Compensatory membrane expression of the V-ATPase B2 subunit isoform in renal medullary intercalated cells of B1-deficient mice

Abstract

Compensatory membrane expression of the V-ATPase B2 subunit isoform in renal medullary intercalated cells of B1-deficient mice. Am J Physiol Renal Physiol 293: F1915-F1926, 2007. First published September 26, 2007; doi:10.1152/ajprenal.00160.2007.-Mice deficient in the ATP6V1B1 ("B1") subunit of the vacuolar protonpumping ATPase (V-ATPase) maintain body acid-base homeostasis under normal conditions, but not when exposed to an acid load. Here, compensatory mechanisms involving the alternate ATP6V1B2 ("B2") isoform were examined to explain the persistence of baseline pH regulation in these animals. By immunocytochemistry, the mean pixel intensity of apical B2 immunostaining in medullary A intercalated cells (A-ICs) was twofold greater in B1Ϫ/Ϫ mice than in B1ϩ/ϩ animals, and B2 was colocalized with other V-ATPase subunits. No significant upregulation of B2 mRNA or protein expression was detected in B1Ϫ/Ϫ mice compared with wild-type controls. We conclude that increased apical B2 staining is due to relocalization of B2-containing V-ATPase complexes from the cytosol to the plasma membrane. Recycling of B2-containing holoenzymes between these domains was confirmed by the intracellular accumulation of B1-deficient V-ATPases in response to the microtubule-disrupting drug colchicine. V-ATPase membrane expression is further supported by the presence of "rod-shaped" intramembranous particles seen by freeze fracture microscopy in apical membranes of normal and B1-deficient A-ICs. Intracellular pH recovery assays show that significant (28 -40% of normal) V-ATPase function is preserved in medullary ICs from B1Ϫ/Ϫ mice. We conclude that the activity of apical B2-containing V-ATPase holoenzymes in A-ICs is sufficient to maintain baseline acid-base homeostasis in B1-deficient mice. However, our results show no increase in cell surface V-ATPase activity in response to metabolic acidosis in ICs from these animals, consistent with their inability to appropriately acidify their urine under these conditions. proton pump; immunofluorescence; pH homeostasis; urinary acidification; Atp6v1b1Ϫ/Ϫ mice THE MAIN MEDIATOR OF INTRACELLULAR organelle acidification in eukaryotic cells and of proton (H ϩ ) secretion along the distal renal nephron is the ubiquitous vacuolar proton-pumping ATPase (vacuolar, or V-type, H ϩ -ATPase, or V-ATPase). The V-ATPase is a complex enzyme, consisting of two large sectors or domains (V 0 , the transmembrane domain involved in H ϩ translocation, and V 1 , the cytosolic domain, responsible for hydrolyzing ATP), which together contain at least 13 distinct

Renal β-intercalated cells maintain body fluid and electrolyte balance

“…Atp6v1b1 disruption leads to markedly reduced proton pump activity in α-ICs (19,20) and possibly also in β-ICs. Pharmacologic blockade of the pump has been shown to inhibit pendrin-dependent Cl -absorption by cortical collecting duct (CCDs) (21).…”

“…It has been shown that the B2 subunit, which is normally co-expressed with the B1 subunit in ICs, compensates for the lack of the B1 subunit of the H + -ATPase in α-ICs and that its activity is sufficient to maintain acid base homeostasis in Atp6v1b1 -/-mice under basal conditions but not in conditions of acid load (20,35) or stimulation of angiotensin II (36). Our data showing no electroneutral NaCl transport in the CCD of Atp6v1b1 -/-mice suggested that the B2 isoform is not capable of compensating for the absence of B1 in β-ICs under all circumstances (normal-and low-salt diet), conditions that may require angiotensin II mediated regulation of H + -ATPases.…”

Evaluation of Bone Marrow- and Brain-Derived Neural Stem Cells in Therapy of Central Nervous System Autoimmunity

“…To quantify the amount of MBP expression, pixel intensity of MBP immunofluorescence was measured according to previous reports. 26,27 MBP pixel intensity was measured at random areas of disease lesions at low power in the same image size by using ImageJ software (NIH ImageJ; National Institutes of Health, Bethesda, MD). Total axon numbers did not differ significantly among measured areas.…”

Regulation of luminal acidification in the male reproductive tract via cell–cell crosstalk

“…For example, can another a isoform replace the mutated a4 in the proximal tubule and inner ear? Such isoform replacement occurs in the absence of B1 in B1-knockout (KO) mice, where the compensatory insertion of B2 with the plasma membrane-bound V-ATPase allows VATPase-dependent proton transport to occur across the membrane of epididymal clear cells and renal intercalated cells (Da Silva et al, 2007a;Paunescu et al, 2007). Accordingly, male mice lacking the B1 subunit do not develop dRTA and are not infertile (Da Silva et al, 2007a).…”