Quantum Dot Fluorescent Imaging: Using Atomic Structure Correlation Studies to Improve Photophysical Properties.

Efforts to study intricate, higher-order cellular functions have called for fluorescence imaging under physiologically relevant conditions such as tissue systems in simulated native buffers. This endeavor has presented novel challenges for fluorescent probes initially designed for use in simple buffers and monolayer cell culture. Among current fluorescent probes, semiconductor nanocrystals, or quantum dots (QDs), offer superior photophysical properties that are the products of their nanoscale architectures and chemical formulations. While their high brightness and photostability are ideal for these biological environments, even state of the art QDs can struggle under certain physiological conditions. A recent method correlating electron microscopy ultrastructure with single-QD fluorescence has begun to highlight subtle structural defects in QDs once believed to have no significant impact on photoluminescence (PL). Specific defects, such as exposed core facets, have been shown to quench QD PL in physiologically accurate conditions. For QD-based imaging in complex cellular systems to be fully realized, mechanistic insight and structural optimization of size and PL should be established. Insight from single QD resolution atomic structure and photophysical correlative studies provides a direct course to synthetically tune QDs to match these challenging environments.

A mammalian-specific Alex3/Gαq protein complex regulates mitochondrial trafficking, dendritic complexity, and neuronal survival.

Mitochondrial dynamics and trafficking are essential to provide the energy required for neurotransmission and neural activity. We investigated how G protein-coupled receptors (GPCRs) and G proteins control mitochondrial dynamics and trafficking. The activation of Gαq inhibited mitochondrial trafficking in neurons through a mechanism that was independent of the canonical downstream PLCß pathway. Mitoproteome analysis revealed that Gαq interacted with the Eutherian-specific mitochondrial protein armadillo repeat-containing X-linked protein 3 (Alex3) and the Miro1/Trak2 complex, which acts as an adaptor for motor proteins involved in mitochondrial trafficking along dendrites and axons. By generating a CNS-specific Alex3 knockout mouse line, we demonstrated that Alex3 was required for the effects of Gαq on mitochondrial trafficking and dendritic growth in neurons. Alex3-deficient mice had altered amounts of ER stress response proteins, increased neuronal death, motor neuron loss, and severe motor deficits. These data revealed a mammalian-specific Alex3/Gαq mitochondrial complex, which enables control of mitochondrial trafficking and neuronal death by GPCRs.

Processing of stalled replication forks in Bacillus subtilis.

Accurate DNA replication and transcription elongation are crucial for preventing the accumulation of unreplicated DNA and genomic instability. Cells have evolved multiple mechanisms to deal with impaired replication fork progression, challenged by both intrinsic and extrinsic impediments. The bacterium Bacillus subtilis, which adopts multiple forms of differentiation and development, serves as an excellent model system for studying the pathways required to cope with replication stress to preserve genomic stability. This review focuses on the genetics, single molecule choreography, and biochemical properties of the proteins that act to circumvent the replicative arrest allowing the resumption of DNA synthesis. The RecA recombinase, its mediators (RecO, RecR, and RadA/Sms) and modulators (RecF, RecX, RarA, RecU, RecD2, and PcrA), repair licensing (DisA), fork remodelers (RuvAB, RecG, RecD2, RadA/Sms, and PriA), Holliday junction resolvase (RecU), nucleases (RnhC and DinG), and translesion synthesis DNA polymerases (PolY1 and PolY2) are key functions required to overcome a replication stress, provided that the fork does not collapse.

Quantitative morphological analysis of InP-based quantum dots reveals new insights into the complexity of shell growth.

The incorporation of quantum dots in display technology has fueled a renewed interest in InP-based quantum dots, but difficulty controlling the Zn chemistry during shelling has stymied thick, even ZnSe shell growth. The characteristic uneven, lobed morphology of Zn-based shells is difficult to assess qualitatively and measure through traditional methods. Here, we present a methodological study utilizing quantitative morphological analysis of InP/ZnSe quantum dots to analyze the impact of key shelling parameters on InP core passivation and shell epitaxy. We compare conventional hand-drawn measurements with an open-source semi-automated protocol to showcase the improved precision and speed of this method. Additionally, we find that quantitative morphological assessment can discern morphological trends in morphologies that qualitative methods cannot. In conjunction with ensemble fluorescence measurements, we find that changes to shelling parameters that promote even shell growth often do so at the cost of core homogeneity. These results indicate that the chemistry of passivating the core and promoting shell growth must be balanced carefully to maximize brightness while maintaining emission color-purity.

