The survival of cells in the mammalian inner medulla requires an adaptive response to the hypertonic environment. This adaptation to changes in tonicity initially involves the activation of protein kinases and ultimately the upregulation of proteins that allow for cellular accumulation of inert osmolytes. It is now evident that a more complex array of both genes and proteins is involved in the adaptive response. The present proposal intends to examine the repertoire of genes and proteins that are both increased and decreased in cells adapted to live in hypertonic conditions (600 and 900 mOsm/kg) and in cells acutely (24- 48 hours) exposed to increased tonicity. In this setting the role of various signaling pathways in the genetic regulation will be assessed by employing inhibitors of such pathways as well as transfected cells with both gain and loss of function mutations. The in-vitro results will also be extended to the study in-vivo in rodents in various states of water balance. Both genomic and proteomic approaches are contemplated to unveil as full a repertoire as the presently available techniques allow. In its second aim the proposal will examine the mechanism whereby the subunit of Na-K-ATPase is upregulated by hypertonicity and how the protein provides a survival advantage in these conditions. More specifically, the transcriptional regulation of the gene will be studied. We hypothesize that the effect of JNK kinase to enhance gamma subunit synthesis is regulated at the transcriptional level. The hypothesis will be studied by employing inhibitors of JNK kinase, gain and loss of function transfectents of the kinase and knockout mice deficient in JNK1 and JNK2. The translational regulation will focus on the role of PI3 kinase in the process. The role of the subunit in osmoprotection will be studied by comparing the effects of hypertonicity in wild type cells to that of cells rendered incapable of synthesizing the protein by a silencing RNA technique. This approach could be extended to the study of relevant genes unveiled in the genomics-proteomic studies. Effects on alteration in cell Na concentration, uptake of an inert osmolyte (inositol) and finally cell survival itself will be assessed. These experiments should define genes and proteins involved in osmoregulation and particularly unveil the role of the subunit as a vital component in the adaption to the hypertonic environment of the mammalian inner medulla.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Cellular and Molecular Biology of the Kidney Study Section (CMBK)
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Ketchum, Christian J
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University of Colorado Denver
Internal Medicine/Medicine
Schools of Medicine
United States
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Andres-Hernando, Ana; Lanaspa, Miguel A; Li, Nanxing et al. (2010) Effects of 2-bromoethanamine on TonEBP expression and its possible role in induction of renal papillary necrosis in mice. Toxicol Sci 118:510-20
Lanaspa, Miguel A; Andres-Hernando, Ana; Rivard, Christopher J et al. (2009) ZAC1 is up-regulated by hypertonicity and decreases sorbitol dehydrogenase expression, allowing accumulation of sorbitol in kidney cells. J Biol Chem 284:19974-81
Berl, Tomas (2009) How do kidney cells adapt to survive in hypertonic inner medulla? Trans Am Clin Climatol Assoc 120:389-401
Klawitter, Jost; Rivard, Christopher J; Brown, Lewis M et al. (2008) A metabonomic and proteomic analysis of changes in IMCD3 cells chronically adapted to hypertonicity. Nephron Physiol 109:p1-10
Andres-Hernando, Ana; Lanaspa, Miguel A; Rivard, Christopher J et al. (2008) Nucleoporin 88 (Nup88) is regulated by hypertonic stress in kidney cells to retain the transcription factor tonicity enhancer-binding protein (TonEBP) in the nucleus. J Biol Chem 283:25082-90
Lanaspa, Miguel A; Andres-Hernando, Ana; Rivard, Christopher J et al. (2008) Hypertonic stress increases claudin-4 expression and tight junction integrity in association with MUPP1 in IMCD3 cells. Proc Natl Acad Sci U S A 105:15797-802
Rivard, Christopher J; Brown, Lewis M; Almeida, Nestor E et al. (2007) Expression of the calcium-binding protein S100A4 is markedly up-regulated by osmotic stress and is involved in the renal osmoadaptive response. J Biol Chem 282:6644-52
Lanaspa, Miguel A; Almeida, Nestor E; Andres-Hernando, Ana et al. (2007) The tight junction protein, MUPP1, is up-regulated by hypertonicity and is important in the osmotic stress response in kidney cells. Proc Natl Acad Sci U S A 104:13672-7
Jimenez, Carlos; Cossio, Belen R; Rivard, Christopher J et al. (2007) Cell division in the unicellular microalga Dunaliella viridis depends on phosphorylation of extracellular signal-regulated kinases (ERKs). J Exp Bot 58:1001-11
Capasso, Juan M; Rivard, Christopher J; Berl, Tomas (2006) Silencing and overexpression of the gamma-subunit of Na-K-ATPase directly affect survival of IMCD3 cells in response to hypertonic stress. Am J Physiol Renal Physiol 291:F1142-7

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