The kidney collecting system arises through branching morphogenesis of the ureteric bud (UB), a process that we have been able to replicate in vitro by culturing the isolated UB in the presence of a metanephric mesenchyme cell conditioned medium. Our purification of proteins from this conditioned medium during the previous funding period has demonstrated that multiple soluble factors regulate UB branching. Here we argue that the ability of these growth factors to generate clear distinctions between UB tips and stalks is critical for normal collecting system development. We therefore seek to identify the cellular and molecular mechanisms by which particular growth factors work to generate tips and stalks (SA1). Since we have found that different sets of growth factors give very distinct patterns of isolated UB morphogenesis--""""""""fine branching (tips and stalks),"""""""" """"""""stalk-centric"""""""" and """"""""tip-centric"""""""" phenotypes-we will use these clearly different conditions to identify molecules and mechanisms that lead to tip and stalk specification. In the proposed studies, we plan to connect the growth factors we have identified to cell surface signaling (via GFRalpha1 and lipid rafts) and intracellular signaling pathways (via Rho/ROCK), as well as intracellular protein sorting pathways (especially those involving ER chaperones), necessary for appropriate expression of tip and stalk-specific gene products. The logic of these connections, based largely on work completed during the previous funding period, is developed in detail in the application. (SA1 techniques: isolated UB culture, density gradient fractionation, transfections, selective down regulation of mRNA, laser capture microdissection, microarrays). In addition, we have identified at least one remaining potent activity for UB branching in vitro and aim to purify and characterize it. (SA2 techniques: column chromatography, mass spectrometry, protein expression). Together, the proposed studies should provide a much more complete picture of the soluble factors that regulate UB branching and how they act at the cell surface and intracellularly to generate tips and stalks during collecting system development. The experiments are based on considerable preliminary data and follow directly from work completed during the previous funding period. We have proven expertise in the techniques necessary to complete the project.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Urologic and Kidney Development and Genitourinary Diseases Study Section (UKGD)
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Hoshizaki, Deborah K
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University of California San Diego
Schools of Medicine
La Jolla
United States
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Gallegos, Thomas F; Kouznetsova, Valentina; Kudlicka, Krystyna et al. (2012) A protein kinase A and Wnt-dependent network regulating an intermediate stage in epithelial tubulogenesis during kidney development. Dev Biol 364:11-21
Bush, Kevin T; Crawford, Brett E; Garner, Omai B et al. (2012) N-sulfation of heparan sulfate regulates early branching events in the developing mammary gland. J Biol Chem 287:42064-70
Shah, Mita M; Sakurai, Hiroyuki; Gallegos, Thomas F et al. (2011) Growth factor-dependent branching of the ureteric bud is modulated by selective 6-O sulfation of heparan sulfate. Dev Biol 356:19-27
Garner, Omai B; Bush, Kevin T; Nigam, Kabir B et al. (2011) Stage-dependent regulation of mammary ductal branching by heparan sulfate and HGF-cMet signaling. Dev Biol 355:394-403
Rosines, Eran; Johkura, Kohei; Zhang, Xing et al. (2010) Constructing kidney-like tissues from cells based on programs for organ development: toward a method of in vitro tissue engineering of the kidney. Tissue Eng Part A 16:2441-55
Tee, James B; Choi, Yohan; Shah, Mita M et al. (2010) Protein kinase A regulates GDNF/RET-dependent but not GDNF/Ret-independent ureteric bud outgrowth from the Wolffian duct. Dev Biol 347:337-47
Shah, Mita M; Sakurai, Hiroyuki; Sweeney, Derina E et al. (2010) Hs2st mediated kidney mesenchyme induction regulates early ureteric bud branching. Dev Biol 339:354-65
Crawford, Brett E; Garner, Omai B; Bishop, Joseph R et al. (2010) Loss of the heparan sulfate sulfotransferase, Ndst1, in mammary epithelial cells selectively blocks lobuloalveolar development in mice. PLoS One 5:e10691
Shah, Mita M; Tee, James B; Meyer, Tobias et al. (2009) The instructive role of metanephric mesenchyme in ureteric bud patterning, sculpting, and maturation and its potential ability to buffer ureteric bud branching defects. Am J Physiol Renal Physiol 297:F1330-41
Nigam, Sanjay K; Shah, Mita M (2009) How does the ureteric bud branch? J Am Soc Nephrol 20:1465-9

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