A New Cycler for Automated Peritoneal Dialysis to Provide Efficient Dialysis and Improved Sleep Quality.

INTRODUCTION: Automated peritoneal dialysis (APD) employs cyclers to control inflow and outflow of the dialysis fluid to the patient's abdomen. To allow more patients to use this modality, cyclers should support the achievement of an adequate dialysis dose and be easy to use, cost-effective, and silent. The new SILENCIA cycler (Fresenius Medical Care, Bad Homburg, Germany), designed to improve these characteristics in comparison to its predecessor device, was evaluated in this respect in a prospective study. METHODS: This cross-over study comprised two 2-week study periods, separated by a 3-week training phase. First, patients underwent APD with their current cycler (PD-NIGHT [Fresenius Medical Care, Bad Homburg, Germany] or HomeChoice Pro [Baxter, Deerfield, IL, USA] as control), followed by training on the SILENCIA cycler. Then, patients were switched to the SILENCIA cycler. During each treatment period, we collected data on total Kt/Vurea, ultrafiltration (UF) volume, patient-reported outcomes (sleep quality, among others), and device handling. RESULTS: Sixteen patients were enrolled; 2 patients terminated the study prematurely before study intervention, 1 patient due to a protocol violation. In 13 patients, total Kt/Vurea and UF could be evaluated. Neither Kt/Vurea nor UF differed significantly between control and SILENCIA cyclers. Out of 10 patients answering the questionnaire on sleep quality after the 2-week phase with the SILENCIA cycler, sleep quality improved in 5 patients; in the other patients, sleep quality was rated unchanged compared to the previously used cycler. The average reported sleep time was 5.9 ± 1.8 h with the PD-NIGHT, 7.2 ± 2.1 h with HomeChoice Pro, and 8.0 ± 1.6 h with the SILENCIA cycler. All patients were much or very much satisfied with the new cycler. CONCLUSION: The SILENCIA cycler delivers adequate urea clearance and UF. Importantly, sleep quality improved, possibly related to less caution messages and alarms.

Bacillus subtilis RadA/Sms-Mediated Nascent Lagging-Strand Unwinding at Stalled or Reversed Forks Is a Two-Step Process: RadA/Sms Assists RecA Nucleation, and RecA Loads RadA/Sms.

Replication fork rescue requires Bacillus subtilis RecA, its negative (SsbA) and positive (RecO) mediators, and fork-processing (RadA/Sms). To understand how they work to promote fork remodeling, reconstituted branched replication intermediates were used. We show that RadA/Sms (or its variant, RadA/Sms C13A) binds to the 5'-tail of a reversed fork with longer nascent lagging-strand and unwinds it in the 5'→3' direction, but RecA and its mediators limit unwinding. RadA/Sms cannot unwind a reversed fork with a longer nascent leading-strand, or a gapped stalled fork, but RecA interacts with and activates unwinding. Here, the molecular mechanism by which RadA/Sms, in concert with RecA, in a two-step reaction, unwinds the nascent lagging-strand of reversed or stalled forks is unveiled. First, RadA/Sms, as a mediator, contributes to SsbA displacement from the forks and nucleates RecA onto single-stranded DNA. Then, RecA, as a loader, interacts with and recruits RadA/Sms onto the nascent lagging strand of these DNA substrates to unwind them. Within this process, RecA limits RadA/Sms self-assembly to control fork processing, and RadA/Sms prevents RecA from provoking unnecessary recombination.

High Plasma Levels of Fibroblast Growth Factor 23 Are Associated with Increased Risk of COVID-19 in End-Stage Renal Disease Patients on Hemodialysis: Results of a Prospective Cohort.

