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1.
Nucleic Acids Res ; 2024 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-39380503

RESUMO

SARS-CoV-2 nucleocapsid (N) protein is a structural component of the virus with essential roles in the replication and packaging of the viral RNA genome. The N protein is also an important target of COVID-19 antigen tests and a promising vaccine candidate along with the spike protein. Here, we report a compact stem-loop DNA aptamer that binds tightly to the N-terminal RNA-binding domain of SARS-CoV-2 N protein. Crystallographic analysis shows that a hexanucleotide DNA motif (5'-TCGGAT-3') of the aptamer fits into a positively charged concave surface of N-NTD and engages essential RNA-binding residues including Tyr109, which mediates a sequence-specific interaction in a uracil-binding pocket. Avid binding of the DNA aptamer allows isolation and sensitive detection of full-length N protein from crude cell lysates, demonstrating its selectivity and utility in biochemical applications. We further designed a chemically modified DNA aptamer and used it as a probe to examine the interaction of N-NTD with various RNA motifs, which revealed a strong preference for uridine-rich sequences. Our studies provide a high-affinity chemical probe for the SARS-CoV-2 N protein RNA-binding domain, which may be useful for diagnostic applications and investigating novel antiviral agents.

2.
bioRxiv ; 2024 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-39386696

RESUMO

Liquid-liquid phase separation of proteins and nucleic acids into condensate phases is a versatile mechanism for ensuring compartmentalization of cellular biochemistry. RNA molecules play critical roles in these condensates, particularly in transcriptional regulation and stress responses, exhibiting a wide range of thermodynamic and dynamic behaviors. However, deciphering the molecular grammar that governs the stability and dynamics of protein-RNA condensates remains challenging due to the multicomponent and heterogeneous nature of these biomolecular mixtures. In this study, we employ atomistic simulations of twenty distinct mixtures containing minimal RNA and peptide fragments to dissect the phase-separating affinities of all twenty amino acids in the presence of RNA. Our findings elucidate chemically specific interactions, hydration profiles, and ionic effects that synergistically promote or suppress protein-RNA phase separation. We map a ternary phase diagram of interactions, identifying four distinct groups of residues that promote, maintain, suppress, or disrupt protein-RNA clusters.

3.
Methods Enzymol ; 705: 311-345, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39389668

RESUMO

In recent years, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become ever more apparent. This growing awareness and lack of inhibitory drugs has created a distinct need for biochemical tools that can be used to identify and characterize potential inhibitors of this family of enzymes. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay. The RADD assay provides a rapid, real-time fluorescence readout of APOBEC3 DNA deamination and serves as a crucial addition to the existing APOBEC3 biochemical and cellular toolkit. This method improves upon contemporary DNA deamination assays by offering a more rapid and quantifiable readout as well as providing a platform that is readily adaptable to a high-throughput format for inhibitor discovery. In this chapter we provide a detailed guide for the usage of the RADD assay for the characterization of APOBEC3 enzymes and potential inhibitors.


Assuntos
DNA , Transferência Ressonante de Energia de Fluorescência , Humanos , Transferência Ressonante de Energia de Fluorescência/métodos , DNA/metabolismo , Desaminação , Citidina Desaminase/metabolismo , Citidina Desaminase/genética , Ensaios Enzimáticos/métodos , Inibidores Enzimáticos/farmacologia , Desaminases APOBEC/metabolismo
4.
J Phys Chem Lett ; : 10811-10817, 2024 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-39432826

RESUMO

Liquid-liquid phase separation of proteins and nucleic acids into condensate phases is a versatile mechanism for ensuring the compartmentalization of cellular biochemistry. RNA molecules play critical roles in these condensates, particularly in transcriptional regulation and stress responses, exhibiting a wide range of thermodynamic and dynamic behaviors. However, deciphering the molecular grammar that governs the stability and dynamics of protein-RNA condensates remains challenging due to the multicomponent and heterogeneous nature of condensates. In this study, we employ atomistic simulations of 20 distinct mixtures containing minimal RNA and peptide fragments which allows us to dissect the phase-separating affinities of all 20 amino acids in the presence of RNA. Our findings elucidate chemically specific interactions, hydration profiles, and ionic effects that synergistically promote or suppress protein-RNA phase separation. We map a ternary phase diagram of interactions, identifying four distinct groups of residues that promote, maintain, suppress, and disrupt protein-RNA clusters.

