Evolution of an ancient protein function involved in organized multicellularity in animals.

To form and maintain organized tissues, multicellular organisms orient their mitotic spindles relative to neighboring cells. A molecular complex scaffolded by the GK protein-interaction domain (GKPID) mediates spindle orientation in diverse animal taxa by linking microtubule motor proteins to a marker protein on the cell cortex localized by external cues. Here we illuminate how this complex evolved and commandeered control of spindle orientation from a more ancient mechanism. The complex was assembled through a series of molecular exploitation events, one of which - the evolution of GKPID's capacity to bind the cortical marker protein - can be recapitulated by reintroducing a single historical substitution into the reconstructed ancestral GKPID. This change revealed and repurposed an ancient molecular surface that previously had a radically different function. We show how the physical simplicity of this binding interface enabled the evolution of a new protein function now essential to the biological complexity of many animals

Plasma exosome profile in st-elevation myocardial infarction patients with and without out-of-hospital cardiac arrest

The identification of new biomarkers allowing an early and more accurate characterization of patients with ST-segment elevation myocardial infarction (STEMI) is still needed, and exosomes represent an attractive diagnostic tool in this context. However, the characterization of their protein cargo in relation to cardiovascular clinical manifestation is still lacking. To this end, 35 STEMI patients (17 experiencing resuscitated out-of-hospital cardiac arrest (OHCA-STEMI) and 18 uncomplicated) and 32 patients with chronic coronary syndrome (CCS) were enrolled. Plasma exosomes were characterized by the nanoparticle tracking analysis and Western blotting. Exosomes from STEMI patients displayed a higher concentration and size and a greater expression of platelet (GPIIb) and vascular endothelial (VE-cadherin) markers, but a similar amount of cardiac troponin compared to CCS. In addition, a difference in exosome expression of acute-phase proteins (ceruloplasmin, transthyretin and fibronectin) between STEMI and CCS patients was found. GPIIb and brain-associated marker PLP1 accurately discriminated between OHCA and uncomplicated STEMI. In conclusion, the exosome profile of STEMI patients has peculiar features that differentiate it from that of CCS patients, reflecting the pathophysiological mechanisms involved in STEMI. Additionally, the exosome expression of brain-and platelet-specific markers might allow the identification of patients experiencing ischemic brain injury in STEMI

Cardiac magnetic resonance predictors of left ventricular remodelling following acute ST elevation myocardial infarction: The VavirimS study

Left ventricular (LV) remodelling (REM) ensuing after ST-elevation myocardial infarction (STEMI), has typically been studied by echocardiography, which has limitations, or cardiac magnetic resonance (CMR) in early phase that may overestimate infarct size (IS) due to tissue edema and stunning. This prospective, multicenter study investigated LV-REM performing CMR in the subacute phase, and 6 months after STEMI

The Legionella effector WipB is a translocated Ser/Thr phosphatase that targets the host lysosomal nutrient sensing machinery

Legionella pneumophila infects human alveolar macrophages and is responsible for Legionnaire’s disease, a severe form of pneumonia. L. pneumophila encodes more than 300 putative effectors, which are translocated into the host cell via the Dot/Icm type IV secretion system. These effectors highjack the host’s cellular processes to allow bacterial intracellular growth and replication. Here we adopted a multidisciplinary approach to investigate WipB, a Dot/Icm effector of unknown function. The crystal structure of the N-terminal domain at 1.7 Å resolution comprising residues 25 to 344 revealed that WipB harbours a Ser/Thr phosphatase domain related to the eukaryotic phospho-protein phosphatase (PPP) family. The C-terminal domain (residues 365–524) is sufficient to pilot the effector to acidified LAMP1-positive lysosomal compartments, where WipB interacts with the v-ATPase and the associated LAMTOR1 phosphoprotein, key components of the lysosomal nutrient sensing (LYNUS) apparatus that controls the mammalian target of rapamycin (mTORC1) kinase complex at the lysosomal surface. We propose that WipB is a lysosome-targeted phosphatase that modulates cellular nutrient sensing and the control of energy metabolism during Legionella infection

Activity of endovesical gemcitabine in BCG-refractory bladder cancer patients: a translational study

