Rotating with the brakes on and other unresolved features of the vacuolar ATPase

The rotary ATPase family is comprised of the ATP synthase (F-ATPase), vacuolar ATPase (V-ATPase) and acrahael ATPase (A-ATPase). These either predominantly utilise a proton gradient for ATP synthesis or use ATP to produce a proton gradient, driving secondary transport and acidifying organelles. With advances in electron microscopy (EM) has come a significant increase in our understanding of the rotary ATPase family. Following the sub nm resolution reconstructions of both the F and V-ATPase the secondary structure organisation of the elusive subunit a has now been resolved, revealing a novel helical arrangement. Despite these significant developments in our understanding of the rotary ATPases there are still a number of unresolved questions about the mechanism, regulation, and overall architecture, which this mini-review aims to highlight and discuss

Characterization of the N-ATPase, a distinct, laterally transferred Na+-translocating form of the bacterial F-type membrane ATPase

An analysis of the distribution of the Na+-translocating ATPases/ATP synthases among microbial genomes identified an atypical form of the F1Fo-type ATPase that is present in the archaea Methanosarcina barkeri and M.acetivorans, in a number of phylogenetically diverse marine and halotolerant bacteria and in pathogens Burkholderia spp. In complete genomes, representatives of this form (referred to here as N-ATPase) are always present as second copies, in addition to the typical proton-translocating ATP synthases. The N-ATPase is encoded by a highly conserved atpDCQRBEFAG operon and its subunits cluster separately from the equivalent subunits of the typical F-type ATPases. N-ATPase c subunits carry a full set of sodium-binding residues, indicating that most of these enzymes are Na+-translocating ATPases that likely confer on their hosts the ability to extrude Na+ ions. Other distinctive properties of the N-ATPase operons include the absence of the delta subunit from its cytoplasmic sector and the presence of two additional membrane subunits, AtpQ (formerly gene 1) and AtpR (formerly gene X). We argue that N-ATPases are an early-diverging branch of membrane ATPases that, similarly to the eukaryotic V-type ATPases, do not synthesize ATP

Inhibition of Iron Uptake Is Responsible for Differential Sensitivity to V-ATPase Inhibitors in Several Cancer Cell Lines

Many cell lines derived from tumors as well as transformed cell lines are far more sensitive to V-ATPase inhibitors than normal counterparts. The molecular mechanisms underlying these differences in sensitivity are not known. Using global gene expression data, we show that the most sensitive responses to HeLa cells to low doses of V-ATPase inhibitors involve genes responsive to decreasing intracellular iron or decreasing cholesterol and that sensitivity to iron uptake is an important determinant of V-ATPase sensitivity in several cancer cell lines. One of the most sensitive cell lines, melanoma derived SK-Mel-5, over-expresses the iron efflux transporter ferroportin and has decreased expression of proteins involved in iron uptake, suggesting that it actively suppresses cytoplasmic iron. SK-Mel-5 cells have increased production of reactive oxygen species and may be seeking to limit additional production of ROS by iron

Extracellular and Luminal pH Regulation by Vacuolar H⁺-ATPase Isoform Expression and Targeting to the Plasma Membrane and Endosomes

Plasma membrane vacuolar H+ -ATPase (pm-V-ATPase) activity of tumor cells is a major factor in control of cytoplasmic and extracellular pH and metastatic potential, but the isoforms involved and the factors governing plasma membrane recruitment remain uncertain. Here, we examined expression, distribution and activity of V- ATPase isoforms in invasive prostate adenocarcinoma (PC-3) cells. Isoforms 1 and 3 were the most highly expressed forms of membrane subunit a, with a1 and a3 the dominant plasma membrane isoforms. Correlation between pm-V-ATPase activity and invasiveness was limited, but RNAi knockdown of either a isoform did slow cell proliferation and inhibit invasion in vitro. Isoform a1 was recruited to the cell surface from the early endosome/recycling complex pathway, its knockdown arresting transferrin receptor (TfR) recycling. Isoform a3 was associated with the late endosomal/lysosomal compartment. Both a isoforms associated with accessory protein Ac45, knockdown of which stalled transit of a1 and Tf-TfR, decreased proton efflux and reduced cell growth and invasiveness, this latter effect at least partly due to decreased delivery of the membrane-bound matrix metalloproteinase MMP-14 to the plasma membrane. These data indicate that in prostatic carcinoma cells, a1 and a3 isoform populations predominate in different compartments where they maintain different luminal pH. Ac45 plays a central role in navigating the V-ATPase to the plasma membrane, and hence is an important factor in expression of the invasive phenotype