End-stage renal disease (ESRD) patients are a population with high rates of COVID-19 and mortality. These patients present a low response to anti-SARS-CoV-2 immunization, which is associated with immune dysfunction. ESRD patients also present high plasma titers of Fibroblast Growth Factor 23 (FGF23), a protein hormone that reduces immune response in vivo and in vitro. Increased FGF23 levels associate with higher infection-related hospitalizations and adverse infectious outcomes. Thus, we evaluated whether ESRD patients with high FGF23 titers have an increased rate of SARS-CoV-2 infection. METHODS: We performed a prospective cohort of ESRD patients in hemodialysis who had measurements of plasma intact FGF23 in 2019. We determined COVID-19 infections, hospitalizations, and mortality between January 2020 and December 2021. RESULTS: We evaluated 243 patients. Age: 60.4 ± 10.8 years. Female: 120 (49.3%), diabetes: 110 (45.2%). During follow-up, 45 patients developed COVID-19 (18.5%), 35 patients were hospitalized, and 12 patients died (mortality rate: 26.6%). We found that patients with higher FGF23 levels (defined as equal or above median) had a higher rate of SARS-CoV-2 infection versus those with lower levels (18.8% versus 9.9%; Hazard ratio: 1.92 [1.03-3.56], p = 0.039). Multivariate analysis showed that increased plasma FGF23 was independently associated with SARS-CoV-2 infection and severe COVID-19. DISCUSSION: Our results suggest that high plasma FGF23 levels are a risk factor for developing COVID-19 in ESRD patients. These data support the potential immunosuppressive effects of high circulating FGF23 as a factor implicated in the association with worse clinical outcomes. Further data are needed to confirm this hypothesis.

Triagem de Manchester: Perceções dos enfermeiros sobre os seus contributos e fatores que a influenciam / Manchester Triage: Nurses' perceptions of its contributions and influencing factors / Triaje de Mánchester: Percepción de los enfermeros sobre su contribución y factores que influyen en ella

Resumo Enquadramento: A Triagem de Manchester é essencial nos serviços de urgência, permitindo o estabelecimento da prioridade clínica do utente e o tempo recomendado até à observação médica, podendo ser condicionada por vários fatores. Objetivos: Conhecer a perceção dos enfermeiros sobre os contributos da Triagem de Manchester e fatores influenciadores. Metodologia: Estudo exploratório descritivo, de cariz qualitativo, com entrevistas semiestruturadas a 10 enfermeiros de dois serviços de urgência de Portugal, entre janeiro e fevereiro de 2020, submetidas a análise de conteúdo. Resultados: Emergiram três áreas temáticas - contributos da Triagem de Manchester, fatores que influenciam a sua realização e sugestões de melhoria. Conclusão: Identificaram-se como contributos da Triagem de Manchester - estabelecimento de prioridades, atendimento ao utente e melhoria do funcionamento do serviço; e como fatores que influenciam a realização da triagem: recursos humanos e materiais, estrutura física, protocolo de Triagem de Manchester, gestão do serviço e relacionados com o utente e com o enfermeiro. Sugerem-se melhorias em - gestão do serviço, protocolo de Triagem de Manchester e seleção dos enfermeiros.

Abstract Background: The Manchester Triage (MT) is essential in emergency services, setting the patient's clinical priority and the recommended time until medical assessment. Different factors can condition it. Objective: To know the nurses' perceptions of the contributions of the MT and its influencing factors. Methodology: This is an exploratory, descriptive, and qualitative study, with semi-structured interviews with ten nurses from two Portuguese emergency departments, conducted between January and February 2020 and submitted to content analysis. Results: Three thematic areas emerged - The contributions of the MT, Factors influencing its performance, and Suggestions for improvement. Conclusion: The contributions of the MT regarded priority setting, patient service, and the improvement of service operation. Factors that influenced triage performance were human and material resources, physical structures, MT protocol, service management, and patient- and nurse-related factors. Service management, MT protocol, and nurse selection were areas suggested for improvement.

Resumen Marco contextual: El Triaje de Mánchester es fundamental en los servicios de urgencias, ya que permite establecer la prioridad clínica del paciente y el tiempo recomendado hasta la atención médica, que puede estar condicionada por diversos factores. Objetivos: Conocer la percepción de los enfermeros sobre las aportaciones del Triaje de Mánchester y los factores que influyen. Metodología: Estudio descriptivo exploratorio, de carácter cualitativo, con entrevistas semiestructuradas a 10 enfermeros de dos servicios de urgencias de Portugal entre enero y febrero de 2020, sometidos a un análisis de contenido. Resultados: Surgieron tres áreas temáticas: contribuciones del Triaje de Mánchester, factores que influyen en su aplicación y sugerencias de mejora. Conclusión: Se identificaron como contribuciones del Triaje de Mánchester, establecimiento de prioridades, atención al usuario y mejora del funcionamiento del servicio; y como factores que influyen en la realización del triaje, recursos humanos y materiales, estructura física, protocolo del Triaje de Mánchester, gestión del servicio y relación con el usuario y el enfermero. Se sugieren mejoras en la gestión del servicio, el protocolo Triaje de Mánchester y la selección de enfermeros.

