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Found 36319 matches. Displaying 81-90
Yang L, Chan AKN, Miyashita K, Delaney CD, Wang X, Li HZ, Pokharel SP, Li S, Li ML, Xu XB, Lu W, Liu Q, Mattson N, Chen KY, Wang JH, Yuan YC, Horne D, Rosen ST, Soto-Feliciano Y, Feng ZH, Hoshii T, Xiao G, Muschen M, Chen JJ, Armstrong SA, Chen CW
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High-resolution characterization of gene function using single-cell CRISPR tiling screen

NATURE COMMUNICATIONS 2021 JUL 1; 12(1):? Article 4063
Identification of novel functional domains and characterization of detailed regulatory mechanisms in cancer-driving genes is critical for advanced cancer therapy. To date, CRISPR gene editing has primarily been applied to defining the role of individual genes. Recently, high-density mutagenesis via CRISPR tiling of gene-coding exons has been demonstrated to identify functional regions in genes. Furthermore, breakthroughs in combining CRISPR library screens with single-cell droplet RNA sequencing (sc-RNAseq) platforms have revealed the capacity to monitor gene expression changes upon genetic perturbations at single-cell resolution. Here, we present "sc-Tiling," which integrates a CRISPR gene-tiling screen with single-cell transcriptomic and protein structural analyses. Distinct from other reported single-cell CRISPR screens focused on observing gene function and gene-to-gene/enhancer-to-gene regulation, sc-Tiling enables the capacity to identify regulatory mechanisms within a gene-coding region that dictate gene activity and therapeutic response. Identifying functional domains and genetic regulatory mechanisms is essential for developing new therapies. Here the authors present sc-Tiling, single-cell high-density CRISPR tiling screening for functional domain characterization.
Sten TH, Li RF, Otopalik A, Ruta V
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Sexual arousal gates visual processing during Drosophila courtship

NATURE 2021 JUL 22; 595(7868):549-553
Long-lasting internal arousal states motivate and pattern ongoing behaviour, enabling the temporary emergence of innate behavioural programs that serve the needs of an animal, such as fighting, feeding, and mating. However, how internal states shape sensory processing or behaviour remains unclear. In Drosophila, male flies perform a lengthy and elaborate courtship ritual that is triggered by the activation of sexually dimorphic P1 neurons(1-5), during which they faithfully follow and sing to a female(6,7). Here, by recording from males as they court a virtual 'female', we gain insight into how the salience of visual cues is transformed by a male's internal arousal state to give rise to persistent courtship pursuit. The gain of LC10a visual projection neurons is selectively increased during courtship, enhancing their sensitivity to moving targets. A concise network model indicates that visual signalling through the LC10a circuit, once amplified by P1-mediated arousal, almost fully specifies a male's tracking of a female. Furthermore, P1 neuron activity correlates with ongoing fluctuations in the intensity of a male's pursuit to continuously tune the gain of the LC10a pathway. Together, these results reveal how a male's internal state can dynamically modulate the propagation of visual signals through a high-fidelity visuomotor circuit to guide his moment-to-moment performance of courtship.
Blachere NE, Hacisuleyman E, Darnell RB
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Vaccine Breakthrough Infections with SARS-CoV-2 Variants REPLY

NEW ENGLAND JOURNAL OF MEDICINE 2021 JUL 8; 385(2):?
Lavin RC, Johnson C, Ahn YM, Kremiller KM, Sherwood M, Patel JS, Pan Y, Russo R, MacGilvary NJ, Giacalone D, Kevorkian YL, Zimmerman MD, Glickman JF, Freundlich JS, Tan SM
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Targeting Mycobacterium tuberculosis response to environmental cues for the development of effective antitubercular drugs

