p38 Kinase Activity Is Essential for Osmotic Induction of mRNAs for HSP70 and Transporter for Organic Solute Betaine in Madin-Darby Canine Kidney Cells

Abstract

In renal cells, hypertonicity induces genes for heat shock proteins (HSP70, ␣B-crystallin), as well as enzymes and transporters directly involved in the metabolism and transport of protective organic osmolytes. While heat shock proteins are induced by many stresses including osmotic stress, the induction of the osmolytes genes appears to be specific to osmotic stress. These two adaptive mechanisms allow kidney cells to survive and function in the hypertonic environment that exists on routine basis in kidney medulla. In mammalian cells, hypertonicity induces three mitogen-activated protein kinase pathways: ERK (extracellular regulated kinase), JNK (Jun N-terminal kinase), and p38. ERK activation by osmotic stress is a consistent finding in many cells, but it is not essential for transcriptional regulation of mRNA for transporter of organic osmolyte betaine. While the growth of yeast cells on NaCl-supplemented medium is dependent on HOG1 pathway, it is still unclear which pathway mediates the adaptation to osmotic stress in mammalian cells. Here, we show that inhibition of p38 kinase activity, using the specific inhibitor SB203580 (4-(fluorophenyl)-2-(4-methylsulfonylphenyl)-5-(4-pyridyl) imidazole), abolishes the hypertonicity-mediated induction of mRNAs for HSP70 and betaine transporter in Madin-Darby canine kidney cells. The inhibition is dose-dependent and correlates with the in situ activity of native p38 kinase, determined as MAPKAPK-2 activity in cell extracts. As reported previously, the activities of ERK-1 and -2 were not affected by SB203580, but surprisingly, inhibition of native p38 kinase activity correlates with up-regulation of native JNK-1 activity in osmotically stressed cells. p38 mRNA is induced by hypertonic stress and is attenuated with p38 kinase inhibition. We also find that thermal induction of HSP70 mRNA is not affected by p38 kinase inhibition. Such findings suggest that p38 kinase activity is essential for the induction of genes involved in the adaptation of mammalian cells to osmotic stress and that the increased activity of JNK-1 during p38 kinase inhibition is consistent with regulation of JNK-1 by p38 kinase in osmotically stressed cells. In addition, the transduction pathways mediating HSP70 mRNA induction by different stresses appear to be divergent; osmotic induction of HSP70 is p38 kinase-dependent, while thermal induction is not.During water deprivation, the extracellular osmolality in the mammalian renal medulla can exceed 3000 mosmol/kg of H 2 O. While such an osmolality is incompatible with the survival of cells from other organs, kidney cells tolerate it well. The survival of kidney medullary cells in hypertonic environment is essential for the generation of the concentrating gradient, which is key for maintenance of body solute and water homeostasis.Many organisms, including bacteria, yeast, plants, and animals, adapt to sustained hyperosmotic stress by accumulating osmotically active organic solutes (compatible organic osmolytes) (1). These compounds do not perturb cellula...

Adaptive Increase of Amino Acid Transport System A Requires ERK1/2 Activation

“…A p38-dependent pathway was recently implicated in the hypertonic induction of betaine and myoinositol transport in human monocytes (43) and of betaine transport in Madin-Darby canine kidney cells (44). However, although cell volume changes were not monitored in either of those reports, cell transfer to hypertonic medium is expected to cause an abrupt shrinkage rather than a gradual and progressive decrease of cell volume, as observed here.…”

“…The p38 inhibitor SB203580 (1 M), added to the extracellular medium 60 min before amino acid-free incubation and maintained thereafter, produced a 60% inhibition of adaptive increase in system A transport activity. 2 However, the interpretation of this result is made difficult by the marked activation of JNK1 promoted by the inhibitor in our cells 2 as in other models (44). ERK1/2 kinases are markedly activated when cultured human fibroblasts (this report) or other cells (20 -22) are incubated under hypertonic conditions.…”

Abnormal p38 mitogen-activated protein kinase signalling in human and experimental diabetic nephropathy

“…High glucose stimulation of cultured proximal tubular epithelial cells results in p38-dependent angiotensinogen production and p38-independent hypertrophy [37,38]. In addition, high glucose levels can increase the hypertonicity of the tubular fluid, resulting in greater osmotic stress to tubules, which is known to induce p38 phosphorylation in cultured tubular cells [39]. In the current study, dilated tubules in diabetic kidneys appeared to have an increased proportion of p-p38+ cells, suggesting that increased phosphorylation of tubular p38 also occurs in response to tubular injury.…”

Independent Signaling Pathways Regulate Cellular Turgor during Hyperosmotic Stress and Appressorium-Mediated Plant Infection by Magnaporthe grisea

“…The resulting accumulation of glycerol in yeast cells maintains cellular turgor even after exposure to severe hyperosmotic stress. Elements of the HOG pathway have also been found in mammals, Drosophila , and plants (Han et al, 1994(Han et al, , 1998Popping et al, 1996) and shown either to be required for hyperosmotic adaptation (Sheikh Hamad et al, 1998) or to be capable of complementing corresponding yeast mutants (Kumar et al, 1995; Han et al, 1998).…”