Reconstrucción ancestral de una B-lactamasa y comparativa con sus homólogos actuales / Ancestral reconstruction of a B-lactamase and comparison with its extant proteins

Las B-lactamasas son proteínas de origen bacteriano que se caracterizan por hidrolizar los antibióticos B-lactámicos, confiriendo resistencia microbiana ante estos. Son una familia heterogénea de proteínas muy relevantes desde el punto de vista sanitario debido a la facilidad que presentan para adquirir resistencia a nuevos fármacos por su alta capacidad de evolución.La evolución in vitro de estas proteínas ha servido, no solo para desarrollar su caracterización y mejorar su conocimiento, sino como una nueva línea de investigación que permite identificar de manera predictiva residuos implicados en la adquisición de resistencia frente antibióticos. Al mismo tiempo, el método de reconstrucción ancestral de proteínas se ha revelado como una herramienta novedosa y útil para comprender la evolución de las β- lactamasas y entender algunas de sus características como es su promiscuidad. En este trabajo, se ha realizado un estudio de B-lactamasas ancestrales reconstruidas a partir de la filogenia de B-lactamasas existentes de clase A. De las cuatro proteínas ancestrales estudiadas, se ha obtenido una que es funcional y se ha comparado su actividad hidrolítica con la de cuatro de sus homólogos actuales frente a ocho fármacos B-lactámicos. Se ha comprobado que esta proteína ancestral tiene una actividad frente aantibióticos más generalista que cualquier de las proteínas actuales estudiadas. Además, la proteína ancestral activa mostró más resistencia frente a uno de los fármacos utilizados que el resto de B-lactamasas existentes. Finalmente se han discutido estos resultados y a partir de ellos se argumenta por qué las secuencias ancestrales reconstruidas pueden ser un punto de partida muy atractivo a la hora de realizar evolución dirigida de proteínas para la obtención de proteínas de interés biotecnológico. (AU)

The B-lactamases are proteins of bacterial origin that are characterized by hydrolyzing antibiotics B-lactams, conferring microbial resistance against them. They are a heterogeneous family of proteins very relevant from a health point of view due to the ease they present to acquire resistance to new drugs due to their high capacity for evolution.The in vitro evolution of these proteins has served not only to develop their characterization and improve their knowledge, but as a new line of research that allows to predictively identify residues involved in the acquisition of antibiotic resistance.At the same time, the method of ancestral protein reconstruction has been revealed as a novel and useful tool to understand the evolution of B-lactamases and understand some of their characteristics such as their promiscuity.In this work, a study of ancestral B-lactamases reconstructed from the phylogeny of existing class A B-lactamases has been carried out. Of the four ancestral proteins studied, one has been obtained that is functional and has compared its hydrolytic activity with that of four of its current counterparts against eight β-lactam drugs. This ancestral protein has been shown to have a more generalistic antibiotic activity than any of the current proteins studied. In addition, the active ancestral protein showed more resistance to one of the drugs used than the rest of B-lactamases existing. Finally these results have been discussed and from them it is argued why reconstructed ancestral sequences can be a very attractive starting point when it comes to direct evolution of proteins for obtaining proteins of biotechnological interest.

Complete Genome Sequence of Gordonia rubripertincta Bacteriophage Hexbug Suggests Potential for a New CT Subcluster.

Through the SEA-PHAGES program at Tufts University, a bacteriophage infecting Gordonia rubripertincta NRRL B-16540 was isolated and characterized. Hexbug is a lytic phage and is currently one of 44 phages belonging to cluster CT. The Hexbug genome shares >96% nucleotide identity with cluster CT phage Orla.

[Effectiveness of regional citrate anticoagulation in continuous renal replacement therapy]. / Efectividad de anticoagulación regional con citrato en terapia de reemplazo renal continua.

