Investigating the physiological roles of YqjA and YghB: members of the conserved DedA membrane protein family in Escherichia coli

The aim of this dissertation is to characterize the physiological roles of E. coli YqjA and YghB, which belong to the ancient and conserved DedA protein family of hitherto unknown functions. YqjA and YghB are inner membrane (IM) proteins with 61% amino acid identity and share partially redundant functions. Such characterization is achieved by addressing the phenotypes exhibited by strain BC202 with in-frame deletions of yqjA and yghB. BC202 exhibits cell division defects, inability to grow at temperatures above 42°C and an altered phospholipid composition. The cell division defect of BC202 is attributed to the inefficient secretion of periplasmic amidases AmiA and AmiC by the twin arginine transport (Tat) pathway into the periplasm. This inefficient secretion by the Tat pathway is likely related to the loss of proton motive force (PMF) as demonstrated by fluorescence microscopy. In addition, several envelope stress response pathways are induced in BC202 under permissive growth conditions as demonstrated using lacZ fusions. The activation of the stress response pathways in BC202 is largely independent of its cell division and temperature sensitive phenotypes, and demonstrates compromised envelope integrity. The temperature sensitivity of BC202 is also related to the loss of PMF and is rescued by overexpression of the multidrug transporter MdfA or by growth in acidic media. The altered phospholipid composition of BC202 is unrelated to its cell division defect and temperature sensitivity and likely results from an adaptive response to compromised IM integrity. Together, these results suggest possible role(s) of YqjA/YghB in cellular PMF homeostasis and IM quality control either directly as proton transporters or indirectly as regulators of extant cellular mechanisms which participate in the homeostasis of PMF

Multiple envelope stress response pathways are activated in an Escherichia coli strain with mutations in two members of the deda membrane protein family

We have reported that simultaneous deletion of two Escherichia coli genes, yqjA and yghB, encoding related and conserved inner membrane proteins belonging to the DedA protein family results in a number of intriguing phenotypes, including temperature sensitivity at 42°C, altered membrane lipid composition, and cell division defects. We sought to characterize these and other phenotypes in an effort to establish a function for this protein family in E. coli. Here, using reporter assays, we show that the major envelope stress response pathways Cpx, Psp, Bae, and Rcs are activated in strain BC202 (W3110; yqjA yghB) at the permissive growth temperature of 30°C. We previously demonstrated that 10mMMg2+, 400mMNaCl, and overexpression of tatABC are capable of restoring normal growth to BC202 at elevated growth temperatures. Deletion of the cpxR gene from BC202 results in the loss of the ability of these supplements to restore growth at 42°C. Additionally, we report that the membrane potential of BC202 is significantly reduced and that cell division and growth can be restored either by expression of the multidrug transporter MdfA from a multicopy plasmid or by growth at pH 6.0. Together, these results suggest that the DedA family proteins YqjA and YghB are required for general envelope maintenance and homeostasis of the proton motive force under a variety of growth conditions. © 2013, American Society for Microbiology

New functions for the ancient DedA membrane protein family

The DedA protein family is a highly conserved and ancient family of membrane proteins with representatives in most sequenced genomes, including those of bacteria, archaea, and eukarya. The functions of the DedA family proteins remain obscure. However, recent genetic approaches have revealed important roles for certain bacterial DedA family members in membrane homeostasis. Bacterial DedA family mutants display such intriguing phenotypes as cell division defects, temperature sensitivity, altered membrane lipid composition, elevated envelope-related stress responses, and loss of proton motive force. The DedA family is also essential in at least two species of bacteria: Borrelia burgdorferi and Escherichia coli. Here, we describe the phylogenetic distribution of the family and summarize recent progress toward understanding the functions of the DedA membrane protein family. © 2013, American Society for Microbiology

Inefficient Tat-Dependent Export of Periplasmic Amidases in an Escherichia coli Strain with Mutations in Two DedA Family Genes▿

The DedA family genes are found in most bacterial genomes. Two of these proteins are Escherichia coli YqjA and YghB, predicted inner membrane proteins of unknown function sharing 61% amino acid identity. The E. coli single deletion mutants are largely without phenotype, but the double mutant (BC202; ΔyqjA::Tetr ΔyghB::Kanr) is characterized by incomplete cell division, temperature sensitivity, and altered phospholipid levels (K. Thompkins et al., J. Bacteriol. 190:4489-4500, 2008). In this report, we have better characterized the cell division chaining defect of BC202. Fluorescence recovery after photobleaching indicates that 58% of the cells in chains are compartmentalized by at least a cytoplasmic membrane. Green fluorescent protein fusions to the cell division proteins FtsZ, ZipA, FtsI, FtsL, and FtsQ are correctly localized to new septation sites in BC202. Periplasmic amidases AmiC and AmiA, secreted by the twin arginine transport (Tat) pathway, are localized to the cytoplasm in BC202. Overexpression of AmiA, AmiC, or AmiB, a periplasmic amidase secreted via the general secretory pathway, restores normal cell division but does not suppress the temperature sensitivity of BC202, indicating that YghB and YqjA may play additional roles in cellular physiology. Strikingly, overexpression of the Tat export machinery (TatABC) results in normal cell division and growth at elevated temperatures. These data collectively suggest that the twin arginine pathway functions inefficiently in BC202, likely due to the altered levels of membrane phospholipids in this mutant. These results underscore the importance of membrane composition in the proper function of the Tat protein export pathway

