Regina Saum

The vacuolar (H(+))-ATPases are ATP-dependent proton pumps that acidify intracellular compartments and, in some cases, transport protons across the plasma membrane of eukaryotic cells. Intracellular V-ATPases play an important role in normal physiological processes such as receptor-mediated endocytosis, intracellular membrane trafficking, pro-hormone processing, protein degradation, and the coupled uptake of small molecules, such as neurotransmitters. They also function in the entry of various pathogenic agents, including many envelope viruses, like influenza virus, and toxins, like anthrax toxin. Plasma membrane V-ATPases function in renal pH homeostasis, bone resorption and sperm maturation, and various disease processes, including renal tubular acidosis, osteopetrosis, and tumor metastasis. V-ATPases are composed of a peripheral V(1) domain containing eight different subunits that is responsible for ATP hydrolysis and an integral V(0) domain containing six different subunits that translocates protons. In mammalian cells, most of the V-ATPase subunits exist in multiple isoforms which are often expressed in a tissue specific manner. Isoforms of one of the V(0) subunits (subunit a) have been shown to possess information that targets the V-ATPase to distinct cellular destinations. Mutations in isoforms of subunit a lead to the human diseases osteopetrosis and renal tubular acidosis. A number of mechanisms are employed to regulate V-ATPase activity in vivo, including reversible dissociation of the V(1) and V(0) domains, control of the tightness of coupling of proton transport and ATP hydrolysis, and selective targeting of V-ATPases to distinct cellular membranes. Isoforms of subunit a are involved in regulation both via the control of coupling and via selective targeting. This review will begin with a brief introduction to the function, structure, and mechanism of the V-ATPases followed by a discussion of the role of V-ATPase subunit isoforms and the mechanisms involved in regulation of V-ATPase activity.

N(epsilon)-acetyl-beta-lysine is a unique compatible solute found in methanogenic archaea grown at high salinities. Deletion of the genes that encode the lysine-2,3-aminomutase (ablA) and the beta-lysine acetyltransferase (ablB) abolished the production of N(epsilon)-acetyl-beta-lysine in Methanosarcina mazei Gö1. The mutant grew well at low and intermediate salinities. Interestingly, growth at high salt (800 mM NaCl) was only slowed down but not impaired demonstrating that in M. mazei Gö1 N(epsilon)-acetyl-beta-lysine is not essential for growth at high salinities. Nuclear magnetic resonance (NMR) analysis revealed an increased glutamate pool in the mutant. In addition to alpha-glutamate, a novel solute, alanine, was produced. The intracellular alanine concentration was as high as 0.36 +/- 0.05 micromol (mg protein)-1 representing up to 18% of the total solute pool at 800 mM NaCl. The cellular alanine concentration increased with the salinity of the medium and decreased in the presence of glycine betaine in the medium, indicating that alanine is used as compatible solute by M. mazei Gö1.

There is a long-standing discussion in the literature, based on biochemical and genomic data, whether some archaeal species may have two structurally and functionally distinct ATP synthases in one cell: the archaeal A(1)A(O) together with the bacterial F(1)F(O) ATP synthase. To address a potential role of the bacterial F(1)F(O) ATP synthase, we have exchanged the F(1)F(O) ATPase gene cluster in Methanosarcina acetivorans against a puromycin resistance cassette. Interestingly, the mutant was able to grow with no difference in growth kinetics to the wild type, and cellular ATP contents were identical in the wild type and the mutant. These data demonstrate that the F(1)F(O) ATP synthase is dispensable for the growth of M. acetivorans.