Neuropsychological Performance in Alzheimer's Disease versus Late-Life Depression: A Systematic Review and Meta-Analysis.

OBJECTIVE: Despite decades of research, neuropsychological tests (NPTs) that clearly differentiate between Alzheimer's disease (AD) and late-life depression (LLD) have yet to be agreed upon. Given this gap in knowledge and the rapid deployment of disease-modifying drugs for the two disorders, accurate clinical diagnosis using evidence-based assessment is essential. This study aims to systematically examine the literature to identify NPTs that would be able to differentiate AD and LLD. METHOD: Databases and bibliographies were searched to identify articles for analysis. Two major inclusion criteria were that the studies compared neuropsychological functioning of AD versus LLD using normed NPTs and provided data for effect size calculation. Risk of bias was minimized by having independent coders for all steps in the review. RESULTS: Forty-one studies met inclusion criteria (N = 2,797) and provided effect sizes for tests that were classified as belonging to 15 domains of functioning. The two groups were well differentiated by tasks of delayed contextual verbal memory as compared to immediate or non-contextual memory, recognition cueing, confrontation naming, visuospatial construction, and conceptualization. Specific NPTs that appear to be useful for differential diagnosis include the Rey Auditory Verbal Learning Test-Delayed Recognition; Boston Naming Test; the Dementia Rating Scale's memory, conceptualization, and construction subscales; and the CERAD Constructional Praxis. CONCLUSIONS: The NPTs highlighted in this systematic review could be used as a relatively simple and cost-effective method to differentiate between patients with cognitive dysfunction due to AD versus LLD.

Neuropsychological Assessment and Screening in Heart Failure: a Meta-Analysis and Systematic Review.

A variety of neuropsychological changes secondary to heart failure have been documented in the literature. However, what remains unclear are which neuropsychological abilities are the most impacted by heart failure and what tests have the sensitivity to measure that impact. Eight databases were searched for articles that examined the neuropsychological functioning of patients with heart failure. Some of the inclusion criteria were articles had to have a heart failure group with a demographically comparable control group and standardized neuropsychological testing. Exclusion criteria included articles with a heart failure group with any other type of major organ failure, or comparisons that were between different classes of heart failure rather than between a heart failure and non-heart failure group. A total of 33 articles met the inclusion criteria (total heart failure sample n = 8900) and provided effect size data for 20 neuropsychological domains. All observed domain-level differences between heart failure and non-heart failure groups were statistically significant, except for simple motor functioning and confrontation naming. The greatest differences in performance were in executive functioning, global cognition, complex psychomotor speed, and verbal memory. The highest effect sizes came from Trail-Making Test-Part B, CAMCOG, Symbol Digit Modality Test, and California Verbal Learning Test. The neuropsychological patterns of heart failure suggested diffuse cognitive involvement, with higher-level processes being most affected. It is important to track neurocognition in this clinical population since neuropsychological impairment is prevalent, and screening measures appear to be reliable. Such screening and further assessment would inform future medical treatment and may improve patient care management.

Neuropsychological functioning in opioid use disorder: A research synthesis and meta-analysis.

BACKGROUND: Research has demonstrated that patients with opioid use disorders (OUD; including both opioid abuse and/or dependence) have poorer neuropsychological functioning compared to healthy controls; however, the pattern and robustness of the findings remain unknown. OBJECTIVES: This study meta-analyzed the results from previous research examining the neuropsychological deficits associated with opioids across 14 neurocognitive domains. METHOD: Articles comparing patients with OUD to healthy controls were selected based on detailed inclusion/exclusion criteria and variables of interest were coded. In total, 61 studies were selected for the analyses. These consisted of 2580 patients with OUD and 2102 healthy control participants (15.9% female). Drug-related variables were analyzed as potential moderators. RESULTS: The largest effect size difference in neuropsychological performance was observed in complex psychomotor ability. With the exception of the motor and processing speed domains, which showed no group differences, small-to-medium effect sizes were associated with all neurocognitive domains examined. Meta-regression revealed that increases in the length of abstinence were associated with decreases in effect sizes of the complex psychomotor domain. Additionally, attentional ability predicted effect size differences in executive functioning as well as verbal memory ability. Although the majority of meta-analyzed studies demonstrated significant differences between patients with OUD and controls, the average raw scores for patients with OUD in these studies typically fell within the normal range. CONCLUSION: The pattern of neuropsychological performance among patients with OUD appears to reflect mild generalized cognitive dysfunction, with a large effect in complex psychomotor abilities.

