TFIID Enables RNA Polymerase II Promoter-Proximal Pausing.

RNA polymerase II (RNAPII) transcription is governed by the pre-initiation complex (PIC), which contains TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, RNAPII, and Mediator. After initiation, RNAPII enzymes pause after transcribing less than 100 bases; precisely how RNAPII pausing is enforced and regulated remains unclear. To address specific mechanistic questions, we reconstituted human RNAPII promoter-proximal pausing in vitro, entirely with purified factors (no extracts). As expected, NELF and DSIF increased pausing, and P-TEFb promoted pause release. Unexpectedly, the PIC alone was sufficient to reconstitute pausing, suggesting RNAPII pausing is an inherent PIC function. In agreement, pausing was lost upon replacement of the TFIID complex with TATA-binding protein (TBP), and PRO-seq experiments revealed widespread disruption of RNAPII pausing upon acute depletion (t = 60 min) of TFIID subunits in human or Drosophila cells. These results establish a TFIID requirement for RNAPII pausing and suggest pause regulatory factors may function directly or indirectly through TFIID.

Water quality monitoring and modeling for an urban storm drainage channel in Thane, India.

The absence of a sewer system and inadequate wastewater treatment plants results in a discharge of untreated wastewater to the urban drainage channels and pollutes receiving waters. Field visits were carried out to observe water quality parameters such as dissolved oxygen (DO), biochemical oxygen demand (BOD), and chemical oxygen demand (COD) in an urban drainage system (Kolshet drain) in Thane City, Mumbai Metropolitan Region, India. Dye-tracing studies using rhodamine WT dye were used for computing the velocity, discharge, and dispersion coefficient of the drain. The data analysis shows that the BOD and COD values in the drain are higher than the permissible limits (30 mg L-1 for BOD and 250 mg L-1 for COD), which is not suitable for disposal to any receiving water body. Also, the DO was less than the permissible limit of a minimum of 3 mg L-1 (for the survival of aquatic life). It is seen that the higher BOD load significantly reduced the DO throughout the drain. The Water Quality Analysis Simulation Program (WASP 8.32, 2019) developed by the US Environmental Protection Agency (USEPA) has been used for the simulation of the DO and BOD in the drainage channel. The model simulates an appropriate estimate of the expected variation of DO and BOD at points of interest. The modeling for the Kolshet drain is expected to enable better estimates of the wastewater parameters and the pollution transport in the drain for planning purposes.

Response to botulinum toxin may predict response to peroral pyloromyotomy in patients with gastroparesis.

BACKGROUND : Patients with gastroparesis who have undergone prior intrapyloric botulinum toxin injection (BTI) may seek an opinion regarding peroral pyloromyotomy (POP). There are only two small reports assessing the role of BTI as a predictor for successful treatment with POP. METHODS: We performed a retrospective cohort study to assess whether symptomatic improvement after BTI predicts a response to POP. We included 119 patients who had undergone both BTI and POP at Cleveland Clinic Ohio or Cleveland Clinic Florida from January 2016 to September 2019. RESULTS: 65.5 % of patients had symptomatic improvement after BTI. Gastroparesis Cardinal Symptom Index (GCSI) scores were available for 74 patients, with 64 % achieving a response to POP, defined as a decrease in mean GCSI ≥ 1. In multivariable analysis, response to BTI (odds ratio [OR] 7.7 [95 %CI 2.2-26.1]) and higher pre-POP GCSI score (OR 2.3 [95 %CI 1.2-4.6]) were independent predictors of response to POP. CONCLUSIONS: Clinical improvement after BTI is a predictor of response to POP in patients with gastroparesis. This information may aid in improving patient selection for POP.

TAF8 regions important for TFIID lobe B assembly or for TAF2 interactions are required for embryonic stem cell survival.