PLOS BIOLOGY 2021 JUL; 19(7):? Article e3001355
Sensing and response to environmental cues, such as pH and chloride (Cl-), is critical in enabling Mycobacterium tuberculosis (Mtb) colonization of its host. Utilizing a fluorescent reporter Mtb strain in a chemical screen, we have identified compounds that dysregulate Mtb response to high Cl- levels, with a subset of the hits also inhibiting Mtb growth in host macrophages. Structure-activity relationship studies on the hit compound "C6," or 2-(4-((2-(ethylthio)pyrimidin-5-yl)methyl)piperazin-1-yl)benzo[d]oxazole, demonstrated a correlation between compound perturbation of Mtb Cl- response and inhibition of bacterial growth in macrophages. C6 accumulated in both bacterial and host cells, and inhibited Mtb growth in cholesterol media, but not in rich media. Subsequent examination of the Cl- response of Mtb revealed an intriguing link with bacterial growth in cholesterol, with increased transcription of several Cl--responsive genes in the simultaneous presence of cholesterol and high external Cl- concentration, versus transcript levels observed during exposure to high external Cl- concentration alone. Strikingly, oral administration of C6 was able to inhibit Mtb growth in vivo in a C3HeB/FeJ murine infection model. Our work illustrates how Mtb response to environmental cues can intersect with its metabolism and be exploited in antitubercular drug discovery.
Felix S, Handem S, Nunes S, Paulo AC, Candeias C, Valente C, Simoes AS, Almeida ST, Tavares DA, Brito-Avo A, de Lencastre H, Sa-Leao R
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Impact of private use of the 13-valent pneumococcal conjugate vaccine (PCV13) on pneumococcal carriage among Portuguese children living in urban and rural regions

VACCINE 2021 JUL 22; 39(32):4524-4533
In Portugal, the 13-valent pneumococcal conjugate vaccine (PCV13) was commercially available between 2010 and 2015, following a decade of private use of PCV7. We evaluated changes on serotype distribution and antimicrobial susceptibility of pneumococci carried by children living in two regions of Portugal (one urban and one rural). Three epidemiological periods were defined: pre-PCV13 (2009-2010), early-PCV13 (2011-2012), and late-PCV13 (2015-2016). Nasopharyngeal samples (n = 4,232) were obtained from children 0-6 years old attending day-care centers. Private use of PCVs was very high in both regions (>75%). Pneumococcal carriage remained stable and high over time (62.1%, 62.4% and 61.6% (p = 0.909) in the urban region; and 59.8%, 62.8%, 59.5% (p = 0.543) in the rural region). Carriage of PCV7 serotypes remained low (5.3%, 7.8% and 4.3% in the urban region; and 2.5%, 3.7% and 4.8% in the rural region). Carriage of PCV13 serotypes not targeted by PCV7 decreased in both the urban (16.4%, 7.3%, and 1.6%; p < 0.001) and rural regions (13.2%, 7.8%, and 1.9%; p < 0.001). This decline was mostly attributable to serotype 19A (14.1%, 4.4% and 1.3% in the urban region; and 11.1%, 3.6% and 0.8% in the rural region, both p < 0.001). Serotype 3 declined over time in the urban region (10.1%, 4.4%, 0.8%; p < 0.001) and had no obvious trend in the rural region (4.2%, 6.7%, 2.4%; p = 0.505). Serotype 6C decreased in both regions while serotypes 11D, 15A/B/C, 16F, 21, 22F, 23A/B, 24F, 35F, and NT were the most prevalent in the late-PCV13 period. Intermediate resistance to penicillin and non-susceptibility to erythromycin decreased significantly in both regions (19.5%, 13.3%, and 9.3%; and 25.4%, 25.9%, and 13.4%; both p < 0.001, respectively in the urban region; and 12.4%, 11.1%, and 2.8% (p < 0.001); and 15.3%, 14.7%, and 9.2% (p = 0.037), respectively, in the rural region). In conclusion, private use of PCV13 led to significant changes on the pneumococcal population carried by children in Portugal. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/).
Huber T, Goldman O, Epstein AE, Stella G, Sakmar TP
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Principles and practice for SARS-CoV-2 decontamination of N95 masks with UV-C