Intravesical gemcitabine (Gem) has shown promising activity against transitional cell carcinomas (TCC) of the bladder, with moderate urinary toxicity and low systemic absorption. The present phase II study evaluated the activity of biweekly intravesical treatment with Gem using a scheme directly derived from in vitro preclinical studies. Patients with Bacille Calmette-Guérin (BCG) -refractory Ta G3, T1 G1-3 TCC underwent transurethral bladder resection and then intravesical instillation with 2000 mg Gem diluted in 50 ml saline solution on days 1 and 3 for 6 consecutive weeks. Thirty-eight (95%) of the 40 patients showed persistent negative post-treatment cystoscopy and cytology 6 months after Gem treatment, while the remaining 2 patients relapsed at 5 and 6 months. At a median follow-up of 28 months, recurrences had occurred in 14 patients. Among these, four had downstaged (T) disease, three had a lower grade (G) lesion and three had a reduction in both T and G. Urinary and systemic toxicity was very low, with no alterations in biochemical profiles. In conclusion, biweekly instillation of Gem proved active in BCG-refractory Ta G3, T1 G1-3 TCC. Our results highlight the importance of preclinical studies using in vitro systems that adequately reproduce the conditions of intravesical clinical treatment to define the best therapeutic schedule

Exploring the Prognostic Performance of MECKI Score in Heart Failure Patients with Non-Valvular Atrial Fibrillation Treated with Edoxaban

Introduction: Risk stratification in heart failure (HF) is essential for clinical and therapeutic management. The Metabolic Exercise test data combined with Cardiac and Kidney Indexes (MECKI) score is a validated prognostic model for assessing cardiovascular risk in HF patients with reduced ejection fraction (HFrEF). From the validation of the score, the prevalence of HF patients treated with direct oral anticoagulants (DOACs), such as edoxaban, for non-valvular atrial fibrillation (NVAF) has been increasing in recent years. This study aims to evaluate the reliability of the MECKI score in HFrEF patients treated with edoxaban for NVAF. Materials and Methods: This study included consecutive outpatients with HF and NVAF treated with edoxaban (n = 83) who underwent a cardiopulmonary exercise test (CPET). They were matched by propensity score with a retrospective group of HFrEF patients with NVAF treated with vitamin K antagonists (VKAs) from the MECKI score registry (n = 844). The study endpoint was the risk of cardiovascular mortality, urgent heart transplantation, or Left Ventricle Assist Device (LVAD) implantation. Results: Edoxaban patients were treated with a more optimized HF therapy and had different clinical characteristics, with a similar MECKI score. After propensity score, 77 patients treated with edoxaban were successfully matched with the MECKI-VKA control cohort. In both groups, MECKI accurately predicted the composite endpoint with similar area under the curves (AUC = 0.757 vs. 0.829 in the MECKI-VKA vs. edoxaban-treated group, respectively, p = 0.452). The two populations’ survival appeared non-significantly different at the 2-year follow-up. Conclusions: this study confirms the prognostic accuracy of the MECKI score in HFrEF patients with NVAF treated with edoxaban, showing improved predictive power compared to VKA-treated patients

Age at menopause in Latin America

OBJECTIVE: To assess the age at menopause (AM) in Latin America urban areas. DESIGN: A total of 17,150 healthy women, aged 40 to 59 years, accompanying patients to healthcare centers in 47 cities of 15 Latin American countries, were surveyed regarding their age, educational level, healthcare coverage, history of gynecological surgery, smoking habit, presence of menses, and the use of contraception or hormone therapy at menopause. The AM was calculated using logit analysis. RESULTS: The mean age of the entire sample was 49.4 ± 5.5 years. Mean educational level was 9.9 ± 4.5 years, and the use of hormone therapy and oral contraception was 22.1% and 7.9%, respectively. The median AM of women in all centers was 48.6 years, ranging from 43.8 years in Asuncion (Paraguay) to 53 years in Cartagena de Indias (Colombia). Logistic regression analysis determined that women aged 49 living in cities at 2,000 meters or more above sea level (OR = 2.0, 95% CI: 1.4-2.9, P less than 0.001) and those with lower educational level (OR = 1.9, 95% CI: 1.3-2.8, P less than 0.001) or living in countries with low gross national product (OR = 2.1, 95% CI: 1.5-2.9, P less than 0.001) were more prone to an earlier onset of menopause. CONCLUSIONS: The AM varies widely in Latin America. Lower income and related poverty conditions influence the onset of menopause. © 2006 by The North American Menopause Society

Spatio-temporal variability in underwater light climate in a turbid river-floodplain system. Driving factors and estimation using Secchi disc