The human general transcription factor TFIID is composed of the TATA-binding protein (TBP) and 13 TBP-associated factors (TAFs). In eukaryotic cells, TFIID is thought to nucleate RNA polymerase II (Pol II) preinitiation complex formation on all protein coding gene promoters and thus, be crucial for Pol II transcription. TFIID is composed of three lobes, named A, B, and C. A 5TAF core complex can be assembled in vitro constituting a building block for the further assembly of either lobe A or B in TFIID. Structural studies showed that TAF8 forms a histone fold pair with TAF10 in lobe B and participates in connecting lobe B to lobe C. To better understand the role of TAF8 in TFIID, we have investigated the requirement of the different regions of TAF8 for the in vitro assembly of lobe B and C and the importance of certain TAF8 regions for mouse embryonic stem cell (ESC) viability. We have identified a region of TAF8 distinct from the histone fold domain important for assembling with the 5TAF core complex in lobe B. We also delineated four more regions of TAF8 each individually required for interacting with TAF2 in lobe C. Moreover, CRISPR/Cas9-mediated gene editing indicated that the 5TAF core-interacting TAF8 domain and the proline-rich domain of TAF8 that interacts with TAF2 are both required for mouse embryonic stem cell survival. Thus, our study defines distinct TAF8 regions involved in connecting TFIID lobe B to lobe C that appear crucial for TFIID function and consequent ESC survival.

Bariatric Surgery Reduces Cancer Risk in Adults With Nonalcoholic Fatty Liver Disease and Severe Obesity.

BACKGROUND & AIMS: Nonalcoholic fatty liver disease (NAFLD) is associated with obesity and increased risk of cancer. The impacts of bariatric surgery on cancer risk in NAFLD patients are unknown. We investigated the effect of bariatric surgery on cancer risk in patients with NAFLD and severe obesity using the MarketScan database. METHODS: We conducted a retrospective cohort study of 18 to 64 years old newly diagnosed NAFLD patients with severe obesity between 2007 and 2017. We used Cox proportional hazard models to examine the association between bariatric surgery, modeled as a time-varying covariate, and the risks of any cancer and obesity-related cancer, while accounting for confounding using inverse probability of treatment weighting (IPTW). RESULTS: A total of 98,090 patients were included in the study, 33,435 (34.1%) received bariatric surgery. In those without surgery, 1898 incident cases of cancer occurred over 115,890.11 person-years of follow-up, compared with 925 cancer cases over 67,389.82 person-years among surgery patients (crude rate ratio, 0.84; 95% CI, 0.77- 0.91). The IPTW-adjusted risk of any cancer and obesity-related cancer was reduced by 18% (hazard ratio, 0.82; 95% CI, 0.76-0.89) and 25% (hazard ratio, 0.65; 95% CI, 0.56-0.75), respectively, in patients with versus without bariatric surgery. The adjusted risks of any cancer and obesity-related cancer were significantly lower in cirrhotic versus non-cirrhotic patients who underwent surgery. In cancer-specific models, bariatric surgery was associated with significant risk reductions for colorectal, pancreatic, endometrial, thyroid cancers, hepatocellular carcinoma, and multiple myeloma. CONCLUSION: Bariatric surgery was associated with significant reductions in the risks of any cancer and obesity-related cancer in NAFLD patients with severe obesity.

The Use of Pectoralis Blocks in Breast Surgery: A Practice Advisory and Narrative Review from the Society for Ambulatory Anesthesia (SAMBA).

Although pectoralis (PECS) blocks are commonly used for breast surgery, recommendations regarding the efficacy of these blocks have thus far not been developed by any professional anesthesia society. Given the potential impact of PECS blocks on analgesia after outpatient breast surgery, The Society for Ambulatory Anesthesia (SAMBA) convened a task force to develop a practice advisory on the use of this analgesic technique. In this practice advisory, we compare the efficacy of PECS blocks with systemic analgesia, local infiltration anesthesia, and paravertebral blockade. Our objectives were to advise on two clinical questions. (1) Does PECS-1 and/or -2 blockade provide more effective analgesia for breast-conserving surgery than either systemic analgesics or surgeon-provided local infiltration anesthesia? (2) Does PECS-1 and/or -2 blockade provide equivalent analgesia for mastectomy compared with a paravertebral block (PVB)? Among patients undergoing breast-conserving surgery, PECS blocks moderately reduce postoperative opioid use, prolong time to analgesic rescue, and decrease postoperative pain scores when compared with systemic analgesics. SAMBA recommends the use of a PECS-1 or -2 blockade in the absence of systemic analgesia (Strength of Recommendation A). No evidence currently exists that strongly favors the use of PECS blocks over surgeon-performed local infiltration in this surgical population. SAMBA cannot recommend PECS blocks over surgical infiltration (Strength of Recommendation C). For patients undergoing a mastectomy, a PECS block may provide an opioid-sparing effect similar to that achieved with PVB; SAMBA recommends the use of a PECS block if a patient is unable to receive a PVB (Strength of Recommendation A).

