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Ma, Y.

Publications and source records attributed to Ma, Y..

At least 307 records · Page 17Linked to original sources

TSD: Transformers for Seizure Detection

Epilepsy is a common neurological disorder that sub-stantially deteriorates patients safety and quality of life. Electroencephalogram (EEG) has been the golden-standard technique for diagnosing this brain disorder and has played an essential role in epilepsy monitoring and disease management. It is extremely laborious and challenging, if not practical, for physicians and expert humans to annotate all recorded signals, particularly in long-term monitoring. The annotation process often involves identifying signal segments with suspected epileptic seizure features or other abnormalities and/or known healthy features. Therefore, automated epilepsy detection becomes a key clinical need because it can greatly improve clinical practices efficiency and free up human expert time to attend to other important tasks. Current automated seizure detection algorithms generally face two challenges: (1) models trained for specific patients, but such models are patient-specific, hence fail to generalize to other patients and real-world situations; (2) seizure detection models trained on large EEG datasets have low sensitivity and/or high false positive rates, often with an area under the receiver operating characteristic (AUROC) that is not high enough for potential clinical applicability. This paper proposes Transformers for Seizure Detection, which we refer to as TSD in this manuscript. A Transformer is a deep learning architecture based on an encoder-decoder structure and on attention mechanisms, which we apply to recorded brain signals. The AUROC of our proposed model has achieved 92.1%, tested with Temple Universitys publically available electroencephalogram (EEG) seizure corpus dataset (TUH). Additionally, we highlight the impact of input domains on the models performance. Specifically, TSD performs best in identifying epileptic seizures when the input domain is a time-frequency. Finally, our proposed model for seizure detection in inference-only mode with EEG recordings shows outstanding performance in classifying seizure types and superior model initialization.

bioengineering↗

Lung infection by P. aeruginosa induces neuroinflammation and blood-brain barrier dysfunction in mice

BackgroundSevere lung infection can lead to brain dysfunction and neurobehavioral disorders. The mechanisms that regulate the lung-brain axis of inflammatory response to respiratory infection are incompletely understood. This study examined the effects of lung infection causing systemic and neuroinflammation as a potential mechanism contributing to blood-brain barrier (BBB) leakage and behavioral impairment. MethodsPneumonia was induced in adult C57BL/6 mice by intratracheal inoculation of Pseudomonas aeruginosa (PA). Solute extravasation, histology, immunofluorescence, RT-PCR, multiphoton imaging and neurological testing were performed in this study. ResultsLung infection caused alveolar-capillary barrier injury as indicated by leakage of plasma proteins across pulmonary microvessels and histopathological characteristics of pulmonary edema (alveolar wall thickening, microvessel congestion, and neutrophil infiltration). PA also caused significant BBB dysfunction characterized by leakage of different sized molecules across cerebral microvessels and a decreased expression of cell-cell junctions (VE-cadherin, claudin-5) in the brain. BBB leakage peaked at 24 hours and lasted for 7 days post-inoculation. Additionally, mice with lung infection displayed hyperlocomotion and anxiety-like behaviors. To test whether cerebral dysfunction was caused by PA directly or indirectly, we measured bacterial load in multiple organs. While PA loads were detected in the lungs up to 7 days post-inoculation, bacteria were not detected in the brain as evidenced by negative cerebral spinal fluid (CSF) cultures and lack of distribution in different brain regions or isolated cerebral microvessels. However, mice with PA lung infection demonstrated increased mRNA expression in the brain of pro-inflammatory cytokines (IL-1{beta}, IL-6, and TNF-), chemokines (CXCL-1, CXCL-2) and adhesion molecules (VCAM-1 and ICAM-1) along with CD11b+ cell recruitment, corresponding to their increased blood levels of white cells (polymorphonuclear cells) and cytokines. To confirm the direct effect of cytokines on endothelial permeability, we measured cell-cell adhesive barrier resistance and junction morphology in mouse brain microvascular endothelial cell monolayers, where administration of IL-1{beta} induced a significant reduction of barrier function coupled with tight junction (TJ) diffusion and disorganization. Combined treatment with IL-1{beta} and TNF augmented the barrier injury. ConclusionsThese results suggest that lung bacterial infection causes cerebral microvascular leakage and neuroinflammation via a mechanism involving cytokine-induced BBB injury.