BACKGROUND: Anticoagulation in continuous renal replacement therapy (CRRT) is essential to counteract the coagulation cascade activation, induced by the dialysis circuit. Heparin is the most widely used anticoagulant, followed by regional citrate anticoagulation (RCA). AIM: To determine the effectiveness and safety of anticoagulant treatment with citrate in CRRT. MATERIAL AND METHODS: Retrospective study of adults in CRRT hospitalized between the years 2014 and 2020 in critical units, who required change to RCA according to established protocols. RESULTS: We studied 24 patients aged 63 ± 13 years (12 females). The reasons for admission were acute kidney injury (AKI) in 80% and stage 5 chronic kidney disease in 20%. The indication of RCA in 75% of patients was by coagulation of more than 3 circuits in 24 hours. The duration of the circuit in RCA was 18.5 ± 4.8 hours versus 11.9 ± 4.9 hours with heparin (p < 0.0001). There were 19 mild complications that did not affect the RCA. CONCLUSIONS: RCA is feasible to perform, it is a safe and efficient procedure if it is protocolized, allowing a longer duration of the dialysis circuit.

Clinical Efficacy of SARS-CoV-2 Vaccination in Hemodialysis Patients.

Introduction: The COVID-19 pandemic is a global public health problem. Patients with end-stage renal disease on hemodialysis are at a higher risk of infection and mortality than the general population. Worldwide, a vaccination campaign has been developed that has been shown to reduce severe infections and deaths in the general population. However, there are currently limited data on the clinical efficacy of vaccinations in the hemodialysis population. Methods: A national multicenter observational cohort was performed in Chile to evaluate the clinical efficacy of anti-SARS-CoV-2 vaccination in end-stage renal disease patients on chronic hemodialysis from February 2021 to August 2021. In addition, the BNT162b2 (Pfizer-BioNTech) and CoronaVac (Sinovac) vaccines were evaluated. The efficacy of vaccination in preventing SARS-CoV-2 infection, hospitalizations, and deaths associated with COVID-19 was determined. Results: A total of 12,301 patients were evaluated; 10,615 (86.3%) received a complete vaccination (2 doses), 490 (4.0%) received incomplete vaccination, and 1196 (9.7%) were not vaccinated. During follow-up, 1362 (11.0%) patients developed COVID-19, and 150 died (case fatality rate: 11.0%). The efficacy of the complete vaccination in preventing infection was 18.1% (95% confidence interval [CI]:11.8-23.8%), and prevention of death was 66.0% (95% CI:60.6-70.7%). When comparing both vaccines, BNT162b2 and CoronaVac were effective in reducing infection and deaths associated with COVID-19. Nevertheless, the BNT162b2 vaccine had higher efficacy in preventing infection (42.6% vs. 15.0%) and deaths (90.4% vs. 64.8%) compared to CoronaVac. Conclusion: The results of our study suggest that vaccination against SARS-CoV-2 in patients on chronic hemodialysis was effective in preventing infection and death associated with COVID-19.

Survival study and factors associated with mortality in Chilean patients on peritoneal dialysis infected with SARS-CoV-2.

The Covid-19 pandemic has been responsible for millions of deaths worldwide. Patients with comorbidities- such as those on peritoneal dialysis (PD)- present higher morbidity and mortality than the general population. We prospectively evaluated all Chilean patients on PD (48 centres) and followed those who had Covid-19 from the beginning of the Covid-19 pandemic in Chile (March 2020) to January 2021 (start of vaccination campaign). We described demographic history, comorbidities, factors related to infection, need for hospitalisation and death due to Covid-19. During the study period, 106 adults on PD were infected by SARS-CoV-2, with a mean age of 53.1 (±16.3) and of which 53.9% were female. From that group, 54.8% required hospitalisation and 24.5% (n = 26) died due to Covid-19. Most of the patients (63.4%) were infected at home and 22.8% during hospitalisation for other reasons. There was a significant association for Covid-19 mortality with: being ≥60 years old, diabetes, time on PD ≥5 years, need for hospitalisation and hospital-acquired infection. At 90 days of follow-up, all deaths associated to Covid-19 occurred before 40 days. We conclude that patients on PD without Covid-19 vaccination have a high mortality and need for hospitalisation associated to Covid-19. To avoid this negative outcome, it is necessary to intensify strategies to avoid contagion, especially in those ≥60 years old, with diabetes and/or ≥5 years spent on PD.

