Control of vacuole membrane homeostasis by a resident PI-3,5-kinase inhibitor

Lysosomes have an important role in cellular protein and organelle quality control, metabolism, and signaling. On the surface of lysosomes, the PIKfyve/Fab1 complex generates phosphatidylinositol 3,5-bisphosphate, PI-3,5-P2, which is critical for lysosomal membrane homeostasis during acute osmotic stress and for lysosomal signaling. Here, we identify the inverted BAR protein Ivy1 as an inhibitor of the Fab1 complex with a direct influence on PI-3,5- P2 levels and vacuole homeostasis. Ivy1 requires Ypt7 binding for its function, binds PI-3,5-P2, and interacts with the Fab1 kinase. Colocalization of Ivy1 and Fab1 is lost during osmotic stress. In agreement with Ivy1’s role as a Fab1 regulator, its overexpression blocks Fab1 activity during osmotic shock and vacuole fragmentation. Conversely, loss of Ivy1, or lateral relocalization of Ivy1 on vacuoles away from Fab1, results in vacuole fragmentation and poor growth. Our data suggest that Ivy1 modulates Fab1- mediated PI-3,5-P2 synthesis during membrane stress and may allow adjustment of the vacuole membrane environment

Endoplasmic reticulum–plasma membrane contacts: Principals of phosphoinositide and calcium signaling

The endoplasmic reticulum (ER) forms an extensive network of membrane contact sites with intra-cellular organelles and the plasma membrane (PM). Interorganelle contacts have vital roles in membrane lipid and ion dynamics. In particular, ER–PM contacts are integral to numerous inter-cellular and intra-cellular signaling pathways including phosphoinositide lipid and calcium signaling, mechanotransduction, metabolic regulation, and cell stress responses. Accordingly, ER–PM contacts serve important signaling functions in excitable cells including neurons and muscle and endocrine cells. This review highlights recent advances in our understanding of the vital roles for ER–PM contacts in phosphoinositide and calcium signaling and how signaling pathways in turn regulate proteins that form and function at ER–PM contacts

Building ER-PM contacts: keeping calm and ready on alarm

It is well over half a century since contacts between organelles such as the endoplasmic reticulum (ER), mitochondria, and the plasma membrane (PM) were first observed in electron microscopy studies. Still, these early images of seemingly rare organelle interactions continue to capture the attention and curiosity of cell biologists even today. From seminal studies first proposing roles for organelle cross talk in excitable cells, the field has now expanded to cover nearly all aspects of eukaryotic cell biology, from calcium and membrane lipid transport to vesicular trafficking, cell signaling, metabolism, and homeostasis. This review highlights recent discoveries pointing to vital roles for ER-PM contacts in membrane lipid dynamics and organization

Phosphoinositide kinase signaling controls ER-PM cross-talk

Membrane lipid dynamics must be precisely regulated for normal cellular function, and disruptions in lipid homeostasis are linked to the progression of several diseases. However, little is known about the sensory mechanisms for detecting membrane composition and how lipid metabolism is regulated in response to membrane stress. We find that phosphoinositide (PI) kinase signaling controls a conserved PDK-TORC2-Akt signaling cascade as part of a homeostasis network that allows the endoplasmic reticulum (ER) to modulate essential responses, including Ca(2+)-regulated lipid biogenesis, upon plasma membrane (PM) stress. Furthermore, loss of ER-PM junctions impairs this protective response, leading to PM integrity defects upon heat stress. Thus PI kinase-mediated ER-PM cross-talk comprises a regulatory system that ensures cellular integrity under membrane stress conditions

Specialized ER membrane domains for lipid metabolism and transport

The endoplasmic reticulum (ER) is a highly organized organelle that performs vital functions including de novo membrane lipid synthesis and transport. Accordingly, numerous lipid biosynthesis enzymes are localized in the ER membrane. However, it is now evident that lipid metabolism is sub-compartmentalized within the ER and that lipid biosynthetic enzymes engage with lipid transfer proteins (LTPs) to rapidly shuttle newly synthesized lipids from the ER to other organelles. As such, intimate relationships between lipid metabolism and lipid transfer pathways exist within the ER network. Notably, certain LTPs enhance the activities of lipid metabolizing enzymes; likewise, lipid metabolism can ensure the specificity of LTP transfer/exchange reactions. Yet, our understanding of these mutual relationships is still emerging. Here, we highlight past and recent key findings on specialized ER membrane domains involved in efficient lipid metabolism and transport and consider unresolved issues in the field

Diagnostic Utility of the Impact of Event Scale-Revised in Two Samples of Survivors of War