Two inhibitors of yeast plasma membrane ATPase 1 (ScPma1p): toward the development of novel antifungal therapies.

Given that many antifungal medications are susceptible to evolved resistance, there is a need for novel drugs with unique mechanisms of action. Inhibiting the essential proton pump Pma1p, a P-type ATPase, is a potentially effective therapeutic approach that is orthogonal to existing treatments. We identify NSC11668 and hitachimycin as structurally distinct antifungals that inhibit yeast ScPma1p. These compounds provide new opportunities for drug discovery aimed at this important target.

Neuropsychological comparisons of cocaine versus methamphetamine users: A research synthesis and meta-analysis.

BACKGROUND: Previous meta-analytical research examining cocaine and methamphetamine separately suggests potentially different neuropsychological profiles associated with each drug. In addition, neuroimaging studies point to distinct structural changes that might underlie differences in neuropsychological functioning. OBJECTIVES: This meta-analysis compared the effect sizes identified in cocaine versus methamphetamine studies across 15 neuropsychological domains. METHOD: Investigators searched and coded the literature examining the neuropsychological deficits associated with a history of either cocaine or methamphetamine use. A total of 54 cocaine and 41 methamphetamine studies were selected, yielding sample sizes of 1,718 and 1,297, respectively. Moderator analyses were conducted to compare the two drugs across each cognitive domain. RESULTS: Data revealed significant differences between the two drugs. Specifically, studies of cocaine showed significantly larger effect-size estimates (i.e., poorer performance) in verbal working memory when compared to methamphetamine. Further, when compared to cocaine, methamphetamine studies demonstrated significantly larger effect sizes in delayed contextual verbal memory and delayed visual memory. CONCLUSION: Overall, cocaine and methamphetamine users share similar neuropsychological profiles. However, cocaine appears to be more associated with working memory impairments, which are typically frontally mediated, while methamphetamine appears to be more associated with memory impairments that are linked with temporal and parietal lobe dysfunction.

Gray matter abnormalities in opioid-dependent patients: A neuroimaging meta-analysis.

BACKGROUND: Prior research utilizing whole-brain neuroimaging techniques has identified structural differences in gray matter in opioid-dependent individuals. However, the results have been inconsistent. OBJECTIVES: The current study meta-analytically examines the neuroimaging findings of studies published before 2016 comparing opioid-dependent individuals to drug-naïve controls. METHOD: Exhaustive search of five databases yielded 12 studies that met inclusion criteria. Anisotropic Effect-Size Seed-Based d Mapping (AES-SDM) was used to analyze the data extracted by three independent researchers. Voxel-based AES-SDM distinguishes increases and decreases in brain matter significant at the whole-brain level. RESULTS: AES-SDM identified the fronto-temporal region, bilaterally, as being the primary site of gray matter deficits associated with opioid use. Moderator analysis revealed that length of opioid use was negatively associated with gray matter in the left cerebellar vermis and the right Rolandic operculum, including the insula. Meta-regression revealed no remaining significant areas of gray matter reductions, except in the precuneus, following longer abstinence from opioids. CONCLUSIONS: Opioid-dependent individuals had significantly less gray matter in several regions that play a key role in cognitive and affective processing. The findings provide evidence that opioid dependence may result in the breakdown of two distinct yet highly overlapping structural and functional systems. These are the fronto-cerebellar system that might be more responsible for impulsivity, compulsive behaviors, and affective disturbances and the fronto-insular system that might account more for the cognitive and decision-making impairments.

Meta-analyses of clinical neuropsychological tests of executive dysfunction and impulsivity in alcohol use disorder.

BACKGROUND: Promising models for cognitive rehabilitation in alcohol treatment rest on a more nuanced understanding of the associated impairments in the multifaceted domains of executive functioning (EF) and impulsivity. OBJECTIVES: This meta-analysis examined the effects of alcohol on the individual subcomponents of EF and impulsivity in recently detoxified participants, including 1) Inhibition & Self-Regulation, 2) Flexibility & Set Shifting, 3) Planning & Problem Solving, 4) Reasoning & Abstraction, and 5) Verbal Fluency. Impulsivity was further examined through an analysis of motor, cognitive, and decisional subcategories. METHOD: Investigators searched, coded, and calculated effect sizes of impairments demonstrated in a broad range of neuropsychological tests for EF. A total of 77 studies were selected covering 48 years of research with a sample size of 5140. RESULTS: Findings ranged from a Hedges' g effect size of 0.803 for Inhibition to a Hedges' g of 0.359 for Verbal Fluency. Results also varied for the individual subcategories of Inhibition, including a large effect size for decisional impulsivity (g = 0.817) and cognitive impulsivity (0.860), and a moderate effect size for motor impulsivity (g = 0.529). The Hayling Test, Wisconsin Card Sorting Test, and Iowa Gambling Task were the measures most sensitive for alcohol effects. CONCLUSION: Planning, problem solving, and inhibitory abilities are significantly affected by alcohol abuse, with decisional and cognitive forms of impulsivity most impacted. Cognitive remediation targeting these deficits might increase the related functions that mediate the ability to moderate or abstain from alcohol, and so lead to improved treatment results.

