (ATF2) in a p38MAPK-dependent manner in murine inner medullary collecting duct (mIMCD-3) cells. In contrast, high doses (10 nm) of BMP7 inhibited p38MAPK activity and phosphorylation of endogenous ATF2. Treatment with BMP7 exerted no significant effect on the levels of the phosphorylated forms of endogenous SAPK/JNK or p44 and p42 (ERK1 and ERK2) protein kinases. To investigate the functional importance of p38MAPK signaling, we showed that SB203580, a p38MAPK inhibitor, blocked the stimulatory effect of BMP7 on mIMCD-3 cell morphogenesis but had no effect on BMP7-dependent inhibition in a three-dimensional culture model. To identify mechanisms by which BMP7-dependent inhibitory signaling suppresses p38MAPK activity, we measured p38MAPK activity in ligand independent mIMCD-3 models of enhanced and suppressed Smad signaling. Basal activity of p38MAPK was decreased in mIMCD-3 cells and in embryonic kidney tissue expressing a constitutively active activin-like kinase receptor, but was increased in mIMCD-3 cells stably expressing a dominant negative form of Smad1. We conclude that BMP7 stimulates renal epithelial cell morphogenesis via p38MAPK and that p38MAPK activity is negatively regulated by Smad1.

BMP Receptor ALK3 Controls Collecting System Development

The molecular signals that regulate growth and branching of the ureteric bud during formation of the renal collecting system are largely undefined. Members of the bone morphogenetic protein (BMP) family signal through the type I BMP receptor ALK3 to inhibit ureteric bud and collecting duct cell morphogenesis in vitro. We investigated the function of the BMP signaling pathway in vivo by generating a murine model of ALK3 deficiency restricted to the ureteric bud lineage (Alk3(UB-/-) mice). At the onset of branching morphogenesis, Alk3(UB-/-) kidneys are characterized by an abnormal primary (1 degrees ) ureteric bud branch pattern and an increased number of ureteric bud branches. However, during later stages of renal development, Alk3(UB-/-) kidneys have fewer ureteric bud branches and collecting ducts than wild-type kidneys. Postnatal Alk3(UB-/-) mice exhibit a dysplastic renal phenotype characterized by hypoplasia of the renal medulla, a decreased number of medullary collecting ducts, and abnormal expression of beta-catenin and c-MYC in medullary tubules. In summary, normal kidney development requires ALK3-dependent BMP signaling, which controls ureteric bud branching.

Integrin-Linked Kinase Mediates Bone Morphogenetic Protein 7-Dependent Renal Epithelial Cell Morphogenesis

ABSTRACT Bone morphogenetic protein 7 (BMP7) stimulates renal branching morphogenesis via p38 mitogen-activated protein kinase (p38MAPK) and activating transcription factor 2 (ATF-2) (M. C. Hu, D. Wasserman, S. Hartwig, and N. D. Rosenblum, J. Biol. Chem. 279:12051-12059, 2004). Here, we demonstrate a novel role for integrin-linked kinase (ILK) in mediating renal epithelial cell morphogenesis in embryonic kidney explants and identify p38MAPK as a target of ILK signaling in a cell culture model of renal epithelial morphogenesis. The spatial and temporal expression of ILK in embryonic mouse kidney cells suggested a role in branching morphogenesis. Adenovirus-mediated expression of ILK stimulated and expression of a dominant negative ILK mutant inhibited ureteric bud branching in embryonic mouse kidney explants. BMP7 increased ILK kinase activity in inner medullary collecting duct 3 (IMCD-3) cells, and adenovirus-mediated expression of ILK increased IMCD-3 cell morphogenesis in a three-dimensional culture model. In contrast, treatment with a small molecule ILK inhibitor or expression of a dominant negative-acting ILK (ILKE359K) inhibited epithelial cell morphogenesis. Further, expression of ILKE359K abrogated BMP7-dependent stimulation. To investigate the role of ILK in BMP7 signaling, we showed that ILK overexpression increased basal and BMP7-induced levels of phospho-p38MAPK and phospho-ATF-2. Consistent with its inhibitory effects on IMCD-3 cell morphogenesis, expression of ILKE359K blocked BMP7-dependent increases in phospho-p38MAPK and phospho-ATF-2. Inhibition of p38MAPK activity with the specific inhibitor, SB203580, failed to inhibit BMP7-dependent stimulation of ILK activity, suggesting that ILK functions upstream of p38MAPK during BMP7 signaling. We conclude that ILK functions in a BMP7/p38MAPK/ATF-2 signaling pathway and stimulates epithelial cell morphogenesis.

Glypican-3 modulates inhibitory Bmp2-Smad signaling to control renal development in vivo.