The underwater light climate has important effects on primary producers. The aim of this research was to evaluate its variability in a turbid river-floodplain system. Photosynthetically active radiation (PAR) was measured in the Middle Paraná River during different hydrological phases to (a) analyse the photosynthetically active radiation attenuation coefficient (k) and euphotic depth (Zeu) as well as their associations with optically active components and (b) develop and evaluate indices and regression models based on Secchi disc (SD) measurements to estimate k and Zeu. Values of k were higher in the fluvial system than in the floodplain and during low-water stage than high-water stage. Particulate components controlled the light climate variability. Chromophoric dissolved organic matter and chlorophyll-a had significant effects during floods. The estimation of k and Zeu was sensitive to temporal but not to spatial variations. The highest prediction accuracy was observed when using specific non-linear regressions for each hydrological phase, especially for Zeu estimation (low stage: k = 1.76 × SD−0.80, Zeu = 2.62 × 1/SD−0.80; high stage: k = 2.04 × SD−0.53, Zeu = 2.26 × 1/SD−0.53). The indices k × SD and Zeu/SD were significantly different from those proposed for clear water environments. It is concluded that temporal variations should be considered when estimating k and Zeu in turbid river-floodplain systems because of the temporal heterogeneity in optically active components. Considering that ecological implication of the light climate depends on Zeu:depth ratio, we propose to estimate Zeu instead of k. Finally, indices proposed for clear water environments are not recommended to be applied to turbid environments.Fil: Mayora, Gisela Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto Nacional de Limnología. Universidad Nacional del Litoral. Instituto Nacional de Limnología; ArgentinaFil: Devercelli, Melina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto Nacional de Limnología. Universidad Nacional del Litoral. Instituto Nacional de Limnología; Argentin

Structural insight into the membrane targeting domain of the Legionella deAMPylase SidD

AMPylation, the post-translational modification with adenosine monophosphate (AMP), is catalyzed by effector proteins from a variety of pathogens. Legionella pneumophila is thus far the only known pathogen that, in addition to encoding an AMPylase (SidM/DrrA), also encodes a deAMPylase, called SidD, that reverses SidM-mediated AMPylation of the vesicle transport GTPase Rab1. DeAMPylation is catalyzed by the N-terminal phosphatase-like domain of SidD. Here, we determined the crystal structure of full length SidD including the uncharacterized C-terminal domain (CTD). A flexible loop rich in aromatic residues within the CTD was required to target SidD to model membranes in vitro and to the Golgi apparatus within mammalian cells. Deletion of the loop (??loop) or substitution of its aromatic phenylalanine residues rendered SidD cytosolic, showing that the hydrophobic loop is the primary membrane-targeting determinant of SidD. Notably, deletion of the two terminal alpha helices resulted in a CTD variant incapable of discriminating between membranes of different composition. Moreover, a L. pneumophila strain producing SidD??loop phenocopied a L. pneumophila ??sidD strain during growth in mouse macrophages and displayed prolonged co-localization of AMPylated Rab1 with LCVs, thus revealing that membrane targeting of SidD via its CTD is a critical prerequisite for its ability to catalyze Rab1 deAMPylation during L. pneumophila infection

Inhibition of Host Vacuolar H+-ATPase Activity by a Legionella pneumophila Effector

Legionella pneumophila is an intracellular pathogen responsible for Legionnaires' disease. This bacterium uses the Dot/Icm type IV secretion system to inject a large number of bacterial proteins into host cells to facilitate the biogenesis of a phagosome permissive for its intracellular growth. Like many highly adapted intravacuolar pathogens, L. pneumophila is able to maintain a neutral pH in the lumen of its phagosome, particularly in the early phase of infection. However, in all cases, the molecular mechanisms underlying this observation remain unknown. In this report, we describe the identification and characterization of a Legionella protein termed SidK that specifically targets host v-ATPase, the multi-subunit machinery primarily responsible for organelle acidification in eukaryotic cells. Our results indicate that after being injected into infected cells by the Dot/Icm secretion system, SidK interacts with VatA, a key component of the proton pump. Such binding leads to the inhibition of ATP hydrolysis and proton translocation. When delivered into macrophages, SidK inhibits vacuole acidification and impairs the ability of the cells to digest non-pathogenic E. coli. We also show that a domain located in the N-terminal portion of SidK is responsible for its interactions with VatA. Furthermore, expression of sidK is highly induced when bacteria begin to enter new growth cycle, correlating well with the potential temporal requirement of its activity during infection. Our results indicate that direct targeting of v-ATPase by secreted proteins constitutes a virulence strategy for L. pneumophila, a vacuolar pathogen of macrophages and amoebae