Comparative chemical genomics reveal that the spiroindolone antimalarial KAE609 (Cipargamin) is a P-type ATPase inhibitor.

The spiroindolones, a new class of antimalarial medicines discovered in a cellular screen, are rendered less active by mutations in a parasite P-type ATPase, PfATP4. We show here that S. cerevisiae also acquires mutations in a gene encoding a P-type ATPase (ScPMA1) after exposure to spiroindolones and that these mutations are sufficient for resistance. KAE609 resistance mutations in ScPMA1 do not confer resistance to unrelated antimicrobials, but do confer cross sensitivity to the alkyl-lysophospholipid edelfosine, which is known to displace ScPma1p from the plasma membrane. Using an in vitro cell-free assay, we demonstrate that KAE609 directly inhibits ScPma1p ATPase activity. KAE609 also increases cytoplasmic hydrogen ion concentrations in yeast cells. Computer docking into a ScPma1p homology model identifies a binding mode that supports genetic resistance determinants and in vitro experimental structure-activity relationships in both P. falciparum and S. cerevisiae. This model also suggests a shared binding site with the dihydroisoquinolones antimalarials. Our data support a model in which KAE609 exerts its antimalarial activity by directly interfering with P-type ATPase activity.

Yeast Fex1p Is a Constitutively Expressed Fluoride Channel with Functional Asymmetry of Its Two Homologous Domains.

Fluoride is a ubiquitous environmental toxin with which all biological species must cope. A recently discovered family of fluoride export (FEX) proteins protects organisms from fluoride toxicity by removing it from the cell. We show here that FEX proteins in Saccharomyces cerevisiae function as ion channels that are selective for fluoride over chloride and that these proteins are constitutively expressed at the yeast plasma membrane. Continuous expression is in contrast to many other toxin exporters in yeast, and this, along with the fact that two nearly duplicate proteins are encoded in the yeast genome, suggests that the threat posed by fluoride ions is frequent and detrimental. Structurally, eukaryotic FEX proteins consist of two homologous four-transmembrane helix domains folded into an antiparallel dimer, where the orientation of the two domains is fixed by a single transmembrane linker helix. Using phylogenetic sequence conservation as a guide, we have identified several functionally important residues. There is substantial functional asymmetry in the effect of mutation at corresponding sites in the two domains. Specifically, mutations to residues in the C-terminal domain proved significantly more detrimental to function than did similar mutations in the N-terminal domain. Our data suggest particular residues that may be important to anion specificity, most notably the necessity of a positive charge near the end of TMH1 in the C-terminal domain. It is possible that a cationic charge at this location may create an electrostatic well for fluoride ions entering the channel from the cytoplasm.

C-terminal truncations of the Saccharomyces cerevisiae PMA1 H+-ATPase have major impacts on protein conformation, trafficking, quality control, and function.

The C-terminal tail of yeast plasma membrane (PM) H(+)-ATPase extends approximately 38 amino acids beyond the final membrane-spanning segment (TM10) of the protein and is known to be required for successful trafficking, stability, and regulation of enzyme activity. To carry out a detailed functional survey of the entire length of the tail, we generated 15 stepwise truncation mutants. Eleven of them, lacking up to 30 amino acids from the extreme terminus, were able to support cell growth, even though there were detectable changes in plasma membrane expression, protein stability, and ATPase activity. Three functionally distinct regions of the C terminus could be defined. (i) Truncations upstream of Lys(889), removing more than 30 amino acid residues, yielded no viable mutants, and conditional expression of such constructs supported the conclusion that the stretch from Ala(881) (at the end of TM10) to Gly(888) is required for stable folding and PM targeting. (ii) The stretch between Lys(889) and Lys(916), a region known to be subject to kinase-mediated posttranslational modification, was shown here to be ubiquitinated in carbon-starved cells as part of cellular quality control and to be essential for normal ATPase folding and stability, as well as for autoinhibition of ATPase activity during glucose starvation. (iii) Finally, removal of even one or two residues (Glu(917) and Thr(918)) from the extreme C terminus led to visibly reduced expression of the ATPase at the plasma membrane. Thus, the C terminus is much more than a simple appendage and profoundly influences the structure, biogenesis, and function of the yeast H(+)-ATPase.