5.
Adv Sci (Weinh) ; 11(39): e2403690, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39159128

RESUMO

The generation of pressure perturbations in matter stimulated by pulsed light is a method widely recognized as the photoacoustic or light-induced thermoelastic effect. In a series of psychophysical experiments, the robustness of the tactile perception generated with a variety of light sources is examined: a diverging pulsed laser used for photoacoustic tomography optical parameter oscillation (OPO), a miniature diode laser (MDL), and a commercial digital light processing (DLP) projector. It is demonstrated that participants can accurately detect, categorically describe the sensations, and discern the direction of pulsed light travel. High detection accuracy is reported as follows: (d' = 4.95 (OPO); d' = 2.78 (modulated MDL); d' = 2.99 (DLP)) of the stimulus on glabrous skin coated with a thin layer of dye absorber. For all light sources, the predominant sensation is felt as vibration at the distal phalanx (i.e., fingertip, 55.21-57.29%) and the proximal phalanx (41.67-44.79%). At the fingertip, thermal sensations are perceived less frequently than mechanical ones. Moreover, these haptic effects are preserved under a wide range of pulse widths, spot sizes, optical energies, and wavelengths of the light sources. This form of sensory stimulation demonstrates a generalizable non-contact, non-optogenetic, in situ activation of the mechanosensory system.

6.
Plant J ; 2024 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-39121182

RESUMO

The bilateral-to-radial symmetry transition occurring during the development of the Arabidopsis thaliana female reproductive organ (gynoecium) is a crucial biological process linked to plant fertilization and seed production. Despite its significance, the cellular mechanisms governing the establishment and breaking of radial symmetry at the gynoecium apex (style) remain unknown. To fill this gap, we employed quantitative confocal imaging coupled with MorphoGraphX analysis, in vivo and in vitro transcriptional experiments, and genetic analysis encompassing mutants in two bHLH transcription factors necessary and sufficient to promote transition to radial symmetry, SPATULA (SPT) and INDEHISCENT (IND). Here, we show that defects in style morphogenesis correlate with defects in cell-division orientation and rate. We showed that the SPT-mediated accumulation of auxin in the medial-apical cells undergoing symmetry transition is required to maintain cell-division-oriented perpendicular to the direction of organ growth (anticlinal, transversal cell division). In addition, SPT and IND promote the expression of specific core cell-cycle regulators, CYCLIN-D1;1 (CYC-D1;1) and CYC-D3;3, to support progression through the G1 phase of the cell cycle. This transcriptional regulation is repressed by auxin, thus forming an incoherent feed-forward loop mechanism. We propose that this mechanism fine-tunes cell division rate and orientation with the morphogenic signal provided by auxin, during patterning of radial symmetry at the style.

7.
Int J Exerc Sci ; 17(4): 819-830, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39050402

RESUMO

Fitness testing is employed by some law enforcement agencies to assure performance in occupational tasks. The aim of this study was to investigate associations between musculoskeletal fitness assessment scores and performance in police occupational tasks. Retrospective data from 106 law enforcement officers who completed five musculoskeletal fitness assessments (vertical jump (VJ), hand grip strength, leg back dynamometer, 1-minute push-ups and sit-ups) and three routine occupational tasks (1.22m fence jump (FJ), 8.5m victim drag (VD) with 101kg and a get-up (GU)) were collected. A standard multiple regression was performed to determine if the results in fitness assessments were predictive of performance in the occupational tasks. Models combining all fitness assessments significantly predicted performance in FJ (F(5,88)=12.228, p<0.001; adjusted R2=0.38), VD (F(5,88)=9.407, p<0.001; adjusted R2=0.31) and GU (F(5,87)=14.319, p<0.001; adjusted R2=0.42). Further analysis of individual predictors highlighted that performance in the VJ test was a significant contributor for all models, uniquely predicting 15% of FJ (p<0.001), 4% of VD (p=0.03) and 8% of GU (p=0.001) performance. Grip strength uniquely contributed 3% to performance in the VD (p=0.05) and performance in the sit-up test contributed 8% to GU performance (p=0.001). Performance in police-specific occupational tasks requires a combination of muscular strength, power, and endurance. These musculoskeletal fitness components should be ideally assessed in recruitment and return-to work practices to ensure officers can safely and optimally perform their occupational requirements.

8.
Geohealth ; 8(7): e2023GH000784, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38962698

RESUMO

Machine learning methods have seen increased application to geospatial environmental problems, such as precipitation nowcasting, haze forecasting, and crop yield prediction. However, many of the machine learning methods applied to mosquito population and disease forecasting do not inherently take into account the underlying spatial structure of the given data. In our work, we apply a spatially aware graph neural network model consisting of GraphSAGE layers to forecast the presence of West Nile virus in Illinois, to aid mosquito surveillance and abatement efforts within the state. More generally, we show that graph neural networks applied to irregularly sampled geospatial data can exceed the performance of a range of baseline methods including logistic regression, XGBoost, and fully-connected neural networks.

9.
Mov Disord ; 39(9): 1610-1618, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38859549

RESUMO

BACKGROUND: Current treatments for Tourette syndrome (TS) and persistent tic disorder (PTD) are often insufficiently effective, inaccessible, and frequently associated with adverse events. Thus, we must continue to develop and test effective, accessible, and safe treatment options. OBJECTIVE: We aimed to conduct a pilot randomized controlled trial (RCT) comparing a novel, videoconference-delivered group mindfulness-based intervention for tics (MBIT) to videoconference-delivered group psychoeducation, relaxation, and supportive therapy (PRST) for adults with TS or PTD. METHODS: Thirty-two adults with TS or PTD were randomly assigned to receive 8 weeks of either MBIT or PRST. Tic severity, tic-related impairment, and global improvement were assessed by a trained, independent evaluator who was masked to treatment condition at baseline (week 0), posttreatment (week 9), 1-month follow-up, and 6-month follow-up. All study procedures were conducted online via secure videoconferencing. RESULTS: Twenty-eight participants began treatment and were included in analyses. MBIT, relative to PRST, was associated with a significantly greater decline in tic severity (d = 0.85) and tic-related impairment (d = 0.99) from baseline to posttreatment. Treatment response was significantly higher in MBIT (69%) than in PRST (13%). Neither treatment resulted in serious adverse effects. The durability of treatment outcomes is also reported and discussed. CONCLUSIONS: The results from this pilot RCT suggest that videoconference-delivered group MBIT may be an efficacious, accessible, and safe intervention for adults with tics. Future research is necessary to confirm these preliminary findings. © 2024 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.


Assuntos
Atenção Plena , Transtornos de Tique , Síndrome de Tourette , Humanos , Masculino , Atenção Plena/métodos , Feminino , Adulto , Projetos Piloto , Resultado do Tratamento , Transtornos de Tique/terapia , Síndrome de Tourette/terapia , Pessoa de Meia-Idade , Comunicação por Videoconferência , Tiques/terapia , Adulto Jovem , Psicoterapia de Grupo/métodos
10.
J Biol Chem ; 300(6): 107410, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38796062

RESUMO

Over the past decade, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become increasingly apparent. This growing awareness has created a need for biochemical tools that can be used to identify and characterize potential inhibitors of this enzyme family. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination assay. This assay offers a single-step set-up and real-time fluorescent read-out, and it is capable of providing insights into enzyme kinetics. The assay also offers a high-sensitivity and easily scalable method for identifying APOBEC3 inhibitors. This assay serves as a crucial addition to the existing APOBEC3 biochemical and cellular toolkit and possesses the versatility to be readily adapted into a high-throughput format for inhibitor discovery.


Assuntos
Citidina Desaminase , DNA , Humanos , Desaminação , Citidina Desaminase/metabolismo , DNA/metabolismo , DNA/química , Cinética , Desaminases APOBEC/metabolismo , Inibidores Enzimáticos/farmacologia
11.
bioRxiv ; 2024 May 12.
Artigo em Inglês | MEDLINE | ID: mdl-38766133

RESUMO

Over the past decade, the connection between APOBEC3 cytosine deaminases and cancer mutagenesis has become increasingly apparent. This growing awareness has created a need for biochemical tools that can be used to identify and characterize potential inhibitors of this enzyme family. In response to this challenge, we have developed a Real-time APOBEC3-mediated DNA Deamination (RADD) assay. This assay offers a single-step set-up and real-time fluorescent read-out, and it is capable of providing insights into enzyme kinetics and also offering a high-sensitivity and easily scalable method for identifying APOBEC3 inhibitors. This assay serves as a crucial addition to the existing APOBEC3 biochemical and cellular toolkit and possesses the versatility to be readily adapted into a high-throughput format for inhibitor discovery.

12.
Biomed Opt Express ; 15(3): 1408-1417, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38495713

RESUMO

Assessing cell viability is important in many fields of research. Current optical methods to assess cell viability typically involve fluorescent dyes, which are often less reliable and have poor permeability in primary tissues. Dynamic optical coherence microscopy (dOCM) is an emerging tool that provides label-free contrast reflecting changes in cellular metabolism. In this work, we compare the live contrast obtained from dOCM to viability dyes, and for the first time to our knowledge, demonstrate that dOCM can distinguish live cells from dead cells in murine syngeneic tumors. We further demonstrate a strong correlation between dOCM live contrast and optical redox ratio by metabolic imaging in primary mouse liver tissue. The dOCM technique opens a new avenue to apply label-free imaging to assess the effects of immuno-oncology agents, targeted therapies, chemotherapy, and cell therapies using live tumor tissues.

13.
Am J Trop Med Hyg ; 110(3): 518-528, 2024 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-38320317

RESUMO

Current modeling practices for environmental and sociological modulated infectious diseases remain inadequate to forecast the risk of outbreak(s) in human populations, partly due to a lack of integration of disciplinary knowledge, limited availability of disease surveillance datasets, and overreliance on compartmental epidemiological modeling methods. Harvesting data knowledge from virus transmission (aerosols) and detection (wastewater) of SARS-CoV-2, a heuristic score-based environmental predictive intelligence system was developed that calculates the risk of COVID-19 in the human population. Seasonal validation of the algorithm was uniquely associated with wastewater surveillance of the virus, providing a lead time of 7-14 days before a county-level outbreak. Using county-scale disease prevalence data from the United States, the algorithm could predict COVID-19 risk with an overall accuracy ranging between 81% and 98%. Similarly, using wastewater surveillance data from Illinois and Maryland, the SARS-CoV-2 detection rate was greater than 80% for 75% of the locations during the same time the risk was predicted to be high. Results suggest the importance of a holistic approach across disciplinary boundaries that can potentially allow anticipatory decision-making policies of saving lives and maximizing the use of available capacity and resources.


Assuntos
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/epidemiologia , Estações do Ano , Águas Residuárias , Vigilância Epidemiológica Baseada em Águas Residuárias , Inteligência
14.
Biophys J ; 123(3): 349-360, 2024 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-38163950

RESUMO

Phase separation of biomolecules underlies the formation and regulation of various membraneless condensates in cells. How condensates function reliably while surrounded by heterogeneous and dynamic mixtures of biomolecular components with specific and nonspecific interactions is yet to be understood. Studying multicomponent biomolecular mixtures with designer peptides has recently become an attractive avenue for learning about physicochemical principles governing cellular condensates. In this work, we employed long-timescale atomistic simulations of multicomponent tripeptide mixtures with all residue substitutions to illuminate the nature of direct and water-mediated interactions in a prototypical cellular condensate environment. We find that peptide mixtures form clusters with inverse hydrophobic order. Most multivalent and charged residues are localized in the cluster's core, with a large fraction of nonaromatic hydrophobic residues remaining on the surface. This inverse hydrophobic order in peptide clusters is partly driven by the expulsion of nonspecifically bound water molecules following peptide cluster growth. The growth of clusters is also accompanied by the formation of increasing numbers of specific water-mediated interactions between polar and charged residues. While the present study focused on the condensation of short peptide motifs, the general findings and analysis techniques should be helpful for future studies on larger peptides and protein condensation.


Assuntos
Peptídeos , Separação de Fases , Peptídeos/química , Proteínas , Água
15.
Anat Sci Educ ; 17(3): 468-482, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38213130

RESUMO

Basic competency in radiological imaging is essential for physicians to identify and manage diseases. An optimal place in which to include imaging in the medical curriculum is during anatomy as students can correlate the 3D anatomy from their body donors with the 2D cross-sectional anatomy. The goal of this project was to enhance first-year medical students' knowledge of cross-sectional imaging in the gross anatomy lab and to investigate whether there are benefits to learning cross sectional imaging via scans from body donors versus living individuals. Student participant performance was evaluated on laboratory practical examinations, CT image questions and spatial anatomical knowledge in the thorax and abdomen sections of gross anatomy. Students learned the cross-sectional imaging during dissections where they accessed the images relevant to their study on Pacsbin, a web-based Digital Imaging and Communication in Medicine viewer, via iPads. Results showed no statistically significant differences in practical examination scores, spatial anatomical knowledge, or identification of anatomical structures on CT image questions between participants who learned from images on body donors versus living individuals. In a questionnaire given at the end of the course, participants cited that the CT images improved their anatomical and imaging knowledge and that they felt better prepared to use imaging software and interpret diagnostic imaging results upon entering clerkships. While there were no differences in academic performance between the groups, positive outcomes regarding student perceptions of anatomical and imaging knowledge and preparedness for use of imaging software were identified in this study.


Assuntos
Anatomia , Educação de Graduação em Medicina , Estudantes de Medicina , Humanos , Imageamento post mortem , Anatomia/educação , Avaliação Educacional/métodos , Currículo , Educação de Graduação em Medicina/métodos , Inquéritos e Questionários
16.
Arthritis Rheumatol ; 2023 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-38073013

RESUMO

OBJECTIVE: This study aimed to identify peripheral and salivary gland (SG) biomarkers of response/resistance to B cell depletion based on the novel concise Composite of Relevant Endpoints for Sjögren Syndrome (cCRESS) and candidate Sjögren Tool for Assessing Response (STAR) composite endpoints. METHODS: Longitudinal analysis of peripheral blood and SG biopsies was performed pre- and post-treatment from the Trial of Anti-B Cell Therapy in Patients With Primary Sjögren Syndrome (TRACTISS) combining flow cytometry immunophenotyping, serum cytokines, and SG bulk RNA sequencing. RESULTS: Rituximab treatment prevented the worsening of SG inflammation observed in the placebo arm, by inhibiting the accumulation of class-switched memory B cells within the SG. Furthermore, rituximab significantly down-regulated genes involved in immune-cell recruitment, lymphoid organization alongside antigen presentation, and T cell co-stimulatory pathways. In the peripheral compartment, rituximab down-regulated immunoglobulins  and auto-antibodies together with pro-inflammatory cytokines and chemokines. Interestingly, patients classified as responders  according to STAR displayed significantly higher baseline levels of C-X-C motif chemokine ligand-13 (CXCL13), interleukin (IL)-22, IL-17A, IL-17F, and tumor necrosis factor-α (TNF-α), whereas a longitudinal analysis of serum T cell-related cytokines showed a selective reduction in both STAR and cCRESS responder patients. Conversely, cCRESS response was better associated with biomarkers of SG immunopathology, with cCRESS-responders showing a significant decrease in SG B cell infiltration and reduced expression of transcriptional gene modules related to T cell costimulation, complement activation, and Fcγ-receptor engagement. Finally, cCRESS and STAR response were associated with a significant improvement in SG exocrine function linked to transcriptional evidence of SG epithelial and metabolic restoration. CONCLUSION: Rituximab modulates both peripheral and SG inflammation, preventing the deterioration of exocrine function with functional and metabolic restoration of the glandular epithelium. Response assessed by newly developed cCRESS and STAR criteria was associated with differential modulation of peripheral and SG biomarkers, emerging as novel tools for patient stratification.

17.
Int J Mol Sci ; 24(24)2023 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-38139430

RESUMO

Type 10 17ß-hydroxysteroid dehydrogenase (17ß-HSD10) is the HSD17B10 gene product playing an appreciable role in cognitive functions. It is the main hub of exercise-upregulated mitochondrial proteins and is involved in a variety of metabolic pathways including neurosteroid metabolism to regulate allopregnanolone homeostasis. Deacetylation of 17ß-HSD10 by sirtuins helps regulate its catalytic activities. 17ß-HSD10 may also play a critical role in the control of mitochondrial structure, morphology and dynamics by acting as a member of the Parkin/PINK1 pathway, and by binding to cyclophilin D to open mitochondrial permeability pore. 17ß-HSD10 also serves as a component of RNase P necessary for mitochondrial tRNA maturation. This dehydrogenase can bind with the Aß peptide thereby enhancing neurotoxicity to brain cells. Even in the absence of Aß, its quantitative and qualitative variations can result in neurodegeneration. Since elevated levels of 17ß-HSD10 were found in brain cells of Alzheimer's disease (AD) patients and mouse AD models, it is considered to be a key factor in AD pathogenesis. Since data underlying Aß-binding-alcohol dehydrogenase (ABAD) were not secured from reported experiments, ABAD appears to be a fabricated alternative term for the HSD17B10 gene product. Results of this study would encourage researchers to solve the question why elevated levels of 17ß-HSD10 are present in brains of AD patients and mouse AD models. Searching specific inhibitors of 17ß-HSD10 may find candidates to reduce senile neurodegeneration and open new approaches for the treatment of AD.


Assuntos
17-Hidroxiesteroide Desidrogenases , Doença de Alzheimer , Animais , Humanos , Camundongos , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Álcool Desidrogenase/metabolismo , Doença de Alzheimer/metabolismo , Encéfalo/metabolismo
18.
PLoS Genet ; 19(11): e1011043, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-38033156

RESUMO

A prominent source of mutation in cancer is single-stranded DNA cytosine deamination by cellular APOBEC3 enzymes, which results in signature C-to-T and C-to-G mutations in TCA and TCT motifs. Although multiple enzymes have been implicated, reports conflict and it is unclear which protein(s) are responsible. Here we report the development of a selectable system to quantify genome mutation and demonstrate its utility by comparing the mutagenic activities of three leading candidates-APOBEC3A, APOBEC3B, and APOBEC3H. The human cell line, HAP1, is engineered to express the thymidine kinase (TK) gene of HSV-1, which confers sensitivity to ganciclovir. Expression of APOBEC3A and APOBEC3B, but not catalytic mutant controls or APOBEC3H, triggers increased frequencies of TK mutation and similar TC-biased cytosine mutation profiles in the selectable TK reporter gene. Whole genome sequences from independent clones enabled an analysis of thousands of single base substitution mutations and extraction of local sequence preferences with APOBEC3A preferring YTCW motifs 70% of the time and APOBEC3B 50% of the time (Y = C/T; W = A/T). Signature comparisons with breast tumor whole genome sequences indicate that most malignancies manifest intermediate percentages of APOBEC3 signature mutations in YTCW motifs, mostly between 50 and 70%, suggesting that both enzymes contribute in a combinatorial manner to the overall mutation landscape. Although the vast majority of APOBEC3A- and APOBEC3B-induced single base substitution mutations occur outside of predicted chromosomal DNA hairpin structures, whole genome sequence analyses and supporting biochemical studies also indicate that both enzymes are capable of deaminating the single-stranded loop regions of DNA hairpins at elevated rates. These studies combine to help resolve a long-standing etiologic debate on the source of APOBEC3 signature mutations in cancer and indicate that future diagnostic and therapeutic efforts should focus on both APOBEC3A and APOBEC3B.


Assuntos
Neoplasias da Mama , Humanos , Feminino , Neoplasias da Mama/genética , Mutação , Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Linhagem Celular , DNA/metabolismo , Antígenos de Histocompatibilidade Menor/genética , Citosina/metabolismo
19.
PLoS Comput Biol ; 19(10): e1011503, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37862377

RESUMO

Populations of cells typically maintain a consistent size, despite cell division rarely being precisely symmetrical. Therefore, cells must possess a mechanism of "size control", whereby the cell volume at birth affects cell-cycle progression. While size control mechanisms have been elucidated in a number of other organisms, it is not yet clear how this mechanism functions in plants. Here, we present a mathematical model of the key interactions in the plant cell cycle. Model simulations reveal that the network of interactions exhibits limit-cycle solutions, with biological switches underpinning both the G1/S and G2/M cell-cycle transitions. Embedding this network model within growing cells, we test hypotheses as to how cell-cycle progression can depend on cell size. We investigate two different mechanisms at both the G1/S and G2/M transitions: (i) differential expression of cell-cycle activator and inhibitor proteins (with synthesis of inhibitor proteins being independent of cell size), and (ii) equal inheritance of inhibitor proteins after cell division. The model demonstrates that both these mechanisms can lead to larger daughter cells progressing through the cell cycle more rapidly, and can thus contribute to cell-size control. To test how these features enable size homeostasis over multiple generations, we then simulated these mechanisms in a cell-population model with multiple rounds of cell division. These simulations suggested that integration of size-control mechanisms at both G1/S and G2/M provides long-term cell-size homeostasis. We concluded that while both size independence and equal inheritance of inhibitor proteins can reduce variations in cell size across individual cell-cycle phases, combining size-control mechanisms at both G1/S and G2/M is essential to maintain size homeostasis over multiple generations. Thus, our study reveals how features of the cell-cycle network enable cell-cycle progression to depend on cell size, and provides a mechanistic understanding of how plant cell populations maintain consistent size over generations.


Assuntos
Modelos Teóricos , Células Vegetais , Humanos , Recém-Nascido , Divisão Celular , Ciclo Celular/fisiologia , Tamanho Celular
20.
Appl Environ Microbiol ; 89(10): e0033123, 2023 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-37791775

RESUMO

Nucleic acid-based assays, such as polymerase chain reaction (PCR), that amplify and detect organism-specific genome sequences are a standard method for infectious disease surveillance. However, challenges arise for virus surveillance because of their genetic diversity. Here, we calculated the variability of nucleotides within the genomes of 10 human viral species in silico and found that endemic viruses exhibit a high percentage of variable nucleotides (e.g., 51.4% for norovirus genogroup II). This genetic diversity led to the variable probability of detection of PCR assays (the proportion of viral sequences that contain the assay's target sequences divided by the total number of viral sequences). We then experimentally confirmed that the probability of the target sequence detection is indicative of the number of mismatches between PCR assays and norovirus genomes. Next, we developed a degenerate PCR assay that detects 97% of known norovirus genogroup II genome sequences and recognized norovirus in eight clinical samples. By contrast, previously developed assays with 31% and 16% probability of detection had 1.1 and 2.5 mismatches on average, respectively, which negatively impacted RNA quantification. In addition, the two PCR assays with a lower probability of detection also resulted in false negatives for wastewater-based epidemiology. Our findings suggest that the probability of detection serves as a simple metric for evaluating nucleic acid-based assays for genetically diverse virus surveillance.IMPORTANCENucleic acid-based assays, such as polymerase chain reaction (PCR), that amplify and detect organism-specific genome sequences are employed widely as a standard method for infectious disease surveillance. However, challenges arise for virus surveillance because of the rapid evolution and genetic variation of viruses. The study analyzed clinical and wastewater samples using multiple PCR assays and found significant performance variation among the PCR assays for genetically diverse norovirus surveillance. This finding suggests that some PCR assays may miss detecting certain virus strains, leading to a compromise in detection sensitivity. To address this issue, we propose a metric called the probability of detection, which can be simply calculated in silico using a code developed in this study, to evaluate nucleic acid-based assays for genetically diverse virus surveillance. This new approach can help improve the sensitivity and accuracy of virus detection, which is crucial for effective infectious disease surveillance and control.


Assuntos
Doenças Transmissíveis , Norovirus , Humanos , Norovirus/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , RNA Viral/genética , Nucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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