pharmacology and toxicology↗

Pharmacological depletion of RNA splicing factor RBM39 by indisulam synergizes with PARP inhibitors in high-grade serous ovarian carcinoma

Ovarian high-grade serous carcinoma (HGSC) is the most common and lethal subtype of ovarian cancer with limited therapeutic options. In recent years, PARP inhibitors have demonstrated significant clinical benefits, especially in patients with BRCA1/2 mutations. However, acquired drug resistance and relapse is a major challenge. Therapies disrupting the spliceosome alter cancer transcriptomes and have shown potential to improve PARP inhibitor response. Indisulam (E7070) has been identified as a molecular glue that brings splicing factor RBM39 and DCAF15 E3 ubiquitin ligase in close proximity. Exposure to indisulam induces RBM39 proteasomal degradation through DCAF15-mediated polyubiquitination and subsequent RNA splicing defects. In this study, we demonstrate that loss of RBM39 induces splicing errors in DNA damage repair genes in ovarian cancer, leading to increased sensitivity to PARP inhibitors such as olaparib. Indisulam synergized with olaparib in multiple in vitro models of ovarian cancer regardless of PARP inhibitor sensitivity and improved olaparib response in mice bearing PARP inhibitor-resistant tumors. DCAF15 expression, but not BRCA1/2 mutational status, was essential for the synergy between indisulam and olaparib, suggesting that the combination therapy may benefit patients irrespective of their BRCA1/2 status. These findings demonstrate that combining RBM39 degraders and PARP inhibitors is a promising therapeutic approach to improving PARP inhibitor response in ovarian HGSC. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/524417v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@4cccd5org.highwire.dtl.DTLVardef@1be34aorg.highwire.dtl.DTLVardef@e72b06org.highwire.dtl.DTLVardef@b9c89d_HPS_FORMAT_FIGEXP M_FIG C_FIG One Sentence SummaryWe identified a novel drug combination that may improve PARP inhibitor response and benefit a large group of ovarian cancer patients.

cancer biology↗

A Systematic Survey of Reversibly Covalent Dipeptidyl Inhibitors of the SARS-CoV-2 Main Protease

SARS-CoV-2 is the coronavirus pathogen of the currently prevailing COVID-19 pandemic. It relies on its main protease (MPro) for replication and pathogenesis. MPro is a demonstrated target for the development of antivirals for SARS-CoV-2. Past studies have systematically explored tripeptidyl inhibitors such as nirmatrelvir as MPro inhibitors. However, dipeptidyl inhibitors especially those with a spiro residue at their P2 position have not been systematically investigated. In this work, we synthesized about 30 reversibly covalent dipeptidyl MPro inhibitors and characterized them on in vitro enzymatic inhibition potency, structures of their complexes with MPro, cellular MPro inhibition potency, antiviral potency, cytotoxicity, and in vitro metabolic stability. Our results indicated that MPro has a flexible S2 pocket that accommodates dipeptidyl inhibitors with a large P2 residue and revealed that dipeptidyl inhibitors with a large P2 spiro residue such as (S)-2-azaspiro[4,4]nonane-3-carboxylate and (S)-2-azaspiro[4,5]decane-3-carboxylate have optimal characteristics. One compound MPI60 containing a P2 (S)-2-azaspiro[4,4]nonane-3-carboxylate displayed high antiviral potency, low cellular cytotoxicity, and high in vitro metabolic stability and can be potentially advanced to further preclinical tests.

biochemistry↗

Spike substitution T813S increases Sarbecovirus fusogenicity by enhancing the usage of TMPRSS2.