Coordination of K+ transporters in neurospora: TRK1 is scarce and constitutive, while HAK1 is abundant and highly regulated.

Fungi, plants, and bacteria accumulate potassium via two distinct molecular machines not directly coupled to ATP hydrolysis. The first, designated TRK, HKT, or KTR, has eight transmembrane helices and is folded like known potassium channels, while the second, designated HAK, KT, or KUP, has 12 transmembrane helices and resembles MFS class proteins. One of each type functions in the model organism Neurospora crassa, where both are readily accessible for biochemical, genetic, and electrophysiological characterization. We have now determined the operating balance between Trk1p and Hak1p under several important conditions, including potassium limitation and carbon starvation. Growth measurements, epitope tagging, and quantitative Western blotting have shown the gene HAK1 to be much more highly regulated than is TRK1. This conclusion follows from three experimental results: (i) Trk1p is expressed constitutively but at low levels, and it is barely sensitive to extracellular [K(+)] and/or the coexpression of HAK1; (ii) Hak1p is abundant but is markedly depressed by elevated extracellular concentrations of K(+) and by coexpression of TRK1; and (iii) Carbon starvation slowly enhances Hak1p expression and depresses Trk1p expression, yielding steady-state Hak1p:Trk1p ratios of ∼500:1, viz., 10- to 50-fold larger than that in K(+)- and carbon-replete cells. Additionally, it appears that both potassium transporters can adjust kinetically to sustained low-K(+) stress by means of progressively increasing transporter affinity for extracellular K(+). The underlying observations are (iv) that K(+) influx via Trk1p remains nearly constant at ∼9 mM/h when extracellular K(+) is progressively depleted below 0.05 mM and (v) that K(+) influx via Hak1p remains at ∼3 mM/h when extracellular K(+) is depleted below 0.1 mM.

Tandem phosphorylation of Ser-911 and Thr-912 at the C terminus of yeast plasma membrane H+-ATPase leads to glucose-dependent activation.

In recent years there has been growing interest in the post-translational regulation of P-type ATPases by protein kinase-mediated phosphorylation. Pma1 H(+)-ATPase, which is responsible for H(+)-dependent nutrient uptake in yeast (Saccharomyces cerevisiae), is one such example, displaying a rapid 5-10-fold increase in activity when carbon-starved cells are exposed to glucose. Activation has been linked to Ser/Thr phosphorylation in the C-terminal tail of the ATPase, but the specific phosphorylation sites have not previously been mapped. The present study has used nanoflow high pressure liquid chromatography coupled with electrospray electron transfer dissociation tandem mass spectrometry to identify Ser-911 and Thr-912 as two major phosphorylation sites that are clearly related to glucose activation. In carbon-starved cells with low Pma1 activity, peptide 896-918, which was derived from the C terminus upon Lys-C proteolysis, was found to be singly phosphorylated at Thr-912, whereas in glucose-metabolizing cells with high ATPase activity, the same peptide was doubly phosphorylated at Ser-911 and Thr-912. Reciprocal (14)N/(15)N metabolic labeling of cells was used to measure the relative phosphorylation levels at the two sites. The addition of glucose to carbon-starved cells led to a 3-fold reduction in the singly phosphorylated form and an 11-fold increase in the doubly phosphorylated form. These results point to a mechanism in which the stepwise phosphorylation of two tandemly positioned residues near the C terminus mediates glucose-dependent activation of the H(+)-ATPase.

Role of transmembrane segment M8 in the biogenesis and function of yeast plasma-membrane H(+)-ATPase.