Comparison of no stent fixation, endoscopic suturing, and a novel over-the-scope clip for stent fixation in preventing migration of fully covered self-expanding metal stents: a retrospective comparative study (with video).

BACKGROUND AND AIMS: Endoscopic suturing and over-the-scope clips (OTSCs) are used to prevent migration of fully covered self-expandable metal stents (FCSEMSs). Recently, a dedicated OTSC was developed for securing FCSEMSs. Our primary aim was to compare the frequency of stent migration without stent fixation versus fixation with suturing or OTSCs, and out secondary aims were to compare clinical success, procedure duration, and adverse events. METHODS: A retrospective cohort study evaluated the outcome of stent placement throughout the entire GI tract from 2013 to 2021. Stent migration was determined as stent displacement ≥2 cm endoscopically or radiographically. Clinical success was defined as resolution of indication at follow-up. RESULTS: Four hundred thirty-three procedures were performed, 239 (55%) without fixation, 140 (32%) with suturing, and 54 (12%) with OTSCs. Stent migration rates were 62% without fixation, 57% with suturing, and 35% with OTSCs (P = .013). The median time to stent migration was 3 weeks without fixation, 5 weeks with suturing, and 6 weeks with OTSCs (P = .023). The clinical success rate was 43%. The median procedure time for OTSCs was shorter compared with suturing (42 vs 68 minutes, P = .002). Adverse event rates trended toward being lowest with OTSCs at 9% compared with 21% without fixation and 18% with suturing (P > .05). CONCLUSIONS: OTSCs for stent fixation were found to have significantly lower migration rates compared with no fixation and suturing. Moreover, OTSCs were associated with decreased overall procedure time and total costs per procedure while trending to be associated with fewer adverse events.

Endoscopic ultrasound with combined fine needle aspiration plus biopsy improves diagnostic yield in solid pancreatic masses.

GOALS: Our aim was to compare the diagnostic yield of endoscopic ultrasound guided fine needle aspiration (EUS-FNA) versus combined fine needle aspiration and fine needle biopsy (EUS-FNA + FNB) in the evaluation of solid pancreatic masses (SPMs). BACKGROUND: EUS-FNA and EUS-FNB are established methods to diagnose SPMs. No studies have evaluated the efficacy of combination of both (EUS-FNA + FNB). Our senior author (MRS) hypothesized that combining the two techniques by using a single FNB needle improves diagnostic yield and started combination technique in October 2016. STUDY: Patients who underwent EUS for SPMs by MRS during January 2014-September 2019 were included. They were divided into the EUS-FNA group and EUS-FNA + FNB group. EUS-FNA was performed using a 22 or 25 gauge Expect Slimline needle (Boston Scientific, Marlborough, MA) and EUS-FNA + FNB was performed using a single 22 or 25 gauge Shark-core needle (Medtronics, Minneapolis, MN, USA). Our primary outcome was to compare the diagnostic yield in the two groups. RESULTS: Among 105 patients included, 58 were in the EUS-FNA group and 47 were in the EUS-FNA + FNB group. EUS-FNA + FNB group had significantly higher diagnostic yield and required fewer needle passes compared to EUS-FNA group, 95.7% vs. 77.6%, p = .01: and 4 vs. 5, p = .002; respectively. Procedural duration was similar in both groups but the combined technique required less number of needles per procedure. There was no difference in adverse events in the two groups. CONCLUSION: Our study showed that combined EUS-FNA + FNB had higher diagnostic yield compared to EUS-FNA in SPMs along with less number of needle passes and needles required. Further prospective studies are needed to validate these findings and cost-effectiveness of this strategy.