SARS-CoV Spike (S) protein shares considerable homology with SARS-CoV-2 S, especially in the conserved S2 subunit (S2). S protein mediates coronavirus receptor binding and membrane fusion, and the latter activity can greatly influence coronavirus infection. We observed that SARS-CoV S is less effective in inducing membrane fusion compared with SARS-CoV-2 S. We identify that S813T mutation is sufficient in S2 interfering with the cleavage of SARS-CoV-2 S by TMPRSS2, reducing spike fusogenicity and pseudoparticle entry. Conversely, the mutation of T813S in SARS-CoV S increased fusion ability and viral replication. Our data suggested that residue 813 in the S was critical for the proteolytic activation, and the change from threonine to Serine at 813 position might be an evolutionary feature adopted by SARS-2-related viruses. This finding deepened the understanding of Spike fusogenicity and could provide a new perspective for exploring Sarbecovirus evolution. Author SummaryThe Spike strain of SARS-CoV-2 has accumulated many mutations during its time in circulation, most of which have occurred in the S1 region, and more specifically in the RBD, in an effort to either improve the viruss affinity for the receptor ACE2 or to enhance its ability to evade the immune system. Mutations in the Spike S2 region have more far-reaching effects than those in the S1 region because it is more conserved across sarbecoviruses. By comparing SARS and SARS2, we found that an important substitution at amino acid position 813 in the S2 region (T813S) disrupts the utilization of TMPRSS2 and can significantly influence viral entry into cells. This discovery deepens our knowledge of S proteins and provides new prospects for tracing the evolution of Sarbecoviruses.

microbiology↗

SEC-TMT facilitates quantitative differential analysis of protein interaction networks

A proteins molecular interactions and post-translational modifications (PTMs), such as phosphorylation, can be co-dependent and reciprocally co-regulate each other. Although this interplay is central for many biological processes, a systematic method to simultaneously study assembly-states and PTMs from the same sample is critically missing. Here, we introduce SEC-MX (Size Exclusion Chromatography fractions MultipleXed), a global quantitative method combining Size Exclusion Chromatography and PTM-enrichment for simultaneous characterization of PTMs and assembly-states. SEC-MX enhances throughput, allows phosphopeptide enrichment, and facilitates quantitative differential comparisons between biological conditions. Applying SEC-MX to HEK293 and HCT116 cells, we generated a proof-of-concept dataset mapping thousands of phosphopeptides and their assembly-states. Our analysis revealed intricate relationships between phosphorylation events and assembly-states and generated testable hypotheses for follow-up studies. Overall, we establish SEC-MX as a valuable tool for exploring protein functions and regulation beyond abundance changes.

molecular biology↗

Abnormal expression of Prolyl oligopeptidase (POP) and its catalytic products Ac-SDKP contributes to the ovarian fibrosis change in Polycystic ovary syndrome (PCOS) mice

Polycystic ovary syndrome (PCOS) is an endocrine disorder and metabolic syndrome. Ovarian fibrosis pathological change in PCOS gradually attracted peoples attention. In this study, we constructed PCOS mice model through dehydroepiandrosterone. Sirius red staining showed that the ovarian tissues in PCOS mice had obvious fibrosis. Prolyl oligopeptidase(POP) is a serine protease and N-acetyl-Seryl-aspartyl-Lysyl-proline (Ac-SDKP) is its catalytic products. Studies show that abnormal expression and activity of POP and Ac-SDKP are closely related to tissue fibrosis. We found that the expression of POP and Ac-SDKP was decreased in ovaries of PCOS mice. Further studies showed that POP and Ac-SDKP promoted the expression of Matrix metalloproteinases 2 (MMP-2) expression and decreased the expression of transforming growth factor beta 1 (TGF-{beta}1) in granulosa cells. Hyperandrogenemia is a typical symptom of PCOS. We found that testosterone induced the low expression of POP and MMP2, and high expression of TGF-{beta}1 in granulosa cells. POP overexpression and Ac-SDKP treatment inhibited the effect of testosterone on TGF-{beta}1 and MMP2 in vitro and ovarian fibrosis in PCOS mice model. In conclusion, PCOS ovarian tissue showed obvious fibrosis. Low expression of POP, Ac-SDKP and changes in fibrotic factors contribute to the ovarian pathological fibrosis induced by androgen.

cell biology↗

Transcriptomic landscape and potential therapeutic targets for human testicular aging revealed by single-cell RNA sequencing