BIOPHYSICAL JOURNAL 2021 JUL 20; 120(14):2927-2942
A mainstay of personal protective equipment during the coronavirus disease 2019 pandemic is the N95 filtering facepiece respirator. N95 respirators are commonly used to protect healthcare workers from respiratory pathogens, including the novel coronavirus severe acute respiratory syndrome coronavirus 2, and are increasingly employed by other frontline workers and the general public. Under routine circumstances, these masks are disposable, single-use items, but extended use and reuse practices have been broadly enacted to alleviate critical supply shortages during the coronavirus disease 2019 pandemic. Although extended-time single use presents a low risk of pathogen transfer, repeated donning and doffing of potentially contaminated masks presents increased risk of pathogen transfer. Therefore, efficient and safe decontamination methods for N95 masks are needed to reduce the risk of reuse and mitigate local supply shortages. Here, we review the available literature concerning use of germicidal ultraviolet-C (UV-C) light to decontaminate N95 masks. We propose a practical method for repeated point-of-use decontamination using commercially available UV-C cross-linker boxes from molecular biology laboratories to expose each side of the mask to 800-1200 mJ/cm(2) of UV-C. We measure the dose that penetrated to the interior of the respirators and model the potential germicidal action on coronaviruses. Our experimental results, in combination with modeled data, suggest that such a UV-C treatment cycle should induce a >3-log-order reduction in viral bioburden on the surface of the respirators and a 2-log-order reduction throughout the interior. We find that a dose 50-fold greater does not impair filtration or fit of 3M 8210 N95 masks, indicating that decontamination can be performed repeatedly. As such, UV-C germicidal irradiation is a practical strategy for small-scale point-of-use decontamination of N95s.
Brivanlou AH, Rivron N, Gleicher N
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How will our understanding of human development evolve over the next 10 years

NATURE COMMUNICATIONS 2021 JUL 29; 12(1):? Article 4614
In the next 10 years, the continued exploration of human embryology holds promise to revolutionize regenerative and reproductive medicine with important societal consequences. In this Comment we speculate on the evolution of recent advances made and describe emerging technologies for basic research, their potential clinical applications, and, importantly, the ethical frameworks in which they must be considered.
Sacramento CQ, Fintelman-Rodrigues N, Temerozo JR, Da Silva ADD, Dias SDG, da Silva CD, Ferreira AC, Mattos M, Pao CRR, de Freitas CS, Soares VC, Hoelz LVB, Fernandes TVA, Branco FSC, Bastos MM, Boechat N, Saraiva FB, Ferreira MA, Jockusch S, Wang XT, Tao CAJ, Chien MC, Xie W, Patel D, Garzia A, Tuschl T, Russo JJ, Rajoli RKR, Pedrosa CSG, Vitoria G, Souza LRQ, Goto-Silva L, Guimaraes MZ, Rehen SK, Owen A, Bozza FA, Bou-Habib DC, Ju JY, Bozza PT, Souza TML
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In vitro antiviral activity of the anti-HCV drugs daclatasvir and sofosbuvir against SARS-CoV-2, the aetiological agent of COVID-19

