Biology subjects

Peng, N.

Publications and source records attributed to Peng, N..

10 recordsLinked to original sources

bioRxiv2026

Development and Validation of the Transcranial Magnetic Stimulation Reporting Assessment Tool(TMS-RAT)

HighlightsO_LIWe introduce the TMS-RAT, a reporting (assessment) tool for TMS studies C_LIO_LIDeveloped within a community-informed, iterative process rating 333 TMS studies C_LIO_LIEmpirically evaluated for usability, inter-rater, and test-retest reliability C_LIO_LIA validated subset enables reliable retrospective assessment of reporting C_LIO_LIThe modular structure enables use across a wide range of TMS study designs C_LI BackgroundA standardised tool for comprehensive reporting can improve transparency, support consistent documentation, and enable comparison across transcranial magnetic stimulation (TMS) studies. The most used reporting checklist lacks clear definitions of full reporting and was not initially evaluated for usability or inter-rater reliability. A scoping review of studies using this checklist shows that its items are reported only 50% of the time, suggesting that method descriptions are often incomplete. MethodsWe developed the TMS Reporting Assessment Tool (TMS-RAT), a comprehensive reporting framework that provides clear definitions and examples for its items, covering a wide range of TMS protocols. We tested the usability and reliability of the TMS-RAT by rating all studies published between 1991 and 2025 using afferent conditioning (n = 333), a protocol encompassing many reporting categories identified during tool development. Seventeen independent raters contributed across three development phases, a validation phase, and a retest phase, with naive raters introduced in each phase. Iterative refinements of the tool were informed by inter-rater reliability, qualitative rater feedback, and consultation with external TMS experts. ResultsWe present two versions of the tool: the 72-item TMS-RAT v1.0, designed to guide comprehensive reporting, and the TMS-RAT v1.1, a subset of 50 items with the highest inter-rater (overall AC1 = 0.78, range = [0.60-0.99]) and test-retest reliability (overall AC1 = 0.82, range = [0.65-1.0]), intended for retrospective evaluation of reporting in systematic reviews, meta-analyses. ConclusionThe TMS-RAT is a comprehensive, reliable tool that seeks to improve transparency and reproducibility in TMS research.

neuroscience↗

bioRxiv2025

Aberrant 5-Methylcytosine tRNA Modification Disrupts Proteostasis and Exacerbates Age-Related Osteoporosis

Age-related protein aggregation drives senile osteoporosis. Aberrant tRNA modifications exacerbate the progression, yet mechanisms linking these to bone loss remain unclear. In this study, we identify Nsun2 (m5C methyltransferase) as key regulator: age-dependent Nsun2 downregulation in BMSCs reduces mC levels of tRNAs, destabilizing tRNAs and impairing translation efficiency of specific transcripts. This directly disrupts the protein synthesis of molecular chaperone and pro-osteogenic factors, accelerating misfolded protein aggregates and activating unfolded protein response, inducing BMSCs senescence and impairing osteogenesis. Mice specifically depleted of Nsun2 exhibited reduced bone mass, whereas mice overexpressing Nsun2 alleviated age-associated bone loss. Notably, the exacerbated protein aggregation and bone mass loss in Nsun2-deficient mice were ameliorated following treatment with the molecular chaperone activator-ML346. Remarkably, ML346 administration proved sufficient to reverse age-related functional deficits in aged mice. Overall, our findings demonstrate that aberrant tRNA-m5C modification alters protein synthesis and induces proteostasis collapse, which constitute a novel contributor to the pathogenesis of senile osteoporosis. Additionally, reduction of protein aggregation through the activation of molecular chaperones presents a promising therapeutic strategy for this disease.

cell biology↗

bioRxiv2025

Inhibition of interferon signalling improves rabbit calicivirus replication in biliary organoid cultures

The Rabbit haemorrhagic disease virus (RHDV) was discovered 40 years ago. This highly pathogenic virus threatens the integrity of ecosystems in the European rabbits native range, while in Australia, it is used as a biocontrol tool to manage overabundant populations of feral rabbits (Oryctolagus cuniculus). Little is known about the life cycle of this virus due to the absence of a reliable cell culture system. In 2023, we developed a rabbit liver-derived organoid cell culture system that supports RHDV replication but is unable to sustain serial passaging in culture. Here, we report that the interferon signalling pathway inhibitor Ruxolitinib increases virus replication in organoid-derived monolayer cells and, for the first time, enables the serial passaging of RHDV in cell culture. Four consecutive passages were achieved with viral titres reaching the concentration of the initial virus stock as measured by RT-qPCR. Immunofluorescence analysis showed that more cells are infected in the presence of Ruxolitinib. Furthermore, we noted that cells grew faster and formed healthier monolayers in the presence of the interferon inhibitor. To determine the cellular composition of the monolayers, we used single-cell RNA-sequencing, revealing that our organoids consist largely of RHDV-permissive cholangiocytes. IMPORTANCEIn this work, we describe the use of interferon inhibitor to enhance the permissiveness of our recently developed rabbit liver-derived organoid cell culture to RHDV infection. Interferon inhibitors offer a simple and cost-effective approach to increase the replication of difficult-to-grow viruses in culture systems that are not interferon-deficient.

microbiology↗

bioRxiv2023

MolPhase: An Advanced Phase Separation Predictor and an Investigation of Phytobacterial Effector in Plant