Renal branching morphogenesis, defined as growth and branching of the ureteric bud (UB), is a tightly regulated process controlled by growth factor-dependent tissue interactions. Previously, using in vitro models of branching morphogenesis, we demonstrated that BMP2 signals via its intracellular effectors, SMAD1 and SMAD4, to control UB cell proliferation and branching in a manner modulated by Glypican-3 (GPC3), a cell surface heparan sulfate proteoglycan. Here, we used loss-of-function genetic mouse models to investigate the functions of Bmp2 and Gpc3-Bmp2 interactions in vivo. Progressively greater increases in UB cell proliferation were observed in Bmp2+/-, Smad4+/-, and Bmp2+/-; Smad4+/- mice compared to Wt. This increased cell proliferation was accompanied by a significant increase in UB branching in Smad4+/- and Bmp2+/-;Smad4+/- mice compared to Wt. Reduction of Gpc3 gene dosage also increased UB cell proliferation, an effect that was enhanced in Gpc3+/-;Bmp2+/- mice to an extent greater than the sum of that observed in Gpc3+/- and Bmp2+/- mice. Reduction of both Gpc3 and Bmp2 gene dosage enhanced cell proliferation in the metanephric mesenchyme compared to Wt, an effect not observed in either Bmp2+/- or Gpc3+/- mice. Phosphorylation of SMAD1, a measure of SMAD1 activation, was progressively decreased in Gpc3+/- and Gpc3+/-;Bmp2+/- mice compared to Wt, suggesting that Gpc3 stimulates Bmp2-dependent SMAD signaling in vivo. These results demonstrate that BMP2-SMAD signaling, modulated by GPC3, inhibits renal branching morphogenesis in vivo.

Bone morphogenetic protein signaling in the developing kidney: present and future

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-beta superfamily. A critical role for BMP signaling in the development of the metanephric kidney is supported by a growing number of studies using in vitro assays and in vivo animal models. Here we review current knowledge of BMPs, BMP receptors and regulators of the BMP signaling pathway in the developing kidney. We highlight major gaps in our knowledge of the roles of BMP signaling in the development of the normal and abnormal kidney and identify areas and techniques likely to improve our understanding.

Developmental origins and functions of stromal cells in the normal and diseased mammalian kidney.

The kidney is a model developmental paradigm of vertebrate organogenesis. As in many other organs, kidney development involves reciprocal inductive tissue interactions between multiple cell lineages. The most well defined of these interactions occurs between the ureteric bud and the nephrogenic mesenchyme. A population of mesenchymal cells distinct from nephrogenic precursors and termed stromal cells, have been relatively understudied. Yet existing knowledge indicates that stromal cells are critical regulators in the normal and diseased kidney. This commentary reviews current knowledge regarding the origin and functional roles of the stromal cell population during kidney development. Gaps in our current understanding of renal stromal cells and future directions needed to advance this expanding field of study are highlighted. Developmental Dynamics 243:853–863, 2014.

Wilms' tumor-1: a riddle wrapped in a mystery, inside a kidney

After more than 15 years of intense study, WT1 remains a complex protein with multiple functions and targets. This Commentary discusses new developments and puts past results in perspective.

Canadian Journal of Kidney Health and Disease: a unique launch of a unique journal

Usually inaugural editorials are written by the Editor-in-Chief to describe the scope and vision for the journal to potential authors and readers. This editorial is written by the Editor-in-Chief, the Deputy Editors and the Associate Editors collaboratively as a clear signal that this is a unique and different journal. We will build this journal on a set of principles which are fundamental to improving the outcomes of patients with kidney disease. To that end, we aim to be supportive, to collaborate, to integrate multiple perspectives and to be open to possibilities.RésuméHabituellement, il revient à l’éditeur en chef de rédiger l’éditorial inaugural décrivant la vision et les champs d’intérêts d’un nouveau journal. Le Journal canadien de la santé et de la maladie rénale a choisi de faire les choses autrement. En effet, cet éditorial est le fruit de la collaboration entre l’éditeur en chef et les éditeurs en chef adjoints. Ce journal s’appuie sur des principes qui seront fondamentaux pour améliorer le sort de patients atteints de maladie rénale. Pour y arriver, nous nous engageons à apporter du support aux auteurs, à collaborer, à intégrer différentes perspectives et être ouverts à des nouvelles possibilités.

SOX4 regulates gonad morphogenesis and promotes male germ cell differentiation in mice.

The group C SOX transcription factors SOX4, -11 and -12 play important and mutually overlapping roles in development of a number of organs. Here, we examined the role of SoxC genes during gonadal development in mice. All three genes were expressed in developing gonads of both sexes, predominantly in somatic cells, with Sox4 being most strongly expressed. Sox4 deficiency resulted in elongation of both ovaries and testes, and an increased number of testis cords. While female germ cells entered meiosis normally, male germ cells showed reduced levels of differentiation markers Nanos2 and Dnmt3l and increased levels of pluripotency genes Cripto and Nanog, suggesting that SOX4 may normally act to restrict the pluripotency period of male germ cells and ensure their proper differentiation. Finally, our data reveal that SOX4 (and, to a lesser extent, SOX11 and -12) repressed transcription of the sex-determining gene Sox9 via an upstream testis-specific enhancer core (TESCO) element in fetal gonads, raising the possibility that SOXC proteins may function as transcriptional repressors in a context-dependent manner.