JOURNAL OF ANTIMICROBIAL CHEMOTHERAPY 2021 JUL; 76(7):1874-1885
Background: Current approaches of drug repurposing against COVID-19 have not proven overwhelmingly successful and the SARS-CoV-2 pandemic continues to cause major global mortality. SARS-CoV-2 nsp12, its RNA polymerase, shares homology in the nucleotide uptake channel with the HCV orthologue enzyme NS5B. Besides, HCV enzyme NS5A has pleiotropic activities, such as RNA binding, that are shared with various SARS-CoV-2 proteins. Thus, anti-HCV NS5B and NS5A inhibitors, Like sofosbuvir and daclatasvir, respectively, could be endowed with anti-SARS-CoV-2 activity. Methods: SARS-CoV-2-infected Vero cells, HuH-7 cells, Calu-3 cells, neural stem cells and monocytes were used to investigate the effects of daclatasvir and sofosbuvir. In silico and cell-free based assays were performed with SARS-CoV-2 RNA and nsp12 to better comprehend the mechanism of inhibition of the investigated compounds. A physiologically based pharmacokinetic model was generated to estimate daclatasvir's dose and schedule to maximize the probability of success for COVID-19. Results: Daclatasvir inhibited SARS-CoV-2 replication in Vero, HuH-7 and Calu-3 cells, with potencies of 0.8, 0.6 and 1.1 mu M, respectively. Although Less potent than daclatasvir, sofosbuvir alone and combined with daclatasvir inhibited replication in Calu-3 cells. Sofosbuvir and daclatasvir prevented virus-induced neuronal apoptosis and release of cytokine storm-related inflammatory mediators, respectively. Sofosbuvir inhibited RNA synthesis by chain termination and daclatasvir targeted the folding of secondary RNA structures in the SARS-CoV-2 genome. Concentrations required for partial daclatasvir in vitro activity are achieved in plasma at C-max after administration of the approved dose to humans. Conclusions: Daclatasvir, alone or in combination with sofosbuvir, at higher doses than used against HCV, may be further fostered as an anti-COVID-19 therapy.
Diab NS, Barish S, Dong WL, Zhao SJ, Allington G, Yu XB, Kahle KT, Brueckner M, Jin SC
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Molecular Genetics and Complex Inheritance of Congenital Heart Disease

GENES 2021 JUL; 12(7):? Article 1020
Congenital heart disease (CHD) is the most common congenital malformation and the leading cause of mortality therein. Genetic etiologies contribute to an estimated 90% of CHD cases, but so far, a molecular diagnosis remains unsolved in up to 55% of patients. Copy number variations and aneuploidy account for similar to 23% of cases overall, and high-throughput genomic technologies have revealed additional types of genetic variation in CHD. The first CHD risk genotypes identified through high-throughput sequencing were de novo mutations, many of which occur in chromatin modifying genes. Murine models of cardiogenesis further support the damaging nature of chromatin modifying CHD mutations. Transmitted mutations have also been identified through sequencing of population scale CHD cohorts, and many transmitted mutations are enriched in cilia genes and Notch or VEGF pathway genes. While we have come a long way in identifying the causes of CHD, more work is required to end the diagnostic odyssey for all CHD families. Complex genetic explanations of CHD are emerging but will require increasingly sophisticated analysis strategies applied to very large CHD cohorts before they can come to fruition in providing molecular diagnoses to genetically unsolved patients. In this review, we discuss the genetic architecture of CHD and biological pathways involved in its pathogenesis.
Borah S, Thaller DJ, Hakhverdyan Z, Rodriguez EC, Isenhour AW, Rout MP, King MC, Lusk CP
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Heh2/Man1 may be an evolutionarily conserved sensor of NPC assembly state

MOLECULAR BIOLOGY OF THE CELL 2021 JUL 15; 32(15):1359-1373
Integral membrane proteins of the Lap2-emerin-MAN1 (LEM) family have emerged as important components of the inner nuclear membrane (INM) required for the functional and physical integrity of the nuclear envelope. However, like many INM proteins, there is limited understanding of the biochemical interaction networks that enable LEM protein function. Here, we show that Heh2/Man1 can interact with major scaffold components of the nuclear pore complex (NPC), specifically the inner ring complex (IRC), in evolutionarily distant yeasts. Although an N-terminal domain is required for Heh2 targeting to the INM, we demonstrate that more stable interactions with the NPC are mediated by a C-terminal winged helix (WH) domain, thus decoupling INM targeting and NPC binding. Inhibiting Heh2's interactions with the NPC by deletion of the Heh2 WH domain leads to NPC clustering. Interestingly, Heh2's association with NPCs can also be disrupted by knocking out several outer ring nucleoporins. Thus, Heh2's interaction with NPCs depends on the structural integrity of both major NPC scaffold complexes. We propose a model in which Heh2 acts as a sensor of NPC assembly state, which may be important for NPC quality control mechanisms and the segregation of NPCs during cell division.