BackgroundTesticular aging is known to cause male age-related fertility decline and hypogonadism, but the underlying molecular mechanisms remain unclear. MethodsWe survey the single-cell transcriptomic landscape of testes from young and old men and examine age-related changes in germline and somatic niche cells. ResultsIn-depth evaluation of the gene expression dynamics of germline cells reveals that disturbance of base-excision repair pathway is a major feature of aging spermatogonial stem cells (SSCs), suggesting that defective DNA repair of SSCs may serve as a potential driver for increased de novo germline mutations with age. Further analysis of aging-associated transcriptional changes shows that stress-related changes and apoptotic signaling pathway accumulate in aged somatic cells. We identify age-related impairment of redox homeostasis in aged Leydig cells and find that pharmacological treatment with antioxidants alleviate this cellular dysfunction of Leydig cells and promote testosterone production. Lastly, our results reveal that decreased pleiotrophin (PTN) signaling is a contributing factor for testicular aging. ConclusionsThese findings provide a comprehensive understanding of the cell-type-specific mechanisms underlying human testicular aging at a single-cell resolution, and suggest potential therapeutic targets that may be leveraged to address age-related male fertility decline and hypogonadism. FundingThis work was supported by the National Key Research and Development Program of China (2018YFA0107200, 2018YFA0801404), the National Natural Science Foundation of China (32130046, 82171564, 82101669, 81871110, 81971759), the Key Research and Development Program of Guangdong Province (2019B020234001), the Natural Science Foundation of Guangdong Province, China (2022A1515010371), the Major Project of Medical Science and Technology Development Research Center of National Health Planning Commission, China (HDSL202001000), the Open Project of NHC Key Laboratory of Male Reproduction and Genetics (Family Planning Research Institute of Guangdong Province) (KF202001), the Guangdong Province Regional Joint Fund-Youth Fund Project (2021A1515110921), the China Postdoctoral Science Foundation (2021M703736).

developmental biology↗

Evolutionary trajectory of organelle-derived nuclear DNAs in the Triticum/Aegilops complex species

Organelle-derived nuclear DNAs, nuclear plastid DNAs (NUPTs) and nuclear mitochondrial DNAs (NUMTs), have been identified in plants. Most, if not all, genes residing in NUPTs/NUMTs (NUPGs/NUMGs) are known to be inactivated and pseudogenized. However, the role of epigenetic control in silencing NUPGs/NUMGs and the dynamic evolution of NUPTs/NUMTs with respect to organismal phylogeny remain barely explored. Based on the available nuclear and organellar genomic resources of the Triticum/Aegilops complex species, we investigated the evolutionary fates of NUPTs/NUMTs in terms of their epigenetic silencing and their dynamic occurrence rates in the nuclear diploid genomes and allopolyploid subgenomes. NUPTs and NUMTs possessed similar genomic atlas, including preferential integration to the transposable element-rich intergenic regions and generating sequence variations in the nuclear genome. The global transcriptional silencing of NUPGs/NUMGs with disrupted and intact open reading frames can be mainly attributed to their repressive chromatin states, namely high levels of DNA methylation and low levels of active histone modifications. Phylogenomic analyses suggested that the species-specific and gradual accumulation of NUPTs/NUMTs accompanied the speciation processes. Moreover, based on further pan-genomic analyses, we found significant subgenomic asymmetry in the NUPT/NUMT occurrence, which accumulated during allopolyploid wheat evolution. Our findings provide novel insights into the dynamic evolutionary fates of organelle-derived nuclear DNA in plants.

genomics↗

DEFECTIVE KERNEL1 (DEK1) regulates cellulose synthesis and affects primary cell wall mechanics

The cell wall is one of the defining features of plants, controlling cell shape, regulating growth dynamics and hydraulic conductivity, as well as mediating plants interactions with both the external and internal environments. Here we report that a putative mechanosensitive Cys-protease DEFECTIVE KERNEL1 (DEK1) interacts with cell wall integrity (CWI) pathways and regulation of cellulose synthesis. Our results indicate that DEK1 is an important regulator of cellulose synthesis in epidermal tissue of Arabidopsis thaliana cotyledons during early post-embryonic development. DEK1 is involved in regulation of cellulose synthase complexes (CSCs) by modifying their biosynthetic properties, possibly through interactions with various cellulose synthase regulatory proteins. Mechanical properties of the primary cell wall are altered in DEK1 modulated lines supporting a role in maintenance of CWI. DEK1 affects stiffness of the cell wall and thickness of the cellulose microfibrils bundles in epidermal cell walls of cotyledons.

plant biology↗

Changes of platelet morphology, ultrastructure and function in patients with acute ischemic stroke based on super-resolution microscopy.