The study aimed at examining the diagnostic utility of the Impact of Event Scale-Revised (IES-R) as a screening tool for post-traumatic stress disorder (PTSD) in survivors of war. The IES-R was completed by two independent samples that had survived the war in the Balkans: a sample of randomly selected people who had stayed in the area of former conflict (n = 3,313) and a sample of refugees to Western European countries (n = 854). PTSD was diagnosed using the MINI International Neuropsychiatric Interview. Prevalence of PTSD was 20.1% in the Balkan sample and 33.1% in the refugee sample. Results revealed that when considering a minimum value of specificity of 0.80, the optimally sensitive cut-off score for screening for PTSD in the Balkan sample was 34. In both the Balkan sample and the refugee sample, this cut-off score provided good values on sensitivity (0.86 and 0.89, respectively) and overall efficiency (0.81 and 0.79, respectively). Further, the kappa coefficients for sensitivity for the cut-off of 34 were 0.80 in both samples. Findings of this study support the clinical utility of the IES-R as a screening tool for PTSD in large-scale research studies and intervention studies if structured diagnostic interviews are regarded as too labor-intensive and too costly

TORC1 Determines Fab1 Lipid Kinase Function at Signaling Endosomes and Vacuoles

Organelles of the endomembrane system maintain their identity and integrity during growth or stress conditions by homeostatic mechanisms that regulate membrane flux and biogenesis. At lysosomes and endosomes, the Fab1 lipid kinase complex and the nutrient-regulated target of rapamycin complex 1 (TORC1) control the integrity of the endolysosomal homeostasis and cellular metabolism. Both complexes are functionally connected as Fab1-dependent generation of PI(3,5)P2 supports TORC1 activity. Here, we identify Fab1 as a target of TORC1 on signaling endosomes, which are distinct from multivesicular bodies, and provide mechanistic insight into their crosstalk. Accordingly, TORC1 can phosphorylate Fab1 proximal to its PI3P-interacting FYVE domain, which causes Fab1 to shift to signaling endosomes, where it generates PI(3,5)P2. This, in turn, regulates (1) vacuole morphology, (2) recruitment of TORC1 and the TORC1-regulatory Rag GTPase-containing EGO complex to signaling endosomes, and (3) TORC1 activity. Thus, our study unravels a regulatory feedback loop between TORC1 and the Fab1 complex that controls signaling at endolysosomes

Rapid Diagnostic Algorithms as a Screening Tool for Tuberculosis: An Assessor Blinded Cross-Sectional Study

Background: A major obstacle to effectively treat and control tuberculosis is the absence of an accurate, rapid, and low-cost diagnostic tool. A new approach for the screening of patients for tuberculosis is the use of rapid diagnostic classification algorithms. Methods: We tested a previously published diagnostic algorithm based on four biomarkers as a screening tool for tuberculosis in a Central European patient population using an assessor-blinded cross-sectional study design. In addition, we developed an improved diagnostic classification algorithm based on a study population at a tertiary hospital in Vienna, Austria, by supervised computational statistics. Results: The diagnostic accuracy of the previously published diagnostic algorithm for our patient population consisting of 206 patients was 54% (CI: 47%–61%). An improved model was constructed using inflammation parameters and clinical information. A diagnostic accuracy of 86% (CI: 80%–90%) was demonstrated by 10-fold cross validation. An alternative model relying solely on clinical parameters exhibited a diagnostic accuracy of 85% (CI: 79%–89%). Conclusion: Here we show that a rapid diagnostic algorithm based on clinical parameters is only slightly improved by inclusion of inflammation markers in our cohort. Our results also emphasize the need for validation of new diagnostic algorithms in different settings and patient populations

52-week efficacy and safety of telbivudine with conditional tenofovir intensification at week 24 in HBeAg-positive chronic Hepatitis B

Background and Aims: The Roadmap concept is a therapeutic framework in chronic hepatitis B for the intensification of nucleoside analogue monotherapy based on early virologic response. The efficacy and safety of this approach applied to telbivudine treatment has not been investigated. Methods: A multinational, phase IV, single-arm open-label study (ClinicalTrials.gov ID NCT00651209) was undertaken in HBeAg-positive, nucleoside-naive adult patients with chronic hepatitis B. Patients received telbivudine (600 mg once-daily) for 24 weeks, after which those with undetectable serum HBV DNA (<300 copies/mL) continued to receive telbivudine alone while those with detectable DNA received telbivudine plus tenofovir (300 mg once-daily). Outcomes were assessed at Week 52. Results: 105 patients commenced telbivudine monotherapy, of whom 100 were included in the efficacy analysis. Fifty-five (55%) had undetectable HBV DNA at Week 24 and continued telbivudine monotherapy; 45 (45%) received tenofovir intensification. At Week 52, the overall proportion of undetectable HBV DNA was 93% (93/100) by last-observation-carried-forward analysis (100% monotherapy group, 84% intensification group) and no virologic breakthroughs had occurred. ALT normalization occurred in 77% (87% monotherapy, 64% intensification), HBeAg clearance in 43% (65% monotherapy, 16% intensification), and HBeAg seroconversion in 39% (62% monotherapy, 11% intensification). Six patients had HBsAg clearance. Myalgia was more common in the monotherapy group (19% versus 7%). No decrease in the mean glomerular filtration rate occurred in either treatment group at Week 52. Conclusions: Telbivudine therapy with tenofovir intensification at Week 24, where indicated by the Roadmap strategy, appears effective and well tolerated for the treatment of chronic hepatitis B. Trial Registration: ClinicalTrials.gov NCT0065120