Management of Difficult Choledocholithiasis.

Over 30% of all endoscopic retrograde cholangiography procedures in the US are associated with biliary stone extraction, and over 10-15% of these cases are noted to be complex or difficult. The aim of this review is to define the characteristics of difficult common bile duct stones and provide an algorithmic therapeutic approach to these difficult cases. We describe additional special clinical circumstances in which difficult biliary stones are identified and provide additional management strategies to aid endoscopic stone extraction efforts.

Prevalence, Determinants and Clinical Significance of Cardiac Troponin-I Elevation among Individuals with Hypertensive Emergency: A Prospective Observational Study.

Introduction: Cardiac troponin-I (cTnI) elevation release during hypertensive emergencies (HEs) is a frequent epiphenomenon that may tangle management of individuals being treated for HEs. The primary objective of this study was to determine the prevalence, determinants, and clinical significance of cTnI elevation and secondary objective was to find out the prognostic significance of cTnI elevation in patients admitted for HE in the emergency department (ED) of a tertiary care hospital. Methodology: The investigator has employed the quantitative research approach with a prospective observational descriptive design. The population of this study comprised of 205 adults, which included both males and females, who were more than or 18 years of age. The subjects were selected by non-probability purposive sampling technique. The study was conducted from August 2015 to December 2016 (16 months). Ethical permission was obtained from the Institutional Ethics Committee (IEC), Max Super Speciality Hospital, Saket, New Delhi and well-informed written consents were taken from the subjects. The analysis of data was done with the help of SPSS, version 17.0. Results: Out of 205 patients in the study, cTnI elevation was found in 102 patients (49.8%). Moreover, there was increased duration of stay in the hospital in patient with elevated cTnI level with mean duration stay 1.55 ± 0.82 (p <0.001). In addition, cTnI elevation was associated with increased mortality, 11 out of 102 in an elevated cTnI group (10.8%) with p <0.002. Conclusion: It was found that cTnI elevation in individuals affected by various clinical factors. The authors highlighted a high frequency of mortality among the individuals presented with HE with elevated cTnI level, whereas the presence of cTnI was associated with greater odds of death. How to cite this article: Gupta K, Kiran M, Chhabra S, Mehta M, Kumar N. Prevalence, Determinants and Clinical Significance of Cardiac Troponin-I Elevation among Individuals with Hypertensive Emergency: A Prospective Observational Study. Indian J Crit Care Med 2022;26(7):786-790.

Deep transcriptomic study reveals the role of cell wall biosynthesis and organization networks in the developing shell of peanut pod.