Platelets are important in acute ischemic stroke (AIS) thrombosis. The observation and evaluation of platelet ultrastructure and efficacy of antiplatelet drug in AIS patients is difficult due to microscopic limitations and sensitivity of platelet. The new super-resolution microscope (SIM) can accurately, quickly analyze the platelet ultrastructure and antiplatelet drug in AIS patients. We applied SIM to observe the morphology and ultrastructure of platelets with AIS patients in different state. SIM images were analyzed to specify the dense granules and granules change of platelets in AIS patients. Testing platelet factor 4 (PF4) to reflect platelet releasing function. We observed that platelet activation in AIS patients was greater after stimulation, with granule showing a pattern of parenchymatous masses. SIM images analyzing showed diameter of platelets, average size of granules, area% of granules per field and mean area of granules per platelet in AIS patient were lower than healthy people. Platelet releasing function was suppressed at rest stage and more efficacy release after stimulation. 2MeSamp inhibited parenchymatous masses of granules and reduced PF4 release of platelets after stimulation. According to the results, the structure and function of platelets in AIS patients are indeed altered. Additionally, SIM could be used as a new method to indicates the onset of AIS and assess antiplatelet drugs.

cell biology↗

Reactivating Positive Personality Traits During Sleep Impacts Self-Evaluative Memories

Positive self-view is evident by a bias in favor of positive self-referential processing, as individuals tend to endorse positive characteristics over negative ones when making self-judgments. While research suggests that a positivity bias can contribute to psychological well-being, it remains unclear how to enhance positive self-referential processing. Here, we reported an integrated training procedure that aimed at enhancing individuals positive self-referential processing. Specifically, participants engaged in a cue-approach training task (CAT) during wakefulness where they gave speeded motor responses to positive personality traits. In a subsequent nap, we unobtrusively re-played half of the trained positive traits during participants slow-wave sleep to reactivate memories associated with these positive traits (targeted memory reactivation, TMR). Upon awakening, we found that CAT+TMR enhanced participants positive self-referential processing, as evidenced by faster endorsement of positive traits. Further analysis revealed that this enhancement was associated with specific brainwave patterns during sleep: delta (1-4 Hz) traveling waves moving from posterior to anterior brain regions. These findings demonstrate the potential benefits of integrated wakeful cue-approach training and sleep-based memory reactivation in strengthening positive self-referential processing.

neuroscience↗

Analysis of hormonal regulation of promoter activities of Cannabis sativa prenyltransferase 1 and 4 and salicylic acid mediated regulation of cannabinoid biosynthesis

Cannabis sativa prenyltransferase 4 (CsPT4) and prenyltransferase 1 (CsPT1) have been shown to catalyze the step in the cannabinoid biosynthetic pathway that generates cannabigerolic acid (CBGA), the substrate for the end-point enzymes that generate cannabidiolic acid (CBDA) and tetrahydrocannabinolic acid (THCA). Prior studies from our lab suggest that CBGA production rate-limits the pathway. There is a lack of understanding concerning how important cannabinoid biosynthetic genes are regulated as cannabinoid synthesis increases during female flower development. Both CsPT genes were shown to be highly expressed in flowers. The genes were also found to be present in leaves and roots. GUS staining also detected the promoter activities in leaves of seedlings, and the promoter activities were drastically stronger in the section of the sugar leaves where glandular trichomes are formed. In silico analysis of the two CsPT genes revealed several hormone and transcription factor responsive elements. Dual luciferase assays were conducted to determine whether a hormone could alter the promoter activities of CsPT1 and CsPT4. The results showed that CsPT4 pro was activated following treatment from salicylic acid (SA), gibberellic acid (GA), ethylene, ABA, and cytokinin, while the CsPT1 promoter was activated following SA, ethylene, ABA, and auxin treatment. In parallel studies, a correlation was observed between multiple cannabinoid biosynthetic pathway genes and SA application to the cannabis growing medium, along with a correlation between MeSA floral application and an increase in cannabinoid content. The results from all aspects of this study demonstrated an interaction between certain hormones and cannabinoid synthesis.