Teratoma mediastinal con áreas de malignidad, reporte de un caso y discusión / Mediastinal Teratoma with Malignant Areas: Case Report and Discussion

Resumen Varios tipos de tumores pueden surgir en el mediastino anterior, de los cuales los tumores de células germinales constituyen el 10- 15%, y el más frecuente es el teratoma maduro. El hallazgo de un componente maligno en un teratoma maduro es raro, habiéndose descrito pocos casos de malignidad en forma de carcinoma escamoso, adenocarcinoma, sarcoma o tumores neuroendocrinos. Presentamos el caso de una mujer joven con diagnóstico de teratoma maduro y desarrollo de adenocarcinoma en la pared, lo que confiere un pronóstico desfavorable, con opciones de tratamiento mal definidas dada la excepcionalidad de la enfermedad. Este caso clínico destaca que se requiere un muestreo histopatológico cuidadoso de las áreas sólidas en un teratoma, incluso en pacientes jóvenes cuyas lesiones son más pequeñas. Aunque existen recomendaciones a favor de la quimioterapia, se basan en series de un número limitado de pacientes. La resección completa de la neoplasia y el seguimiento multidisciplinario serán de relevancia para el control de las recidivas locales y a distancia.

Abstract Several types of tumors may occur in the anterior mediastinum, of which germ cell tumors constitute 10-15%, the most frequent being the mature teratoma. The finding of a malignant component in mature teratoma is rare, and few cases of malignancies such as squamous carcinoma, adenocarcinoma, sarcoma, or neuroendocrine tumors have been described. We present the case of a young woman diagnosed with mature teratoma and development of adenocarcinoma within tumor wall, conferring an unfavorable prognosis, with poorly defined treatment options given the exceptional a mature of the disease. This clinical case highlights the fact that careful histopathological sampling of solid areas is required in a teratoma, even in young patients whose lesions are smaller. Although there are recommendations in favor of chemotherapy, they are based on series of a limited number of patients. Complete resection of the neoplasm and multidisciplinary follow-up would be of relevance for the control of local and distant recurrences.

Efectividad de anticoagulación regional con citrato en terapia de reemplazo renal continua

Background: Anticoagulation in continuous renal replacement therapy (CRRT) is essential to counteract the coagulation cascade activation, induced by the dialysis circuit. Heparin is the most widely used anticoagulant, followed by regional citrate anticoagulation (RCA). Aim: To determine the effectiveness and safety of anticoagulant treatment with citrate in CRRT. Material and Methods: Retrospective study of adults in CRRT hospitalized between the years 2014 and 2020 in critical units, who required change to RCA according to established protocols. Results: We studied 24 patients aged 63 ± 13 years (12 females). The reasons for admission were acute kidney injury (AKI) in 80% and stage 5 chronic kidney disease in 20%. The indication of RCA in 75% of patients was by coagulation of more than 3 circuits in 24 hours. The duration of the circuit in RCA was 18.5 ± 4.8 hours versus 11.9 ± 4.9 hours with heparin (p < 0.0001). There were 19 mild complications that did not affect the RCA. Conclusions: RCA is feasible to perform, it is a safe and efficient procedure if it is protocolized, allowing a longer duration of the dialysis circuit.

PcrA Dissociates RecA Filaments and the SsbA and RecO Mediators Counterbalance Such Activity.

PcrA depletion is lethal in wild-type Bacillus subtilis cells. The PcrA DNA helicase contributes to unwinding RNA from the template strand, backtracking the RNA polymerase, rescuing replication-transcription conflicts, and disassembling RecA from single-stranded DNA (ssDNA) by poorly understood mechanisms. We show that, in the presence of RecA, circa one PcrA/plasmid-size circular ssDNA (cssDNA) molecule hydrolyzes ATP at a rate similar to that on the isolated cssDNA. PcrA K37A, which poorly hydrolyses ATP, fails to displace RecA from cssDNA. SsbA inhibits and blocks the ATPase activities of PcrA and RecA, respectively. RecO partially antagonizes and counteracts the negative effect of SsbA on PcrA- and RecA-mediated ATP hydrolysis, respectively. Conversely, multiple PcrA molecules are required to inhibit RecA·ATP-mediated DNA strand exchange (DSE). RecO and SsbA poorly antagonize the PcrA inhibitory effect on RecA·ATP-mediated DSE. We propose that two separable PcrA functions exist: an iterative translocating PcrA monomer strips RecA from cssDNA to prevent unnecessary recombination with the mediators SsbA and RecO balancing such activity; and a PcrA cluster that disrupts DNA transactions, as RecA-mediated DSE.