Of the four transmembrane helices (M4, M5, M6, and M8) that pack together to form the ion-binding sites of P(2)-type ATPases, M8 has until now received the least attention. The present study has used alanine-scanning mutagenesis to map structure-function relationships throughout M8 of the yeast plasma-membrane H(+)-ATPase. Mutant forms of the ATPase were expressed in secretory vesicles and at the plasma membrane for measurements of ATP hydrolysis and ATP-dependent H(+) pumping. In secretory vesicles, Ala substitutions at a cluster of four positions near the extracytoplasmic end of M8 led to partial uncoupling of H(+) transport from ATP hydrolysis, while substitution of Ser-800 (close to the middle of M8) by Ala increased the apparent stoichiometry of H(+) transport. A similar increase has previously been reported following the substitution of Glu-803 by Gln (Petrov, V. et al., J. Biol. Chem. 275:15709-15718, 2000) at a position known to contribute directly to Ca(2+) binding in the Ca(2+)-ATPase of sarcoplasmic reticulum (Toyoshima, C., et al., Nature 405: 647-655, 2000). Four other mutations in M8 interfered with H(+)-ATPase folding and trafficking to the plasma membrane; based on homology modeling, they occupy positions that appear important for the proper bundling of M8 with M5, M6, M7, and M10. Taken together, these results point to a key role for M8 in the biogenesis, stability, and physiological functioning of the H(+)-ATPase.

Effects of C-terminal truncations on trafficking of the yeast plasma membrane H+-ATPase.

Within the large family of P-type cation-transporting ATPases, members differ in the number of C-terminal transmembrane helices, ranging from two in Cu2+-ATPases to six in H+-, Na+,K+-, Mg2+-, and Ca2+-ATPases. In this study, yeast Pma1 H+-ATPase has served as a model to examine the role of the C-terminal membrane domain in ATPase stability and targeting to the plasma membrane. Successive truncations were constructed from the middle of the major cytoplasmic loop to the middle of the extended cytoplasmic tail, adding back the C-terminal membrane-spanning helices one at a time. When the resulting constructs were expressed transiently in yeast, there was a steady increase in half-life from 70 min in Pma1 delta452 to 348 min in Pma1 delta901, but even the longest construct was considerably less stable than wild-type ATPase (t(1/2) = 11 h). Confocal immunofluorescence microscopy showed that 11 of 12 constructs were arrested in the endoplasmic reticulum and degraded in the proteasome. The only truncated ATPase that escaped the ER, Pma1 delta901, traveled slowly to the plasma membrane, where it hydrolyzed ATP and supported growth. Limited trypsinolysis showed Pma1 delta901 to be misfolded, however, resulting in premature delivery to the vacuole for degradation. As model substrates, this series of truncations affirms the importance of the entire C-terminal domain to yeast H+-ATPase biogenesis and defines a sequence element of 20 amino acids in the carboxyl tail that is critical to ER escape and trafficking to the plasma membrane.

Conformational changes of yeast plasma membrane H(+)-ATPase during activation by glucose: role of threonine-912 in the carboxy-terminal tail.

Yeast Pma1 H(+)-ATPase, which belongs to the P-type family of cation-transporting ATPases, is activated up to 10-fold by growth on glucose, and indirect evidence has linked the activation to Ser/Thr phosphorylation within the C-terminal tail. We have now used limited trypsinolysis to map glucose-induced conformational changes throughout the 100 kDa ATPase. In the wild-type enzyme, trypsin cleaves first at Lys-28 and Arg-73 in the extended N-terminal segment (sites T1 and T2); subsequent cleavages occur at Arg-271 between the A domain and M3 (site T3) and at Lys-749 or Lys-754 in the M6-M7 cytoplasmic loop (site T4). Activation by glucose leads to a striking increase in trypsin sensitivity. At the C-terminal end of the protein, the Arg- and Lys-rich tail is shielded from trypsin in membranes from glucose-starved cells (GS) but becomes accessible in membranes from glucose-metabolizing cells (GM). In the presence of orthovanadate, Lys-174 at the boundary between M2 and the A domain also becomes open to cleavage in GM but not GS samples (site T5). Significantly, this global conformational change can be suppressed by mutations at Thr-912, a consensus phosphorylation site near the C-terminus. Substitution by Ala at position 912 leads to a GS-like (trypsin-resistant) state, while substitution by Asp leads to a GM-like (trypsin-sensitive) state. Thus, the present results help to dissect the intramolecular movements that result in glucose activation.

Use of a fluorescent maleimide to probe structure-function relationships in stalk segments 4 and 5 of the yeast plasma-membrane H+-ATPase.