plant biology↗

Formation process of the twinning β-form anhydrous guanine platelets in the scallop eyes

Square shaped twinning guanine microplates with high symmetry are assembled into highly ordered layered patterns and function as image-forming mirrors in the scallop eyes. However, the formation process and biomineralization mechanism of twinning guanine microplatelets are still unclear. Herein, the eyes of juvenile Yesso scallops were investigated to understand the formation mechanism of the twinning {beta}-form anhydrous guanine ({beta}-AG) microplatelets exposing (100) plane. We find form anhydrous guanine (-AG) and single-crystal {beta}-AG nanoplatelets in the very early stage of the eyes of the juvenile scallops, while the -AG was supposed to be formed via amorphous guanine during the sample preparation process. Besides {beta}-AG and -AG, amorphous guanine was found in the eyes of juvenile scallops with size of 2.5 mm according to the Raman spectra. A formation mechanism was proposed for the biogenic twinning guanine platelets. Firstly, amorphous guanine is formed as an intermediate phase, which transforms into single crystalline {beta}-AG nanoplatelets, or, dissolve and recrystallize to single crystalline {beta}-AG nanoplatelets. Then, a second layer of {beta}-AG forms on the top of the original single crystalline {beta}-AG nanoplatelets, forming twinning {beta}-AG nanoplatelets with two c axes with a certain angle, 83{degrees} or 14{degrees}. Each layer of the {beta}-AG nanoplatelets is calculated to be about 14 {+/-} 2 nm. This is the first time to report the formation mechanism of biogenic twinning {beta}-AG microplatelets. Uncovering the formation mechanism of twinning platelets of organic crystals may shed light on the formation of functional synthetic twinning organic crystals in the laboratories.

biochemistry↗

Molecular Mechanism for Bacterial Degradation of Plant Hormone Auxin

Plant-associated bacteria play important regulatory roles in modulating plant hormone auxin levels, affecting the growth and yields of crops. A conserved auxin-degradation (adg) operon was recently identified in the Variovorax genomes, which is responsible for root growth inhibition (RGI) reversion, promoting rhizosphere colonization and root growth. However, the molecular mechanism underlying auxin degradation by Variovorax remains unclear. Here, we systematically screened Variovorax adg operon products and identified two proteins, AdgB and AdgI, that directly associate with auxin indole-3-acetic acid (IAA). Further biochemical and structural studies revealed that AdgB is a highly IAA-specific ABC transporter solute binding protein, likely involved in IAA uptake. AdgI interacts with AdgH to form a functional Rieske non-heme dioxygenase, which works in concert with a FMN-type reductase encoded by gene adgJ to transform IAA into the biologically inactive 2-oxindole-3-acetic acid (oxIAA), representing a new bacterial pathway for IAA inactivation/degradation. Importantly, incorporation of a minimum set of adgH/I/J genes could enable IAA degradation by E. coli, suggesting a promising strategy for repurposing the adg operon for IAA regulation. Together, our study identifies the key components and underlying mechanisms involved in IAA transformation by Variovorax and brings new insights into the bacterial turnover of plant hormones, which would provide the basis for potential applications in rhizosphere optimization and ecological agriculture.

microbiology↗

An alternative malonyl-CoA producing pathway in nature

Malonyl-CoA is a key metabolic intermediate for biosynthesis of diverse cellular molecules and natural products. Carboxylation of acetyl-CoA is almost the unique pathway for malonly-CoA biosynthesis. Biotechnological production of numerous value-added malonyl-CoA-derived chemicals require high intracellular supply of malonyl-CoA. However, because of the central role of acetyl-CoA in primary metabolism, it is difficult to develop flexible strategies to balance malonyl-CoA supply with other cellular metabolism. Here we find that there is a natural alternative malonyl-CoA-producing pathway, in which the key reaction is catalyzed by an -keto acid dehydrogenase complex BkdFGH from Streptomyces avermitilis. This dehydrogenase complex could efficiently catalyze biosynthesis of malonyl-CoA from oxaloacetate in addition to recognizing its native substrate branched-chain -keto acid. Oxaloacetate dehydrogenase (OADH) was shown to play important physiological roles during the regulation of biosynthesis of native malonyl-CoA-derived polyketides in Streptomyces. Furthermore, the oxlaocetate dehydrogenation reaction is thermodynamically superior to acetyl-CoA carboxylation and enable efficient bioproduction of diverse malonyl-CoA-derived chemicals in engineering Escherichia coli. This novel malonyl-CoA source thus has great potential in the biotechnological field.