Bacillus subtilis RecA, DisA, and RadA/Sms Interplay Prevents Replication Stress by Regulating Fork Remodeling.

Reviving Bacillus subtilis spores require the recombinase RecA, the DNA damage checkpoint sensor DisA, and the DNA helicase RadA/Sms to prevent a DNA replication stress. When a replication fork stalls at a template lesion, RecA filaments onto the lesion-containing gap and the fork is remodeled (fork reversal). RecA bound to single-strand DNA (ssDNA) interacts with and recruits DisA and RadA/Sms on the branched DNA intermediates (stalled or reversed forks), but DisA and RadA/Sms limit RecA activities and DisA suppresses its c-di-AMP synthesis. We show that RecA, acting as an accessory protein, activates RadA/Sms to unwind the nascent lagging-strand of the branched intermediates rather than to branch migrate them. DisA limits the ssDNA-dependent ATPase activity of RadA/Sms C13A, and inhibits the helicase activity of RadA/Sms by a protein-protein interaction. Finally, RadA/Sms inhibits DisA-mediated c-di-AMP synthesis and indirectly inhibits cell proliferation, but RecA counters this negative effect. We propose that the interactions among DisA, RecA and RadA/Sms, which are mutually exclusive, contribute to generate the substrate for replication restart, regulate the c-di-AMP pool and limit fork restoration in order to maintain cell survival.

A Novel Biotinylated Homotryptamine Derivative for Quantum Dot Imaging of Serotonin Transporter in Live Cells.

The serotonin transporter (SERT) is the primary target for selective serotonin reuptake inhibitor (SSRI) antidepressants that are thought to exert their therapeutic effects by increasing the synaptic concentration of serotonin. Consequently, probes that can be utilized to study cellular trafficking of SERT are valuable research tools. We have developed a novel ligand (IDT785) that is composed of a SERT antagonist (a tetrahydro pyridyl indole derivative) conjugated to a biotinylated poly ethylene glycol (PEG) via a phenethyl linker. This compound was determined to be biologically active and inhibited SERT-mediated reuptake of IDT307 with the half-maximal inhibitory concentration of 7.2 ± 0.3 µM. We demonstrated that IDT785 enabled quantum dot (QD) labeling of membrane SERT in transfected HEK-293 cultures that could be blocked using the high affinity serotonin reuptake inhibitor paroxetine. Molecular docking studies suggested that IDT785 might be binding to the extracellular vestibule binding site rather than the orthosteric substrate binding site, which could be attributable to the hydrophilicity of the PEG chain and the increased loss of degrees of freedom that would be required to penetrate into the orthosteric binding site. Using IDT785, we were able to study the membrane localization and membrane dynamics of YFP-SERT heterologously expressed in HEK-293 cells and demonstrated that SERT expression was enriched in the membrane edge and in thin cellular protrusions.

[Rhabdomyolysis as the presentation form of COVID-19 infection. Report of one case]. / Rabdomiólisis como presentación inicial de COVID-19. Caso Clínico.

COVID-19 infection causes a systemic inflammatory response, which mainly presents as a febrile syndrome with respiratory involvement. We report a 37-year-old male who consulted for myalgia, nausea and epigastric pain lasting three days. On admission, he had crepitations at the lung bases. The initial laboratory showed a creatine kinase of 62,768 U/L, a LDH of 1,110 IU/L, a creatinine a 2.1 mg/dL, an aspartate aminotransferase of 1,347 IU/L, a D-dimer of 1,140 ng/mL, a ferritin of 1,201 ng/mL and a lymphocyte count of 810 cells/mm3. The chest CT scan was compatible with multifocal pneumonia, suggesting a COVID-19 infection. COVID-19 PCR was positive. The patient was managed with hydration, sodium bicarbonate, ceftriaxone, and azithromycin, with a good clinical response.