BACKGROUND: Peanut (Arachis hypogaea L.) belongs to an exceptional group of legume plants, wherein the flowers are produced aerially, but the pods develop under the ground. In such a unique environment, the pod's outer shell plays a vital role as a barrier against mechanical damage and soilborne pathogens. Recent studies have reported the uniqueness and importance of gene expression patterns that accompany peanut pods' biogenesis. These studies focused on biogenesis and pod development during the early stages, but the late developmental stages and disease resistance aspects still have gaps. To extend this information, we analyzed the transcriptome generated from four pod developmental stages of two genotypes, Hanoch (Virginia-type) and IGC53 (Peruvian-type), which differs significantly in their pod shell characteristics and pathogen resistance. RESULTS: The transcriptome study revealed a significant reprogramming of the number and nature of differentially expressed (DE) genes during shell development. Generally, the numbers of DE genes were higher in IGC53 than in Hanoch, and the R5-R6 transition was the most dynamic in terms of transcriptomic changes. Genes related to cell wall biosynthesis, modification and transcription factors (TFs) dominated these changes therefore, we focused on their differential, temporal and spatial expression patterns. Analysis of the cellulose synthase superfamily identified specific Cellulose synthase (CesAs) and Cellulose synthase-like (Csl) genes and their coordinated interplay with other cell wall-related genes during the peanut shell development was demonstrated. TFs were also identified as being involved in the shell development process, and their pattern of expression differed in the two peanut genotypes. The shell component analysis showed that overall crude fiber, cellulose, lignin, hemicelluloses and dry matter increased with shell development, whereas K, N, protein, and ash content decreased. Genotype IGC53 contained a higher level of crude fiber, cellulose, NDF, ADF, K, ash, and dry matter percentage, while Hanoch had higher protein and nitrogen content. CONCLUSIONS: The comparative transcriptome analysis identified differentially expressed genes, enriched processes, and molecular processes like cell wall biosynthesis/modifications, carbohydrate metabolic process, signaling, transcription factors, transport, stress, and lignin biosynthesis during the peanut shell development between two contrasting genotypes. TFs and other genes like chitinases were also enriched in peanut shells known for pathogen resistance against soilborne major pathogens causing pod wart disease and pod damages. This study will shed new light on the biological processes involved with underground pod development in an important legume crop.

DJMol: An open-source modeling platform for computational chemistry and materials science with a Python interpreter.

We present a modular and extendable software suite, DJMol, for performing molecular simulations and it is demonstrated with DFTB+, Siesta, Atomic Simulation Environment, and OpenMD codes. It supports many of the standard features of an integrated development environment and consists of a structure builder and viewer, which could be connected with these electronic structure codes along with a set of data analyzers. This program comprises Java and Python modules and its libraries to carry out a different set of modeling tasks in materials science and chemistry. By adopting a Python interpreter into the software, a range of scriptable Python codes, such as Pymatgen can be incorporated into this programmable modeling platform. DJMol, through its common application programming interface (API), supports multiple modeling codes in the backend and several post-processing tools. It benefits an experienced user by increasing efficiency, while a nonexpert user by easy to use API.

The SARS-CoV-2 Spike protein disrupts human cardiac pericytes function through CD147 receptor-mediated signalling: a potential non-infective mechanism of COVID-19 microvascular disease.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a broad range of clinical responses including prominent microvascular damage. The capacity of SARS-CoV-2 to infect vascular cells is still debated. Additionally, the SARS-CoV-2 Spike (S) protein may act as a ligand to induce non-infective cellular stress. We tested this hypothesis in pericytes (PCs), which are reportedly reduced in the heart of patients with severe coronavirus disease-2019 (COVID-19). Here we newly show that the in vitro exposure of primary human cardiac PCs to the SARS-CoV-2 wildtype strain or the α and δ variants caused rare infection events. Exposure to the recombinant S protein alone elicited signalling and functional alterations, including: (1) increased migration, (2) reduced ability to support endothelial cell (EC) network formation on Matrigel, (3) secretion of pro-inflammatory molecules typically involved in the cytokine storm, and (4) production of pro-apoptotic factors causing EC death. Next, adopting a blocking strategy against the S protein receptors angiotensin-converting enzyme 2 (ACE2) and CD147, we discovered that the S protein stimulates the phosphorylation/activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) through the CD147 receptor, but not ACE2, in PCs. The neutralisation of CD147, either using a blocking antibody or mRNA silencing, reduced ERK1/2 activation, and rescued PC function in the presence of the S protein. Immunoreactive S protein was detected in the peripheral blood of infected patients. In conclusion, our findings suggest that the S protein may prompt PC dysfunction, potentially contributing to microvascular injury. This mechanism may have clinical and therapeutic implications.

Drug-induced liver injury associated with antiseizure medications from the FDA Adverse Event Reporting System (FAERS).