microbiology↗

Temporary knockdown of p53 during focal limb irradiation increases the development of sarcomas

BackgroundApproximately half of all cancer patients receive radiotherapy and, as cancer survivorship rates increase with more effective therapies, the very low rate of radiation-associated sarcomas is rising. Radiation-associated sarcomas are life-threatening cancers, and radiation exposure is a primary risk factor for sarcoma development. During radiotherapy or other genotoxic cancer therapy for p53 mutant cancers, pharmacological inhibition of p53 has been proposed to ameliorate acute injury of normal tissues. However, enhancing the survival of normal cells that sustain DNA damage by temporarily inhibiting p53 has the potential to increase the risk of cancer development. Here, we use in vivo shRNA technology to examine the consequences of temporarily reducing p53 expression on radiation-induced sarcoma development. MethodsWe utilized a mouse model of radiation-induced sarcoma where mice express a doxycycline (dox)-inducible p53 shRNA to temporarily and reversibly reduce p53 expression. Mice were placed on a dox diet 10 days prior to receiving 30 or 40 Gy hind limb irradiation in a single fraction and then returned to normal chow. Mice were examined weekly for sarcoma development and scored for radiation-induced normal tissue injuries. Radiation-induced sarcomas were harvested and subjected to RNA sequencing. ResultsFollowing single high-dose irradiation, 21% of temporary p53 knockdown animals developed a sarcoma in the radiation field compared to 2% of control animals. Mice with more severe acute injuries in the first 3 months after irradiation had a significantly increased risk of developing late persistent wounds in the soft tissue and bone. Chronic radiation-induced wounds were associated with sarcomagenesis. Examination of muscle stem cells by flow cytometry following hind limb irradiation indicated p53 knockdown preserves muscle stem cells in the irradiated limb, supporting the notion that temporary p53 knockdown at the time of irradiation reduces death of cells with DNA damage which may then persist to develop into a sarcoma. We performed RNA sequencing on 16 radiation-induced sarcomas compared to normal muscle controls. Gene set enrichment analysis revealed upregulation in the sarcomas of genes related to translation, epithelial mesenchymal transition (EMT), inflammation, and the cell cycle versus downregulation of genes related to myogenesis and tumor metabolism. Furthermore, genes with increased copy number such as Met and Cdk4 were overexpressed in tumors. ConclusionsTemporary reduction of p53 during high-dose irradiation increases late effects including tissue injuries and sarcoma development.

cancer biology↗

Quantitative profiling of pseudouridylation landscape in the human transcriptome

Pseudouridine ({Psi}) is an abundant post-transcriptional RNA modification in ncRNA and mRNA. However, transcriptome-wide measurement of individual {Psi} sites remains unaddressed. Here, we develop "PRAISE", via selective chemical labeling of {Psi} by bisulfite to induce nucleotide deletion signature during reverse transcription, to realize quantitative assessment of the {Psi} landscape in the human transcriptome. Unlike traditional RNA/DNA bisulfite treatment, our approach is based on quaternary base mapping and revealed a ~10% median modification level for 2,714 confident {Psi} sites in HEK293T cells. By perturbing pseudouridine synthases, we obtained differential mRNA targets of PUS1, PUS7 and TRUB1, with TRUB1 mRNA targets showing the highest modification stoichiometry. In addition, we identified and quantified known and novel {Psi} sites in mitochondrial mRNA, catalyzed by a mitochondria-localized isoform of PUS1. Collectively, we provide a sensitive and convenient method to measure transcriptome-wide {Psi}; we envision this quantitative approach would facilitate emerging efforts to elucidate the function and mechanism of mRNA pseudouridylation.

molecular biology↗