Localization and regulation of the ATP6V0A4 (a4) vacuolar H+-ATPase subunit defective in an inherited form of distal renal tubular acidosis.
Vacuolar-type H(+)-ATPases (V-H(+)-ATPases) are the major H(+)-secreting protein in the distal portion of the nephron and are involved in net H(+) secretion (bicarbonate generation) or H(+) reabsorption (net bicarbonate secretion). In addition, V-H(+)-ATPases are involved in HCO(3)(-) reabsorption in the proximal tubule and distal tubule. V-H(+)-ATPases consist of at least 13 subunits, the functions of which have not all been elucidated. Mutations in the accessory ATP6V0A4 (a4 isoform) subunit have recently been shown to cause an inherited form of distal renal tubular acidosis in humans. Here, the localization of this subunit in human and mouse kidney was studied and the regulation of expression and localization of this subunit in mouse kidney in response to acid-base and electrolyte intake was investigated. Reverse transcription-PCR on dissected mouse nephron segments amplified a4-specific transcripts in proximal tubule, loop of Henle, distal convoluted tubule, and cortical and medullary collecting duct. a4 protein was localized by immunohistochemistry to the apical compartment of the proximal tubule (S1/S2 segment), the loop of Henle, the intercalated cells of the distal convoluted tubule, the connecting segment, and all intercalated cells of the entire collecting duct in human and mouse kidney. All types of intercalated cells expressed a4. NH(4)Cl or NaHCO(3) loading for 24 h, 48 h, or 7 d as well as K(+) depletion for 7 and 14 d had no influence on a4 protein expression levels in either cortex or medulla as determined by Western blotting. Immunohistochemistry, however, demonstrated a subcellular redistribution of a4 in response to the different stimuli. NH(4)Cl and K(+) depletion led to a pronounced apical staining in the connecting segment, cortical collecting duct, and outer medullary collecting duct, whereas NaHCO(3) loading caused a stronger bipolar staining in the cortical collecting duct. Taken together, these results demonstrate a4 expression in the proximal tubule, loop of Henle, distal tubule, and collecting duct and suggest that under conditions in which increased V-H(+)-ATPase activity is required, a4 is regulated by trafficking but not protein expression. This may allow for the rapid adaptation of V-H(+)-ATPase activity to altered acid-base intake to achieve systemic pH homeostasis. The significance of a4 expression in the proximal tubule in the context of distal renal tubular acidosis will require further clarification.
Stehberger
Paul A
PA
Departments of Cellular and Molecular Physiology, Genetics, and Surgery, Yale University School of Medicine, New Haven, Connecticut, USA.
Schulz
Nicole
N
Finberg
Karin E
KE
Karet
Fiona E
FE
Giebisch
Gerhard
G
Lifton
Richard P
RP
Geibel
John P
JP
Wagner
Carsten A
CA
D000136:Acid-Base Equilibrium; D000141:Acidosis, Renal Tubular / Q000201:enzymology / Q000235:genetics*; D000818:Animals; D006801:Humans; D007668:Kidney / Q000737:chemistry; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D011506:Proteins; D006180:Proton-Translocating ATPases; D025262:Vacuolar Proton-Translocating ATPases / Q000032:analysis / Q000378:metabolism*
D011506:Proteins; C490181:ATP6V0A4 protein, human; C490179:Atp6v0a4 protein, mouse; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases