Biology subjects

Cao, Q.

Publications and source records attributed to Cao, Q..

At least 55 records · Page 3Linked to original sources

bioRxiv2024

Open-source 3D printed manifolds for exposure studies using human airway epithelial cells

RationaleInhalation of airborne stimuli can damage the airway epithelium, increasing the risk of developing respiratory or systemic diseases. In vitro studies using air-liquid interface cell cultures enable controlled investigation of cellular responses to relevant exposures. Commercial in vitro exposure systems provide precise and reproducible dosage but require significant capital investment and are not amenable to customization. Research groups interested in respiratory exposure science may benefit from a more accessible alternative open-source exposure system. We present 3D printed manifolds for applying a range of airborne exposures uniformly across standard, commercially available 6- and 24-well plates with air-liquid interface culture inserts. MethodsA simple chamber-style exposure system and the manifolds were evaluated for exposure uniformity via computational fluid dynamics simulations and deposition of nebulized FITC-labelled dextran. The chamber and manifolds were manufactured using a stereolithography 3D printer. Cannabis concentrate vapor was generated from 3 different vaporizers and applied to well plates using the manifold system. Calu-3 cells were cultured on Transwell inserts and exposed to whole tobacco smoke or room air. ResultsThe manifolds produced less variation in simulated air velocities and physical deposition of FITC-dextran aerosol deposition across well plates compared to those of the chamber-style exposure system. Distinct doses of cannabis concentrate vapour were delivered to well plates with low variation among wells. Whole tobacco smoke exposure using the manifold system induced functional changes in Calu-3 airway epithelial cell barrier function, cytokine production (IL-6 and IL-8), and cell membrane potential. ConclusionsCollectively, our data demonstrate the feasibility and the validity of our open-source 3D printed manifolds for use in studying various respiratory exposures and position our designs as more accessible options in parallel with commercially available systems. All article content is licensed under a Creative Commons Attribution (CC BY-NC 4.0) license (https://creativecommons.org/licenses/by-nc/4.0/).

bioengineering↗

bioRxiv2024

tp53 R217H and R242H Mutant Zebrafish Exhibit Dysfunctional p53 Hallmarks and Recapitulate Li-Fraumeni Syndrome Phenotypes

Li-Fraumeni syndrome (LFS) is a hereditary cancer predisposition syndrome associated with a highly penetrant cancer spectrum characterized by germline TP53 mutations. We characterized the first LFS zebrafish hotspot mutants, tp53 R217H and R242H (human R248H and R273H), and found these mutants exhibit partial-to-no activation of p53 target genes, have a defective G1 cell-cycle checkpoint, and are resistant to apoptosis. Spontaneous tumor development histologically resembling human sarcomas developed as early as 6 months. tp53 R242H mutants had a higher lifetime tumor incidence compared to tp53 null and R217H mutants, suggesting it is a more aggressive mutation. We observed mutation-specific tumor phenotypes across tp53 mutants with associated diverse transcriptomic and DNA methylome profiles, impacting metabolism, cell signalling, and biomacromolecule synthesis and degradation. These tp53 zebrafish mutants demonstrate fidelity to their human counterparts and provide new insights into underlying tumorigenesis mechanisms and kinetics, which will guide targeted therapeutics for LFS.

cancer biology↗

bioRxiv2024

Single field evolution rule governs the dynamics of representational drift in mouse hippocampal dorsal CA1 region.

How the brain reconciles dynamism with stability to balance learning and reliable memory storage has not yet been fully understood. To address the critical question, we longitudinally recorded place cells in the hippocampal dorsal CA1 region over 7 to 56 days, utilizing multiple goal-oriented navigation paradigms across various environments. We found that over 80% of place cells displayed multiple fields, undergoing complex evolution events including field disappearance, formation, and retention. Place fields from the same neuron showed limited coordination ([~]5%), with a preference for synchronized changes. We further uncovered the single field evolution rule: the longer a field remains active, the more likely it is to continue being active; conversely, the longer a field remains inactive, the less likely it is to recover the future fate of a place field depends on its past activity. Mathematical modeling revealed that this rule sufficiently demonstrates the growing stability of the dCA1 spatial representation at the population level.

neuroscience↗

bioRxiv2024

Avian-restrict Salmonella transition to endemicity is accompanied by localized resistome adaptation

Bacterial regional demonstration after global dissemination is an essential pathway for selecting distinct finesses. However, the evolution of the resistome during the transition to endemicity remains unaddressed. Using the most comprehensive whole-genome sequencing dataset of Salmonella enterica serovar Gallinarum (S. Gallinarum) collected from 15 countries, including 45 newly recovered samples from two related local regions, we established the relationship among avian-specific pathogen genetic profiles and localization patterns. Initially, we revealed the international transmission and evolutionary history of S. Gallinarum to recent endemicity through phylogenetic analysis conducted using a spatiotemporal Bayesian framework. Our findings indicate that the independent acquisition of the resistome via the mobilome, primarily through plasmids and transposons shapes a unique antimicrobial resistance profile among different lineages. Notably, the mobilome-resistome combination among distinct lineages exhibits a geographical-specific manner, further supporting a localized endemic mobilome-driven process. Collectively, this study elucidates resistome adaptation in the endemic transition of an avian-specific pathogen, likely driven by the localized farming style, and provides valuable insights for targeted interventions.

microbiology↗

bioRxiv2024

Cellular and Molecular Changes During Aging in MEC: Unveiling the Role of Bglap3 Neurons in Cognitive Aging

Aging-correlated cognitive declines, including deficiencies in spatial orientation and memory, may reflect dysfunction in the hippocampus and medial entorhinal cortex (MEC). However, aging-related changes in MEC at the cellular and molecular levels remain unclear. In this study, we found fewer grid cells with reduced spatial stability in old mice. We compared gene expression profiles between young and old mice using 10x Genomics Visium technology. Among 1664 differentially expressed genes, we discovered Bglap3, a marker gene for subpopulation in MEC Layer III with decreased cell number with age. Silencing of Bglap3+ neurons in young mice impaired the spatial tuning of neurons in MEC and the spatial learning of a new platform location in water maze. These findings help us to understand the cellular and molecular changes in the MEC in healthy aging animals and the changes of Bglap3+ cells in old mice indicating a possible cause of aging-related MEC deficiency.

neuroscience↗

bioRxiv2024

Synovial Sarcoma Chromatin Dynamics Reveal a Continuum in SS18:SSX Reprograming

Synovial sarcoma (SyS) is an aggressive soft-tissue malignancy characterized by a pathognomonic chromosomal translocation leading to the formation of the SS18::SSX fusion oncoprotein. SS18::SSX associates with mammalian BAF complexes suggesting deregulation of chromatin architecture as the oncogenic driver in this tumour type. To examine the epigenomic state of SyS we performed comprehensive multi-omics analysis on 52 primary pre-treatment human SyS tumours. Our analysis revealed a continuum of epigenomic states across the cohort at fusion target genes independent of rare somatic genetic lesions. We identify cell-of-origin signatures defined by enhancer states and reveal unexpected relationships between H2AK119Ub1 and active marks. The number of bivalent promoters, dually marked by the repressive H3K27me3 and activating H3K4me3 marks, has strong prognostic value and outperforms tumor grade in predicting patient outcome. Finally, we identify SyS defining epigenomic features including H3K4me3 expansion associated with striking promoter DNA hypomethylation in which SyS displays the lowest mean methylation level of any sarcoma subtype. We explore these distinctive features as potential vulnerabilities in SyS and identify H3K4me3 inhibition as a promising therapeutic strategy.

cancer biology↗

bioRxiv2024

Normative growth modeling of brain morphology reveals neuroanatomical heterogeneity and biological subtypes in children with ADHD

BackgroundNeuroimaging studies suggest substantial individual heterogeneity in brain phenotypes in attention-deficit/hyperactivity disorder (ADHD). However, how these individual-level brain phenotypes contribute to the identification of ADHD biotypes and whether these biotypes have different treatment outcomes and neurobiological underpinnings remain largely unknown. MethodsWe collected multisite, high-quality structural magnetic resonance imaging data from 1,006 children aged 6-14 years, including 351 children with ADHD and 655 typically developing children. Normative growth models of cortical thickness were established for 219 regions in the typically developing children. Individual-level deviations from these normal references were quantified and clustered to identify ADHD biotypes. We validated the replicability and generalizability of the ADHD biotypes using two independent datasets and evaluated the associations of the biotypes with symptomatic, cognitive, and gene expression profiles, as well as follow-up treatment outcomes. FindingsNo more than 10% of children with ADHD had extreme deviations in cortical thickness in a single region, suggesting high heterogeneity among individuals with ADHD. On the basis of the brain deviation maps, we discovered two robust ADHD biotypes, an infra-normal subtype with cortical thinning associated with ADHD symptoms and a supranormal subtype with cortical thickening associated with cognition. Patients with the infra-normal subtype responded better to methylphenidate than to atomoxetine, although both subtypes showed treatment efficacy. Brain deviations in the infra-normal subtype were explained by the expression levels of genes enriched in presynaptic and axonal development and polygenic risk of ADHD. InterpretationWe identified anatomically distinct, clinically valuable, and biologically informed ADHD subtypes, providing insight into the neurobiological basis of clinical heterogeneity and facilitating a personalized medication strategy for ADHD patients. Panel: Research in contextO_ST_ABSEvidence before this studyC_ST_ABSSubstantial individual heterogeneity in brain phenotypes in attention-deficit/hyperactivity disorder (ADHD) motivates the need to discover homogeneous biotypes. We searched PubMed for research articles on ADHD biotypes using brain MRI published before December 1, 2023, using the search terms ((attention deficit hyperactivity disorder [Title/Abstract]) OR (ADHD [Title/Abstract])) AND ((subtypes [Title/Abstract]) OR (subgroups [Title/Abstract]) OR (subtyping [Title/Abstract])) AND ((MRI [Title/Abstract]) OR (neuroimaging [Title/Abstract]) OR (brain [Title/Abstract])) without language restrictions. Of the eight included studies, two identified ADHD biotypes using structural morphology, four used functional activity, and two used multimodal features. However, none of these studies considered the developmental effect of the brain phenotypes, examined treatment response, or investigated the genetic correlates of the biotypes. Added value of this studyThis study is the first to use individualized brain measures extracted from normative models to investigate ADHD biotypes in a large sample of more than 1,000 children. We identified two reproducible ADHD biotypes, characterized by distinct symptomatic, cognitive, and gene expression profiles, as well as differential treatment responses. This study advances our understanding of the neurobiological basis underlying the clinical heterogeneity of ADHD and highlights the critical need to discover ADHD biotypes using an unbiased and individualized approach. Implications of all the available evidenceThis study revealed remarkable neuroanatomical heterogeneity in ADHD patients and identified anatomically distinct, clinically valuable, and biologically informed ADHD biotypes. Our findings have potential value for the investigation of data-driven biotypes to evaluate treatment efficacy and facilitate personalized treatment. We also highlight the need for future studies to move beyond the understanding of ADHD solely based on the "average patient" perspective.

neuroscience↗

bioRxiv2024

Commonly used software tools produce conflicting and overly-optimistic AUPRC values

The precision-recall curve (PRC) and the area under it (AUPRC) are useful for quantifying classification performance. They are commonly used in situations with imbalanced classes, such as cancer diagnosis and cell type annotation. We evaluated 10 popular tools for plotting PRC and computing AUPRC, which were collectively used in >3,000 published studies. We found the AUPRC values computed by the tools rank classifiers differently and some tools produce overly-optimistic results.

bioinformatics↗

bioRxiv2024

Cardiac Localized Polycystin-2 plays a Functional Role in Natriuretic Peptide Production and its Absence Contributes to Hypertension

Cardiovascular complications are the most common cause of mortality in patients with autosomal dominant polycystic kidney disease (ADPKD). Hypertension is seen in 70% of patients by the age of 30 prior to decline in kidney function. The natriuretic peptides (NPs), atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), are released by cardiomyocytes in response to membrane stretch, increasing urinary excretion of sodium and water. Mice heterozygous for Pkd2 have attenuated NP responses and we hypothesized that cardiomyocyte-localized polycystin proteins contribute to production of NPs. Cardiomyocyte-specific knock-out models of polycystin-2 (PC2), one of the causative genes of ADPKD, demonstrate diurnal hypertension. These mice have decreased ANP and BNP expression in the left ventricle. Analysis of the pathways involved in production, maturation, and activity of NPs identified decreased transcription of CgB, PCSK6, and NFAT genes in cPC2-KOs. Engineered heart tissue with human iPSCs driven into cardiomyocytes with CRISPR/Cas9 KO of PKD2 failed to produce ANP. These results suggest that PC2 in cardiomyocytes are involved in NP production and lack of cardiac PC2 predisposes to a hypertensive volume expanded phenotype, which may contribute to the development of hypertension in ADPKD.

physiology↗

bioRxiv2023

Crosstalk between Gut Sensory Ghrelin Signaling and Adipose Tissue Sympathetic Outflow Regulates Metabolic Homeostasis

The stomach-derived orexigenic hormone ghrelin is a key regulator of energy homeostasis and metabolism in humans. The ghrelin receptor, growth hormone secretagogue receptor 1a (GHSR), is widely expressed in the brain and gastrointestinal vagal sensory neurons, and neuronal GHSR knockout results in a profoundly beneficial metabolic profile and protects against diet-induced obesity (DIO) and insulin resistance. Here we show that in addition to the well characterized vagal GHSR, GHSR is robustly expressed in gastrointestinal sensory neurons emanating from spinal dorsal root ganglia. Remarkably, sensory neuron GHSR deletion attenuates DIO through increased energy expenditure and sympathetic outflow to adipose tissue independent of food intake. In addition, neuronal viral tract tracing reveals prominent crosstalk between gut non-vagal sensory afferents and adipose sympathetic outflow. Hence, these findings demonstrate a novel gut sensory ghrelin signaling pathway critical for maintaining energy homeostasis.

neuroscience↗

bioRxiv2023

Utilization of the genetically encoded calcium indicator Salsa6F in cardiac applications

Calcium signaling is a critical process required for cellular mechanisms such as cardiac contractility. The inability of the cell to properly activate or regulate calcium signaling can lead to contractile dysfunction. In isolated cardiomyocytes, calcium signaling has been primarily studied using calcium fluorescent dyes, however these dyes have limited applicability to whole organs. Here, we crossed the Salsa6f mouse which expresses a genetically encoded ratiometric cytosolic calcium indicator with a cardiomyocyte specific inducible cre to temporally-induce expression and studied cytosolic calcium transients in isolated cardiomyocytes and modified Langendorff heart preparations. Isolated cardiomyocytes expressing Salsa6f or Fluo-4AM loaded were compared. We also crossed the Salsa6f mouse with a floxed Polycystin 2 (PC2) mouse to test the feasibility of using the Salsa6f mouse to measure calcium transients in PC2 heterozygous or homozygous knock out mice. Although there are caveats in the applicability of the Salsa6f mouse, there are clear advantages to using the Salsa6f mouse to measure whole heart calcium signals.

physiology↗

bioRxiv2023

Virus-derived circular RNAs populate hepatitis C virus-infected cells

It is known that pre-mRNAs in eukaryotic cells can be processed to circular RNAs by a back- splicing mechanism. Circular RNAs have great stability and can sequester proteins or small RNAs to exert functions on cellular pathways. Because viruses often exploit host pathways, we explored whether the RNA genome of the cytoplasmic hepatitis C virus is processed to yield virus-derived circRNAs (vcircRNAs). Computational analyses of RNA-seq experiments predicted that the viral RNA genome is fragmented to generate hundreds of vcircRNAs. More than a dozen of them were experimentally verified by rolling-circle amplification. VcircRNAs that contained the viral internal ribosome entry site were found to be translated into novel proteins that displayed pro-viral functions. Furthermore, a highly abundant, non-translated vcircRNA was shown to enhance viral RNA abundance. These findings argue that novel vcircRNA molecules modulate viral amplification in cells infected by a cytoplasmic RNA virus. Significance StatementProcessing of an RNA viral genome into hundreds of circular RNAs provides novel pro-viral functions and can promote translation of novel viral peptides.

microbiology↗

bioRxiv2023

The central nervous system is a potential reservoir and possible origin of drug resistance in HBV

The significance of HBV in cerebrospinal fluid (CSF) is unclear. In the present study, synchronous serum and CSF samples were collected from 13 patients. HBV-DNA, full-length genome, quasispecies, phylogenetic tree, compartmentalization and mutation of reverse transcriptase (RT) region analyses were performed based on PCR and sequencing methods. We found HBV-DNA was detected in the CSF of 3 antiviral-naive patients and one patient after successful antiviral therapy. Complete full-length HBV genomes were isolated from the CSF of 5 patients, including 2 patients with undetectable serum HBV-DNA. Ten patients exhibited distinct CSF-serum quasispecies, 8 harbored independent CSF-serum genetic compartmentalization and phylogenetic trees, and 5 patients presented lamivudine/entecavir-associated resistance mutations in only the CSF. The frequencies of rtL180M and rtM204I/V mutations in both serum and CSF were higher in HIV/HBV-coinfected patients than in HBV-monoinfected patients (serum: rtL180M: 3.9% vs. 0, P = 0.004; rtM204I/V: 21.3% vs. 0, P < 0.001; CSF: rtL180M: 7.6% vs. 0, P = 0.026; rtM204I/V 7.6% vs. 1.6%, P = 0.097). Our data suggested CSF is a potential HBV reservoir, and HBV in CSF harbors distinct evolution and mutation models from that in serum. HIV infection increases the possibility of HBV rtL180M and rtM204I/V mutations in both serum and CSF.

evolutionary biology↗

bioRxiv2023

Implantable flexible multielectrode arrays for multi-site sensing of serotonin tonic levels

Real-time multi-channel measurements of tonic serotonin (5-hydroxytryptamine, 5-HT) concentrations across different brain regions are of utmost importance to the understanding of 5-HTs role in anxiety, depression, and impulse control disorders, which will improve the diagnosis and treatment of these neuropsychiatric illnesses. Chronic sampling of 5-HT is critical in tracking disease development as well as the time course of pharmacological treatments. Despite their value, in vivo chronic multi-site measurements of 5-HT have not been reported. To fill this technological gap, we batch fabricated implantable glassy carbon (GC) microelectrode arrays (MEAs) on a flexible SU-8 substrate to provide an electrochemically stable and biocompatible device/tissue interface. Then, to achieve multi-site detection of tonic 5-HT concentrations, we incorporated the poly(3,4-ethylenedioxythiophene)/functionalized carbon nanotube (PEDOT/CNT) coating on the GC microelectrodes in combination with a new square wave voltammetry (SWV) approach, optimized for selective 5-HT measurement. In vitro, the PEDOT/CNT coated GC microelectrodes achieved high sensitivity towards 5-HT, good fouling resistance in the presence of 5-HT, and excellent selectivity towards the most common neurochemical interferents. In vivo, our PEDOT/CNT-coated GC MEAs were able to successfully detect basal 5-HT concentrations at different locations of the CA2 hippocampal region of mice in both anesthetized and awake head-fixed conditions. Furthermore, the implanted PEDOT/CNT-coated MEA achieved stable detection of tonic 5-HT concentrations for one week. Finally, histology data in the hippocampus shows reduced tissue damage and inflammatory responses compared to stiff silicon probes. To the best of our knowledge, this PEDOT/CNT-coated GC MEA is the first implantable flexible multisite sensor capable of chronic in vivo multi-site sensing of tonic 5-HT. This implantable MEA can be custom-designed according to specific brain region of interests and research questions, with the potential to combine electrophysiology recording and multiple analyte sensing to maximize our understanding of neurochemistry. HighlightsO_LIPEDOT/CNT-coated GC microelectrodes enabled sensitive and selective tonic detection of serotonin (5-HT) using a new square wave voltammetry (SWV) approach C_LIO_LIPEDOT/CNT-coated GC MEAs achieved multi-site in vivo 5-HT tonic detection for one week. C_LIO_LIFlexible MEAs lead to reduced tissue damage and inflammation compared to stiff silicon probes. C_LI

bioengineering↗

bioRxiv2022

Haploid-resolved and chromosome-scale genome assembly in hexa-autoploid sweetpotato (Ipomoea batatas (L.) Lam)

Sweetpotato (Ipomoea batatas (L.) Lam) is the worlds seventh most important food crop by production quantity. Cultivated sweetpotato is a hexaploid (2n = 6x = 90), and its genome (B1B1B2B2B2B2) is quite complex due to polyploidy, self-incompatibility, and high heterozygosity. Here we established a haploid-resolved and chromosome-scale de novo assembly of autohexaploid sweetpotato genome sequences. Before constructing the genome, we created chromosome-scale genome sequences in I. trifida using a highly homozygous accession, Mx23Hm, with PacBio RSII and Hi-C reads. Haploid-resolved genome assembly was performed for a sweetpotato cultivar, Xushu18 by hybrid assembly with Illumina paired-end (PE) and mate-pair (MP) reads, 10X genomics reads, and PacBio RSII reads. Then, 90 chromosome-scale pseudomolecules were generated by aligning the scaffolds onto a sweetpotato linkage map. De novo assemblies were also performed for chloroplast and mitochondrial genomes in I. trifida and sweetpotato. In total, 34,386 and 175,633 genes were identified on the assembled nucleic genomes of I. trifida and sweetpotato, respectively. Functional gene annotation and RNA-Seq analysis revealed locations of starch, anthocyanin, and carotenoid pathway genes on the sweetpotato genome. This is the first report of chromosome-scale de novo assembly of the sweetpotato genome. The results are expected to contribute to genomic and genetic analyses of sweetpotato.

genomics↗

bioRxiv2022

Heterotrophic bacteria drive sulfide oxidation in coastal sediments

Sulfate reduction and sulfur oxidation are very active in coastal sediments. They shape the biogeochemistry and microbial ecology at hot places of organic matter metabolism. Different from the well-studied sulfate reduction, sulfur oxidation in coastal sediments is still full of questions. Herein, we investigated the distribution of reduced sulfur compounds in differently layers of coastal sediments at the Yellow sea and found that sulfide (H2S), sulfane sulfur (S0), and thiosulfate mainly accumulated in anaerobic sediments and were mostly oxidized in anoxic and oxic interface in the sediments and the sea water. Bacterial community analysis indicated that heterotrophic bacteria are dominating species in surface sediments and sea water. Metagenome analysis showed that two sulfur-oxidizing genes encoding sulfide:quinone oxidoreductases (SQR) and persufide dioxygenases (PDO), were sharply more abundant than other sulfur-oxidizing genes in the coastal sediments. Since members of the marine Roseobacter clade were dominant in coastal waters and sediments, we studied the sulfur oxidation pathway in the Roseobacter Ruegeria pomeroyi DSS-3 and found that sulfide:quinone oxidoreductase, persulfide dioxygenase, and sulfite-oxidizing enzyme were the main enzymes for the oxidation of H2S, zerovalent sulfur, and sulfite/thiosulfate. This study, for the first time, clarified the dominating function of heterotrophic bacteria in sulfur oxidation in the coastal sediments and sea water. IMPORTANCECoastal sediments are the most productive ecosystems. We performed the microbial community diversity and metagenomic analysis of seawater and coastal sediments of the Yellow Sea and explored the sulfur oxidation process in them. We found that heterotrophic bacteria are dominating species in surface sediments and sea water, sulfide and sulfane sulfur were mostly oxidized in surface sediments, and the genes encoding SQR, PDO, and SOE are abundant. Using Ruegeria pomeroyi DSS-3 as the model strain, we studied how these enzymes cooperate to oxidize H2S to sulfate. Thus, this research revealed the critical role of heterotrophic bacteria in sulfur oxidation in coastal sediments and sea water.

microbiology↗

bioRxiv2022

Synthetic extremophiles: Species-specific formulations for microbial therapeutics and beyond.

Microorganisms have been used for millennia to produce food and medicine and are now being developed as products themselves to treat disease and boost crop production. However, as required for these new applications, maintaining high viability throughout manufacturing, transportation and use remains a significant challenge requiring sophisticated cold-chains and packaging. In fact, we found that commercial microbial products (probiotics) provide a poor solution to this challenge, in particular for key industrial organisms like E. coli. To overcome this technological gap, we report the development of synthetic extremophiles of industrially important gram-negative bacteria (E. coli Nissle 1917, Ensifer meliloti), gram positive bacteria (Lactobacillus plantarum) and yeast (Saccharomyces boulardii). Specifically, we developed a high throughput pipeline to define species-specific materials that allow these organisms to survive drying, elevated temperatures, organic solvents and even ionizing radiation. We enhanced the stability of E.coli Nissle 1917 by >4 orders of magnitude over commercial formulations and demonstrate the capacity to remain viable while undergoing tableting and pharmaceutical methodologies involving organic solvents. The development of synthetic materials-based enhanced stabilization stands to transform our capacity to apply micro-organisms in extreme environments including those found on Earth as well as in space. One-Sentence SummaryFragile therapeutic bacteria can be made to survive the manufacturing extremes normally reserved for small molecule drugs.

microbiology↗

bioRxiv2022

Transcription-independent hold of the G1/S transition is exploited to cope with DNA replication stress

RB1 (retinoblastoma) members control the G1/S commitment as transcriptional repressors in eukaryotic cells. Here we uncover that an extra copy of RB1 equivalent (WHI7 or WHI5) is sufficient to bypass the indispensability of the central genomic checkpoint kinases Mec1ATR-Rad53CHK1 in Saccharomyces cerevisiae. Mec1-Rad53 directly phosphorylate Whi7/5, antagonizing their nuclear export or protein turnover upon replication stress. Through in vitro reconstitution, we show that Whi7 C-terminus directly binds and hinders S-CDK-Cks1 from processively phosphorylating Sic1. By microfluidic single-cell real-time quantitative imaging, we demonstrate that both Whi7 and Whi5 are required to flatten the degradation curve of the major S-CDK inhibitor Sic1 in vivo. These findings reveal an eclipsed transcription-independent role of Whi7 homologs, which is highlighted by genome integrity checkpoints to hold the G1/S transition instantly as a rapid response to unforeseeable replication threats. Key pointsO_LIWhi7 overexpression bypasses the essential function of Mec1 and Rad53 in a transcription-independent way. C_LIO_LIWhi7 is stabilized by checkpoint-mediated phosphorylation. C_LIO_LIWhi7 binds and hinders S-CDK-Cks1 from multi-phosphorylation of Sci1, thereby prolonging Sic1 degradation and G1/S transition. C_LI

molecular biology↗