Salicylihalamide A Inhibits the V0 Sector of the V-ATPase through a Mechanism Distinct from Bafilomycin A1

Abstract

The newly identified specific V-ATPase inhibitor, salicylihalamide A, is distinct from any previously identified V-ATPase inhibitors in that it inhibits only mammalian V-ATPases, but not those from yeast or other fungi (Boyd, M. R., Farina, C., Belfiore, P., Gagliardi, S., Kim Acidification of intracellular compartments of eukaryotes is essential for many cellular processes, including receptor-mediated endocytosis, protein degradation in lysosomes, processing of hormones, uptake, and storage of neurotransmitters, and entry of many viruses into cells. The control of pH within these intracellular compartments is mediated by vacuolar H ϩ -translocating ATPases, which acidify organelles of both constitutive and regulated secretory pathways (1-5). V-ATPases 1 are also found in the plasma membrane of certain cells where they are responsible for cell type-specific processes, including urinary acidification and osteoclast-mediated bone resorption (6, 7). The V-type ATPases are among the most widely distributed ATP-driven ion pumps in nature, present in all eukaryotic cells and in various bacteria. Within eukaryotic cells, the structure of these proton pumps is highly conserved from yeast to human, as a multiple subunit complex with a molecular mass exceeding 850 kDa. They contain at least 13 different subunits with various copy numbers, which are organized into two distinct domains, a peripheral V 1 domain that is the catalytic sector and a transmembrane V 0 domain that constitutes the proton channel.V-pumps are regulated at various levels from transcription and protein synthesis to the regulation of its enzymatic activity through a variety of mechanisms. The most unique regulation mechanism for V-pumps is the reversible dissociation and association of V 1 and V 0 in response to energy demand, which has been extensively studied and clearly demonstrated in yeast and tobacco hornworm (8 -10). However, whether this reversible dissociation and association of V-ATPase domains exists in mammals as a regulatory mechanism and, if it does, how this process is regulated, is not clear at the present.During the past two decades, the importance of this class of ATPases for many critical cellular functions has become increasingly appreciated. Furthermore, the elucidation of the physiological role of V-pumps has revealed the important role these proteins play in a wide array of pathological processes, such as osteoporosis (6), certain renal diseases (7, 11), HIV infection (12), and tumor metastasis (5). The food vacuole of certain parasites is acidified by V-type proton pumps, and disruption of the acidification of this intracellular compartment results in death of the organism. Thus, the V-type pumps are potential targets for the development of pharmacological agents to treat a variety of diseases.Because of the importance of V-ATPases as a potential therapeutic target, the mechanism by which inhibitors of V-ATPase interfere with pump function has become an area of great scientific interest. Over the past 15 years a few specific VATP...

Fast Synaptic Vesicle Reuse Slows the Rate of Synaptic Depression in the CA1 Region of Hippocampus

“…The rapid reduction in synaptic responses seen after folimycin treatment was only weakly reversible, even after extended wash out and stimulation (data not shown) (Xie et al, 2004). This shortcoming prompted us to explore an alternative reversible means to impair neurotransmitter refilling and assess the kinetics of synaptic vesicle reuse.…”

“…In the next set of experiments, we used another high affinity v-ATPase inhibitor, salicylihalamide A, to test the specificity of the folimycin effect (Xie et al, 2004). Treatment of hippocampal slices with salicylihalamide A (10 nM) resulted in a similar frequency-dependent augmentation of synaptic depression (Fig.…”

Inhibitors of V-ATPases: old and new players

“…In cell tests, all of these substances exhibited IC 50 values in the nanomolar range (Erickson et al, 1997;Huss et al, 2005;Kim et al, 1999;Kunze et al, 1998;McKee et al, 1998) and in the NCI 60-Cell screen, all of them revealed a V-ATPase inhibitor pattern similar to that of the plecomacrolides ) (Brigitte Kunze, personal communication). Assays of their inhibitory specificity for the VATPase carried out with either membrane preparations of human kidney, liver, osteoclast or bovine chromaffin granules Shen et al, 2002), or with purified V-ATPase from insect midgut (Huss et al, 2002;Huss et al, 2005) or bovine clathrin coated vesicles (Xie et al, 2004) clearly showed that the benzolactone enamides are indeed a novel family of highly specific V-ATPase inhibitors, with IC 50 values in the nanomolar range. One of the most astonishing and interesting features of the benzolacton enamides is that they do not inhibit V-ATPases from fungal sources, as was shown so far for salicylihalamide, lobatamides and oximidines with preparations of vacuolar membranes from N. crassa and Streptomyces cerevisiae , and for apicularen with preparations of vacuolar membranes from S. cerevisiae (S. Bockelmann, M.H., B. Kunze and H.W., unpublished results).…”

Genetic and pharmacologic manipulation of vacuolar ATPase: Effects on zymogen activation in pancreatic acini

“…4,10,11 This class of inhibitors includes the lobatomides and salicylihalamides, which were isolated from the marine tunicate worm 12 and marine sponge, 13 respectively and were originally identified as antitumor compounds. 10 Salicylihalamides bind to the V0 complex of the mammalian vATPase and this binding is distinct from the binding site for bafilomycin 14 and concanamycin. 4 In a previous study we showed that treatment of pancreatic acinar cells with bafilomycin or concanamycin blocked cerulein (CER)-induced zymogen activation.…”