Abstract
The papilla of the mammalian kidney must tolerate greatly varying degrees of hyperosmotic stress during urine concentration and depending on whole organism hydration state. To identify proteome adaptations supporting cell function and survival in such a harsh environment we compared the proteome of a) the hyperosmotic renal papilla with that of adjacent iso-osmotic cortex tissue and b) the renal papilla of diuretic versus that of anti-diuretic mice. Though functionally distinct the papilla is in close physical proximity to the renal cortex, an iso-osmotic region. Proteomic differences between the papilla and cortex of C57BL6 mice were identified using two-dimensional gel electrophoresis and MALDI-TOF/TOF mass spectrometry. We found 37 different proteins characteristic of the cortex and 16 proteins over-represented in the papilla. Regional specificity was confirmed by Western blot and further substantiated by immunohistochemistry for selected proteins. Proteins that are characteristic of the renal papilla include αB crystallin, Hsp beta-1, Hsp90, 14-3-3 protein, glutathione S-transferase, aldose reductase, actin and tropomyosin. Gene ontology analysis confirmed a significant increase in molecular functions associated with protein chaperoning and cell stabilization. Proteins over-represented in the cortex were largely related to routine metabolism. During antidiuresis 15 different proteins changed significantly while 18 different proteins changed significantly during diuresis relative to normally hydrated controls. Changes were confirmed by Western blot for selected proteins. Proteins that are significantly altered by diuretic state are associated with cell structure (actin, tubulin), signaling (Rho GDP dissociation inhibitor, abhydrolase domain-containing protein 14B), chaperone functioning (Hsp beta-1, αB crystallin, T complex protein-1) and anti-oxidant functions (α-enolase, GAPDH and LDH). Taken together our study reveals that specific proteins involved in protein folding, cytoskeletal stabilization, antioxidant responses, and stress signaling contribute greatly to the unique hyperosmotic stress resistant phenotype of the kidney papilla.