PURPOSE: Treatment with antiseizure medications (ASMs) confers a risk of drug-induced liver injury (DILI), especially for older ASMs. We sought to quantify recent reports of DILI attributed to both older and newer generation ASMs and survey newly marketed ASMs for hepatotoxicity in a large post-marketing database. METHODS: We queried over 2.6 million adverse event reports made to the FDA Adverse Event Reporting System (FAERS) database between July 1, 2018 and March 31, 2020 for DILI due to ASMs commonly used in clinical practice. Patient characteristics and outcomes were assessed. We calculated the reporting odds ratio (ROR) of DILI for each individual ASM versus all non-ASM reports. RESULTS: A total of 2175 DILI cases were attributed to an ASM during the study period. 97.2% of these were designated as serious reactions, which include death, hospitalization, disability, and other life-threatening outcomes. A number of older and newer generation ASMs were associated with DILI, specifically: carbamazepine (ROR 2.92), phenobarbital (ROR 2.91), oxcarbazepine (ROR 2.58), phenytoin (ROR 2.40), valproate (ROR 2.22), lamotrigine (ROR 2.06), clobazam (ROR 1.67), levetiracetam (ROR 1.56), and diazepam (ROR 1.53). However, increased odds of DILI were not seen with zonisamide, perampanel, stiripentol, lacosamide, clonazepam, pregabalin, felbamate, eslicarbazepine, cannabidiol, topiramate, gabapentin, ethosuximide, brivaracetam, or primidone. Vigabatrin, tiagabine, and rufinamide all had zero reports of DILI. CONCLUSIONS: The majority of newer generation ASMs were not significantly associated with DILI. Future studies utilizing FAERS in conjunction with other data sources will be critical for the ongoing surveillance of DILI, particularly as newly marketed ASMs continue to enter into widespread clinical use.

Predictors of do-not-resuscitate order utilization in decompensated cirrhosis hospitalized patients: A nationwide inpatient cohort study.

INTRODUCTION AND OBJECTIVES: Decompensated cirrhosis carries high inpatient morbidity and mortality. Consequently, advance care planning is an integral aspect of medical care in this patient population. Our study aims to identify do-not-resuscitate (DNR) order utilization and demographic disparities in decompensated cirrhosis patients. PATIENTS OR MATERIALS AND METHODS: Nationwide Inpatient Sample was used to extract the cohort of patients from January 1st, 2016 to December 31st, 2017, based on the most comprehensive and recent data. The first cohort included hospitalized patients with decompensated cirrhosis. The second cohort included patients with decompensated cirrhosis with at least one contraindication for liver transplantation. RESULTS: A cohort of 585,859 decompensated cirrhosis patients was utilized. DNR orders were present in 14.2% of hospitalized patients. DNR utilization rate among patients with relative contraindication for liver transplantation was 15.0%. After adjusting for co-morbid conditions, disease severity, and inpatient mortality, African-American and Hispanic patient populations had significantly lower DNR utilization rates. There were regional, and hospital-level differences noted. Moreover, advanced age, advanced stage of decompensated cirrhosis, inpatient mortality, and relative contraindications for liver transplantation (metastatic neoplasms, dementia, alcohol misuse, severe cardiopulmonary disease, medical non-adherence) were independently associated with increased DNR utilization rates. CONCLUSIONS: The rate of DNR utilization in patients with relative contraindications for liver transplantation was similar to patients without any relative contraindications. Moreover, there were significant demographic and hospital-level predictors of DNR utilization. This information can guide resource allocation in educating patients and their families regarding prognosis and outcome expectations.

Genomic characterization of ZIP genes in pigeonpea (CcZIP) and their expression analysis among the genotypes with contrasting host response to pod borer.

