Computational modeling and prediction of deletion mutants.

In-frame deletion mutations can result in disease. The impact of these mutations on protein structure and subsequent functional changes remain understudied, partially due to the lack of comprehensive datasets including a structural readout. In addition, the recent breakthrough in structure prediction through deep learning demands an update of computational deletion mutation prediction. In this study, we deleted individually every residue of a small α-helical sterile alpha motif domain and investigated the structural and thermodynamic changes using 2D NMR spectroscopy and differential scanning fluorimetry. Then, we tested computational protocols to model and classify observed deletion mutants. We show a method using AlphaFold2 followed by RosettaRelax performs the best overall. In addition, a metric containing pLDDT values and Rosetta ΔΔG is most reliable in classifying tolerated deletion mutations. We further test this method on other datasets and show they hold for proteins known to harbor disease-causing deletion mutations.

SARS-CoV-2 vaccination and risk of severe COVID-19 outcomes in patients with autoimmune hepatitis.

BACKGROUND: Data regarding outcome of Coronavirus disease 2019 (COVID-19) in vaccinated patients with autoimmune hepatitis (AIH) are lacking. We evaluated the outcome of COVID-19 in AIH patients who received at least one dose of Pfizer- BioNTech (BNT162b2), Moderna (mRNA-1273) or AstraZeneca (ChAdOx1-S) vaccine. PATIENTS AND METHODS: We performed a retrospective study on AIH patients with COVID-19. The outcomes of AIH patients who had acute respiratory syndrome coronavirus 2 (SARS-CoV-2) breakthrough infection after at least one dose of COVID-19 vaccine were compared to unvaccinated patients with AIH. COVID-19 outcome was classified according to clinical state during the disease course as: (i) no hospitalization, (ii) hospitalization without oxygen supplementation, (iii) hospitalization with oxygen supplementation by nasal cannula or mask, (iv) intensive care unit (ICU) admission with non-invasive mechanical ventilation, (v) ICU admission with invasive mechanical ventilation or (vi) death, and data was analyzed using ordinal logistic regression. RESULTS: We included 413 (258 unvaccinated and 155 vaccinated) patients (81%, female) with a median age of 52 (range: 17-85) years at COVID-19 diagnosis. The rates of hospitalization were (36.4% vs. 14.2%), need for any supplemental oxygen (29.5% vs. 9%) and mortality (7% vs. 0.6%) in unvaccinated and vaccinated AIH patients with COVID-19. Having received at least one dose of SARS-CoV-2 vaccine was associated with a significantly lower risk of worse COVID-19 severity, after adjusting for age, sex, comorbidities and presence of cirrhosis (adjusted odds ratio [aOR] 0.18, 95% confidence interval [CI], 0.10-0.31). Overall, vaccination against SARS-CoV-2 was associated with a significantly lower risk of mortality from COVID-19 (aOR 0.20, 95% CI 0.11-0.35). CONCLUSIONS: SARS-CoV-2 vaccination significantly reduced the risk of COVID-19 severity and mortality in patients with AIH.

MLK3 mediates impact of PKG1α on cardiac function and controls blood pressure through separate mechanisms.

cGMP-dependent protein kinase 1α (PKG1α) promotes left ventricle (LV) compensation after pressure overload. PKG1-activating drugs improve heart failure (HF) outcomes but are limited by vasodilation-induced hypotension. Signaling molecules that mediate PKG1α cardiac therapeutic effects but do not promote PKG1α-induced hypotension could therefore represent improved therapeutic targets. We investigated roles of mixed lineage kinase 3 (MLK3) in mediating PKG1α effects on LV function after pressure overload and in regulating BP. In a transaortic constriction HF model, PKG activation with sildenafil preserved LV function in MLK3+/+ but not MLK3-/- littermates. MLK3 coimmunoprecipitated with PKG1α. MLK3-PKG1α cointeraction decreased in failing LVs. PKG1α phosphorylated MLK3 on Thr277/Ser281 sites required for kinase activation. MLK3-/- mice displayed hypertension and increased arterial stiffness, though PKG stimulation with sildenafil or the soluble guanylate cyclase (sGC) stimulator BAY41-2272 still reduced BP in MLK3-/- mice. MLK3 kinase inhibition with URMC-099 did not affect BP but induced LV dysfunction in mice. These data reveal MLK3 as a PKG1α substrate mediating PKG1α preservation of LV function but not acute PKG1α BP effects. Mechanistically, MLK3 kinase-dependent effects preserved LV function, whereas MLK3 kinase-independent signaling regulated BP. These findings suggest augmenting MLK3 kinase activity could preserve LV function in HF but avoid hypotension from PKG1α activation.

Comprehensive study of early features in spinocerebellar ataxia 2: delineating the prodromal stage of the disease.

The prodromal phase of spinocerebellar ataxias (SCAs) has not been systematically studied. Main findings come from a homogeneous SCA type 2 (SCA2) population living in Cuba. The aim of this study was to characterize extensively the prodromal phase of SCA2 by several approaches. Thirty-seven non-ataxic SCA2 mutation carriers and its age- and sex-matched controls underwent clinical assessments, including standardized neurological exam, structured interviews and clinical scales, and looking for somatic and autonomic features, as well as a neuropsychological battery, antisaccadic recordings, and MRI scans. Main clinical somatic features of non-ataxic mutation carriers were cramps, sensory symptoms, sleep disorders, and hyperreflexia, whereas predominating autonomic symptoms were pollakiuria/nocturia, constipation, and frequent throat clearing. Cognitive impairments included early deficits of executive functions and visual memory, suggesting the involvement of cerebro-cerebellar-cerebral loops and/or reduced cholinergic basal forebrain input to the cortex. Antisaccadic task revealed impaired oculomotor inhibitory control but preserved ability for error correction. Cognitive and antisaccadic deficits were higher as carriers were closer to the estimated onset of ataxia, whereas higher Scale for the Assessment and Rating of Ataxia (SARA) scores were associated most notably to vermis atrophy. The recognition of early features of SCA2 offers novel insights into the prodromal phase and physiopathological base of the disease, allowing the assessment of its progression and the efficacy of treatments, in particular at early phases when therapeutical options should be most effective.

Early endocrine and molecular changes in metabolic syndrome models.

The twenty-first century arrived in the middle of a global epidemic of metabolic syndrome (MS) and type 2 diabetes mellitus (DM2). It is generally accepted that an excess of nutrients linked to a low physical activity triggers the problem. However, the molecular features that interact to develop the MS are not clear. In an effort to understand and control them, they have been extensively studied, but this goal has not been achieved yet. Nonhuman animal models have been used to explore diet and genetic factors in which experimental conditions are controlled. For example, only one factor in the diet, such as fats or carbohydrates can be modified to better understand a single change that would be impossible in humans. Most of the studies have been done in rodents. However, it is difficult to directly compare them, because experiments are different in more than one variable; genetic strains, amount, and the type of fat used in the diet and sex. Thus, the only possible criteria of comparison are the relevance of the observed changes. We review different animal models and add some original observations on short-term changes in metabolism and beta cells in our own model of adult Wistar rats that are not especially prone to get fat or develop DM2, treated with 20% sucrose in drinking water. One early change observed in pancreatic beta cells is the increase in GLUT2 expression that is located to the membrane of the cells. This change could partially explain the presence of insulin hypersecretion and hyperinsulinemia in these rats. Understanding early changes that lead to MS and in time to pancreatic islet exhaustion is an important biomedical problem that may contribute to learn how to prevent or even reverse MS, before developing DM2.