BB0250 of Borrelia burgdorferi Is a Conserved and Essential Inner Membrane Protein Required for Cell Division ▿ †

The gene bb0250 of Borrelia burgdorferi is a homolog of the dedA family, encoding integral inner membrane proteins that are present in nearly all species of bacteria. To date, no precise function has been attributed to any dedA gene. Unlike many bacterial species, such as Escherichia coli, which has eight dedA genes, B. burgdorferi possesses only one, annotated bb0250, providing a unique opportunity to investigate the functions of the dedA family. Here, we show that bb0250 is able to restore normal growth and cell division to a temperature-sensitive E. coli mutant with simultaneous deletions of two dedA genes, yqjA and yghB, and encodes a protein that localizes to the inner membrane of E. coli. The bb0250 gene could be deleted from B. burgdorferi only after introduction of a promoterless bb0250 under the control of an inducible lac promoter, indicating that it is an essential gene in this organism. Growth of the mutant in the absence of isopropyl-β-d-thiogalactopyranoside resulted in cell death, preceded by cell division defects characterized by elongated cells and membrane bulges, demonstrating that bb0250 is required for proper cell division and envelope integrity. Finally, we show that BB0250 depletion leads to imbalanced membrane phospholipid composition in borrelia. These results demonstrate a strong conservation of function of the dedA gene family across diverse species of Gram-negative bacteria and a requirement for this protein family for normal membrane lipid composition and cell division

COVID-19 infected ST-Elevation myocardial infarction in India (COSTA INDIA)

Objective: To find out differences in the presentation, management and outcomes of COVID-19 infected STEMI patients compared to age and sex-matched non-infected STEMI patients treated during the same period. Methods: This was a retrospective multicentre observational registry in which we collected data of COVID-19 positive STEMI patients from selected tertiary care hospitals across India. For every COVID-19 positive STEMI patient, two age and sex-matched COVID-19 negative STEMI patients were enrolled as control. The primary endpoint was a composite of in-hospital mortality, re-infarction, heart failure, and stroke. Results: 410 COVID-19 positive STEMI cases were compared with 799 COVID-19 negative STEMI cases. The composite of death/reinfarction/stroke/heart failure was significantly higher among the COVID-19 positive STEMI patients compared with COVID-19 negative STEMI cases (27.1% vs 20.7% p value = 0.01); though mortality rate did not differ significantly (8.0% vs 5.8% p value = 0.13). Significantly lower proportion of COVID-19 positive STEMI patients received reperfusion treatment and primary PCI (60.7% vs 71.1% p value=< 0.001 and 15.4% vs 23.4% p value = 0.001 respectively). Rate of systematic early PCI (pharmaco-invasive treatment) was significantly lower in the COVID-19 positive group compared with COVID-19 negative group. There was no difference in the prevalence of high thrombus burden (14.5% and 12.0% p value = 0.55 among COVID-19 positive and negative patients respectively) Conclusions: In this large registry of STEMI patients, we did not find significant excess in in-hospital mortality among COVID-19 co-infected patients compared with non-infected patients despite lower rate of primary PCI and reperfusion treatment, though composite of in-hospital mortality, re-infarction, stroke and heart failure was higher

Impact of COVID-19 on heart failure hospitalization and outcome in India – A cardiological society of India study (CSI–HF in COVID 19 times study – “The COVID C–HF study”)

Objectives: The presentation and outcomes of acute decompensated heart failure (ADHF) during COVID times (June 2020 to Dec 2020) were compared with the historical control during the same period in 2019. Methods: Data of 4806 consecutive patients of acute HF admitted in 22 centres in the country were collected during this period. The admission patterns, aetiology, outcomes, prescription of guideline-directed medical therapy (GDMT) and interventions were analysed in this retrospective study. Results: Admissions for acute heart failure during the pandemic period in 2020 decreased by 20% compared to the corresponding six-month period in 2019, with numbers dropping from 2675 to 2131. However, no difference in the epidemiology was seen. The mean age of presentation in 2019 was 61.75 (±13.7) years, and 59.97 (±14.6) years in 2020. There was a significant decrease in the mean age of presentation (p = 0.001). Also. the proportion of male patients decreased significantly from 68.67% to 65.84% (p = 0.037). The in-hospital mortality for acute heart failure did not differ significantly between 2019 and 2020 (4.19% and 4.,97%) respectively (p = 0.19). The proportion of patients with HFrEF did not change in 2020 compared to 2019 (76.82% vs 75.74%, respectively). The average duration of hospital stay was 6.5 days. Conclusion: The outcomes of ADHF patients admitted during the Covid pandemic did not differ significantly. The length of hospital stay remained the same. The study highlighted the sub-optimal use of GDMT, though slightly improving over the last few years