Zinc (Zn) is a vital micronutrient from the perspective of biofortification and biotic stress endurance in pigeonpea. The ZIP transporters with domain (Pfam: PF02535) regulate uptake and transport of metal ions, including Zn, in consonance with plant metal homeostasis. Genome-wide analysis in pigeonpea identified 19 non-redundant members of ZIP family (CcZIP) that were analyzed for gene structure, conserved motifs and homology besides other structural and biochemical parameters. Intra-specific as well as the inter-specific phylogenetic relationships of these 19 CcZIPs were elucidated by comparison with ZIP proteins of Arabidopsis thaliana, Medicago truncatula, Phaseolus vulgaris and Glycine max. In addition to gene structure, the cis-regulatory elements (CREs) in the promoter region were also identified. It revealed several stress responsive CREs that might be regulatory for differential expression of CcZIP proteins. Expression analysis showed that both CcZIP3 and CcZIP15, having zinc deficiency responsive element, up-regulated in the reproductive leaf tissues and down-regulated in matured green pods of the pod borer resistant genotypes with higher zinc content. Alternately, the expression of CcZIP6 and CcZIP13 was higher in matured green pods than reproductive leaves of the resistant genotypes. These findings on differential expression indicate the possible role of these CcZIPs on the mobilization of Zn from leaves to pods, phloem loading and unloading, and higher accumulation of seed zinc in pod borer resistant genotypes used in this study. Further functional characterization of CcZIP genes could shed light on their role in bio-fortification and genetic improvement to inhibit the pod borer herbivory in pigeonpea. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01111-1.

Molecular Simulations suggest Vitamins, Retinoids and Steroids as Ligands of the Free Fatty Acid Pocket of the SARS-CoV-2 Spike Protein*.

We investigate binding of linoleate and other potential ligands to the recently discovered fatty acid binding site in the SARS-CoV-2 spike protein, using docking and molecular dynamics simulations. Simulations suggest that linoleate and dexamethasone stabilize the locked spike conformation, thus reducing the opportunity for ACE2 interaction. In contrast, cholesterol may expose the receptor-binding domain by destabilizing the closed structure, preferentially binding to a different site in the hinge region of the open structure. We docked a library of FDA-approved drugs to the fatty acid site using an approach that reproduces the structure of the linoleate complex. Docking identifies steroids (including dexamethasone and vitamin D); retinoids (some known to be active in vitro, and vitamin A); and vitamin K as potential ligands that may stabilize the closed conformation. The SARS-CoV-2 spike fatty acid site may bind a diverse array of ligands, including dietary components, and therefore provides a promising target for therapeutics or prophylaxis.

Challenges in developing urban flood resilience in India.

This note presents a short commentary on the recurring urban flooding in India and the challenges the country faces in implementing sustainable drainage practices and developing urban flood resilience. The major incidences of heavy rainfall and the consequent urban flooding and their causes are briefly described. The recent flood disaster management and mitigation measures carried out by the Central, State and Local Government are highlighted. The flood resilience initiatives and challenges are then discussed. The effects of climate change on increased rainfall in shorter durations are now being felt in many Indian cities. The planners are now taking cognizance of this and future flood resilience measures are being planned to incorporate an integrated approach to stormwater management by recommending best management practices in the stormwater manuals. However, a lot still needs to be done to make the cities in India flood resilient. This article is part of the theme issue 'Urban flood resilience'.

Sequential Digestion with Trypsin and Elastase in Cross-Linking Mass Spectrometry.

Cross-linking mass spectrometry has become an important approach for studying protein structures and protein-protein interactions. The amino acid compositions of some protein regions impede the detection of cross-linked residues, although it would yield invaluable information for protein modeling. Here, we report on a sequential-digestion strategy with trypsin and elastase to penetrate regions with a low density of trypsin-cleavage sites. We exploited intrinsic substrate-recognition properties of elastase to specifically target larger tryptic peptides. Our application of this protocol to the TAF4-12 complex allowed us to identify cross-links in previously inaccessible regions.

Genetic code expansion for multiprotein complex engineering.

We present a baculovirus-based protein engineering method that enables site-specific introduction of unique functionalities in a eukaryotic protein complex recombinantly produced in insect cells. We demonstrate the versatility of this efficient and robust protein production platform, 'MultiBacTAG', (i) for the fluorescent labeling of target proteins and biologics using click chemistries, (ii) for glycoengineering of antibodies, and (iii) for structure-function studies of novel eukaryotic complexes using single-molecule Förster resonance energy transfer as well as site-specific crosslinking strategies.