In the yeast plasma-membrane H(+)-ATPase and other P-type ATPases, conformational changes are transmitted between cytoplasmic and membrane-embedded domains via a stalk region composed of cytoplasmic extensions of membrane segments 2, 3, 4, and 5. The present study has used a fluorescent maleimide (Alexa-488) to probe Cys residues introduced into stalk segments 4 and 5 of the yeast enzyme. In the case of S5, Cys substitutions along one face led to a constitutive, 5- to 10-fold activation of the ATPase in the absence of glucose. Based on homology with SERCA Ca(2+)-ATPase, this face is likely to be buried in the interior of the protein, close to the P domain. Three Cys residues on the opposite face of S5 (A668C, S672C, and D676C) were accessible to Alexa-488 under all conditions tested. In addition, three other Cys residues at or near the boundary between the two faces reacted with Alexa-488 only (V665C, L678C) or preferentially (Y689C) in plasma membranes from glucose-metabolizing cells; this result provides the first direct evidence for a change in conformation of S5 during glucose activation. For stalk segment 4, site-directed mutagenesis gave no sign of a role in glucose-dependent regulation. Rather, substitutions at 13 consecutive positions along S4 caused kinetic changes consistent with a shift in equilibrium from E2 to E1. Four Cys residues along this stretch of S4 (Q357C, K362C, S364C, and S368C) reacted with Alexa-488, indicating that they are exposed to the aqueous medium as predicted in the SERCA-based structural model.

Stalk segment 5 of the yeast plasma membrane H(+)-ATPase. Labeling with a fluorescent maleimide reveals a conformational change during glucose activation.

Glucose is well known to cause a rapid, reversible activation of the yeast plasma membrane H(+)-ATPase, very likely mediated by phosphorylation of two or more Ser/Thr residues near the C terminus. Recent mutagenesis studies have shown that glucose-dependent activation can be mimicked constitutively by amino acid substitutions in stalk segment 5 (S5), an alpha-helical stretch connecting the catalytic part of the ATPase with transmembrane segment 5 (Miranda, M., Allen, K. E., Pardo, J. P., and Slayman, C. W. (2001) J. Biol. Chem. 276, 22485-22490). In the present work, the fluorescent maleimide Alexa-488 has served as a probe for glucose-dependent changes in the conformation of S5. Experiments were carried out in a "3C" version of the ATPase, from which six of nine native cysteines had been removed by site-directed mutagenesis to eliminate background labeling by Alexa-488. In this construct, three of twelve cysteines introduced at various positions along S5 (A668C, S672C, and D676C) reacted with the Alexa dye in a glucose-independent manner, as shown by fluorescent labeling of the 100 kDa Pma1 polypeptide and by isolation and identification of the corresponding tryptic peptides. Especially significant was the fact that three additional cysteines reacted with Alexa-488 more rapidly (Y689C) or only (V665C and L678C) in plasma membranes from glucose-metabolizing cells. The results support a model in which the S5 alpha-helix undergoes a significant change in conformation to expose positions 665, 678, and 689 during glucose-dependent activation of the ATPase.

Quality control in the yeast secretory pathway: a misfolded PMA1 H+-ATPase reveals two checkpoints.

The yeast plasma-membrane H(+)-ATPase, encoded by PMA1, is delivered to the cell surface via the secretory pathway and has recently emerged as an excellent system for identifying quality control mechanisms along the pathway. In the present study, we have tracked the biogenesis of Pma1-G381A, a misfolded mutant form of the H(+)-ATPase. Although this mutant ATPase is arrested transiently in the peripheral endoplasmic reticulum, it does not become a substrate for endoplasmic reticulum-associated degradation nor does it appear to stimulate an unfolded protein response. Instead, Pma1-G381A accumulates in Kar2p-containing vesicular-tubular clusters that resemble those previously described in mammalian cells. Like their mammalian counterparts, the yeast vesicular-tubular clusters may correspond to specific exit ports from the endoplasmic reticulum, since Pma1-G381A eventually escapes from them (still in a misfolded, trypsin-sensitive form) to reach the plasma membrane. By comparison with wild-type ATPase, Pma1-G381A spends a short half-life at the plasma membrane before being removed and sent to the vacuole for degradation in a process that requires both End4p and Pep4p. Finally, in a separate set of experiments, Pma1-G381A was found to impose its phenotype on co-expressed wild-type ATPase, transiently retarding the wild-type protein in the ER and later stimulating its degradation in the vacuole. Both effects serve to lower the steady-state amount of wild-type ATPase in the plasma membrane and, thus, can explain the co-dominant genetic behavior of the G381A mutation. Taken together, the results of this study establish Pma1-G381A as a useful new probe for the yeast secretory system.