Brett, Christopher L., Mark Donowitz, and Rajini Rao. Evolutionary origins of eukaryotic sodium/proton exchangers. Am J Physiol Cell Physiol 288: C223-C239, 2005; doi:10.1152/ajpcell.00360.2004.-More than 200 genes annotated as Na ϩ /H ϩ hydrogen exchangers (NHEs) currently reside in bioinformation databases such as GenBank and Pfam. We performed detailed phylogenetic analyses of these NHEs in an effort to better understand their specific functions and physiological roles. This analysis initially required examining the entire monovalent cation proton antiporter (CPA) superfamily that includes the CPA1, CPA2, and NaT-DC families of transporters, each of which has a unique set of bacterial ancestors. We have concluded that there are nine human NHE (or SLC9A) paralogs as well as two previously unknown human CPA2 genes, which we have named HsNHA1 and HsNHA2. The eukaryotic NHE family is composed of five phylogenetically distinct clades that differ in subcellular location, drug sensitivity, cation selectivity, and sequence length. The major subgroups are plasma membrane (recycling and resident) and intracellular (endosomal/TGN, NHE8-like, and plant vacuolar). HsNHE1, the first cloned eukaryotic NHE gene, belongs to the resident plasma membrane clade. The latter is the most recent to emerge, being found exclusively in vertebrates. In contrast, the intracellular clades are ubiquitously distributed and are likely precursors to the plasma membrane NHE. Yeast endosomal ScNHX1 was the first intracellular NHE to be described and is closely related to HsNHE6, HsNHE7, and HsNHE9 in humans. Our results link the appearance of NHE on the plasma membrane of animal cells to the use of the Na ϩ /K ϩ -ATPase to generate the membrane potential. These novel observations have allowed us to use comparative biology to predict physiological roles for the nine human NHE paralogs and to propose appropriate model organisms in which to study the unique properties of each NHE subclass. Naϩ /H ϩ exchanger; NHX; cation proton antiporter; phylogenetic analysis A BASIC PROPERTY OF LIFE is the ability of an organism to regulate cellular pH, volume, and ion composition. The transmembrane exchange of protons for sodium ions (Na ϩ ) is ubiquitous in organisms across all phyla and kingdoms, and underlies fundamental homeostatic mechanisms to control these ions. The family of Na ϩ /H ϩ exchangers (NHEs) plays an important role in diverse physiological processes, including control of cell cycle and cell proliferation (114, 117), transepithelial Na ϩ movement (174), salt tolerance (93, 130), vesicle trafficking, and biogenesis (5, 22). In mammals, NHE dysfunction is associated with pathophysiological conditions that include hypertension, epilepsy, postischemic myocardial arrhythmia, gastric and kidney disease, diarrhea, and glaucoma (36, 106, 174). Drugs such as S8218 and cariporide, which target specific NHE isoforms, are used to reduce the duration of apnea in animal studies and in clinical trials for the prevention of cardiac ischemia-reperfusion injur...

The relationship between endosomal pH and function is well documented in viral entry, endosomal maturation, receptor recycling, and vesicle targeting within the endocytic pathway. However, specific molecular mechanisms that either sense or regulate luminal pH to mediate these processes have not been identified. Herein we describe the use of novel, compartment-specific pH indicators to demonstrate that yeast Nhx1, an endosomal member of the ubiquitous NHE family of Na ؉ /H ؉ exchangers, regulates luminal and cytoplasmic pH to control vesicle trafficking out of the endosome. Loss of Nhx1 confers growth sensitivity to low pH stress, and concomitant acidification and trafficking defects, which can be alleviated by weak bases. Conversely, weak acids cause wild-type yeast to present nhx1⌬ trafficking phenotypes. Finally, we report that Nhx1 transports K ؉ in addition to Na ؉ , suggesting that a single mechanism may responsible for both pH and K ؉ -dependent endosomal processes. This presents the newly defined family of eukaryotic endosomal NHE as novel targets for pharmacological inhibition to alleviate pathological states associated with organellar alkalinization. INTRODUCTIONIt is well established that luminal acidification of the endocytic pathway, including the endosome and lysosome/vacuole, is required for associated cellular function (Mellman et al., 1986;Mellman, 1992). Some examples include ligandreceptor dissociation and recycling of surface receptors, lysosome-mediated protein degradation, H ϩ -driven neurotransmitter loading and pH-dependent recycling of synaptic vesicles (Buckley et al., 2000;Nishi and Forgac, 2002). Similarly, viral pathogen entry and propagation is dependent on the pH gradient across the lumen of the endosome (Harley et al., 2001), and the abnormal lysosomal/endosomal morphologies and associated defective trafficking observed in a subset of lysosomal storage disorders are associated with abnormal changes in luminal pH (Futerman and van Meer, 2004). Pioneering experiments performed by Heuser clearly demonstrated that changes in cellular pH alone severely alter organellar morphology and movement (Heuser, 1989). This phenomenon can be explained by net changes in vesicle trafficking between compartments, as luminal pH can direct vesicle trafficking; thus, elevated pH in the endosome promotes endosome to Golgi vesicle movement (van Weert et al., 1995(van Weert et al., , 1997 also see Nieland et al., 2004). At the molecular level, local increases in pH are believed to be responsible for assembly of vesicle trafficking/sorting machinery in areas of the endosome destined for return to the plasma membrane (Maranda et al., 2001; also see Zeuzem et al., 1992;Aniento et al., 1996). Despite extensive evidence that changes in pH direct trafficking in this pathway, specific molecular mechanisms that control pH itself have not been defined. The ubiquitous Na ϩ /H ϩ exchangers of the NHE family are associated with cellular pH regulation (Orlowski and Grinstein, 2004). Recent phylogenetic analysis of the ...

Genetic studies in yeast, plants, insects and mammals have identified four universally conserved proteins, together called Vps Class C, that are essential for late endosome and lysosome assembly and for numerous endolysosomal trafficking pathways, including the terminal stages of autophagy. Two Vps-C complexes, HOPS and CORVET, incorporate diverse biochemical functions: they tether membranes, stimulate Rab nucleotide exchange, guide SNARE assembly to drive membrane fusion, and possibly act as ubiquitin ligases. Recent studies offer new insight into the complex relationships between Vps-C complexes and their cognate Rab small GTP-binding (G-)proteins at endosomes and lysosomes. Accumulating evidence supports the view that Vps-C complexes implement a regulatory logic that governs endomembrane identity and dynamics.