We introduce MolPhase (http://molphase.sbs.ntu.edu.sg/), an advanced protein phase separation (PS) prediction algorithm that improves accuracy and reliability by utilizing diverse physicochemical features and extensive experimental datasets. MolPhase applies a user-friendly interface to compare distinct biophysical features side-by-side along protein sequences. By additional comparison with structural predictions, MolPhase enables efficient predictions of new phase-separating proteins and guides hypothesis generation and experimental design. Key contributing factors underlying MolPhase include pi-pi interaction, disorder, and prion-like domain. As an example, MolPhase finds that phytobacterial type III effectors (T3Es) are highly prone to homotypic PS, which was experimentally validated in vitro biochemically and in vivo in plants, mimicking their injection and accumulation in the host during microbial infection. In addition, the phase-separation of T3Es were evolved both in vivo and in vitro, suggesting their determinative scaffolding function, though there is a difference in material properties, implying a difference in homotypic and heterotypic macromolecular condensation. Robust integration of MolPhases effective prediction and experimental validation exhibit the potential to evaluate and explore how biomolecule PS functions in biological systems.

biophysics↗

bioRxiv2023

Reprogramming an RNA-guided archaeal endonuclease for genome editing

RNA-guided endonucleases developed for genome editing have been identified exclusively from bacteria until now. Recently, the RNA-guided TnpB endonucleases encoded by bacterial IS200/IS605 and IS607 families have been reprogrammed for genome editing. In fact, these insertion sequence families are also widely distributed in archaea. However, whether archaea-derived TnpB could be used for genome editing remains unclear. Here, we found that TnpB derived from Sulfolobus islandicus REY15A (SisTnpB1) was an Mg2+/Mn2+-dependent RNA-guided endonuclease, and it could cleave DNA next to the 5 TTTAA transposon-associated motif (TAM) to generate double-stranded DNA breaks at the 3-end. We also found that SisTnpB1 was active at 37-85{degrees}C and tolerant to variations of TAM. Moreover, SisTnpB1 could be reprogrammed to edit bacterial cells with an editing efficiency of 100% at 37{degrees}C or 45{degrees}C. In summary, our finding expanded the resources of RNA-guided endonucleases for genome editing, especially those derived from archaea, the third domain of life.

molecular biology↗

bioRxiv2021

Zika virus noncoding RNA cooperates with the viral protein NS5 to inhibit STAT1 phosphorylation and facilitate viral pathogenesis

Zika virus (ZIKV) is a re-emerging pathogenic flavivirus, which causes microcephaly in infants and poses a continuing threat to public health. ZIKV, like all other flaviviruses, produces highly abundant noncoding RNA known as subgenomic flaviviral RNA (sfRNA). Herein we utilized wild-type and mutant ZIKV defective in production of sfRNA to elucidate for the first time how production of sfRNA affects all aspects of ZIKV pathogenesis. We found that in mouse pregnancy model of infection sfRNA is required for trans-placental dissemination of ZIKV and subsequent infection of fetal brain. Using human brain organoids, we showed that sfRNA promotes apoptosis of neural progenitor cells leading to profound cytopathicity and disintegration of organoids. We also found by transcriptome profiling and gene network analysis that in infected human placental cells sfRNA inhibits multiple antiviral pathways and promotes apoptosis with STAT1 identified as a key shared factor linking these two interconnected sfRNA activities. We further showed for the first time that sfRNA inhibits phosphorylation and nuclear translocation of STAT1 by a novel mechanism which involves binding to and stabilizing viral protein NS5. This allows accumulation of NS5 at the levels required for efficient inhibition of STAT1 phosphorylation. Thus, we elucidated the molecular mechanism by which ZIKV sfRNA exerts its functions in vertebrate hosts and discovered a co-operation between viral noncoding RNA and a viral protein as a novel strategy employed by viruses to counteract antiviral responses.

microbiology↗

bioRxiv2020

Aminoglycoside antibiotics inhibit mycobacteriophage infection

Antibiotic resistance is becoming the biggest current threat to global health. At the same time, phage therapy is witnessing a return of interest. The therapeutic use of bacteriophages, that infect and kill bacteria, is well suited to be a good strategy to combat antibiotic resistance. Furthermore, bacteriophages are increasingly used in combination with standard antibiotics against the drug-resistant pathogens. Interestingly, we found that the engineered mycobacteriophage phAE159 and natural phage D29 can not infect the Mycobacterium tuberculosis in the presence of kanamycin, hygromycin or streptomycin, but there is no effect on the phage infection in the presence of spectinomycin. Based on a series of studies and structural analysis of the above four aminoglycoside antibiotics, we can speculate as to the mechanism by which amino sugar group of aminoglycoside was able to selectively inhibit mycobacteriophage DNA replication. This is a rare discovery that broad-spectrum antibiotics inhibit phage infection. We envisioned that this study will provide guidance for people to combine phage and antibiotics to treat M. tuberculosis.Competing Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗

bioRxiv2020

Reprogramming the endogenous type III-A CRISPR-Cas system for genome editing, RNA interference and CRISPRi screening in Mycobacterium tuberculosis

Mycobacterium tuberculosis (M.tb) causes the current leading infectious disease. Examination of the functional genomics of M.tb and development of drugs and vaccines are hampered by the complicated and time-consuming genetic manipulation techniques for M.tb. Here, we reprogrammed M.tb endogenous type III-A CRISPR-Cas10 system for simple and efficient gene editing, RNA interference and screening via simple delivery of a plasmid harboring a mini-CRISPR array, thereby avoiding the introduction of exogenous proteins and minimizing proteotoxicity. We demonstrated that M.tb genes were efficiently and specifically knocked-in/out by this system, which was confirmed by whole-genome sequencing. This system was further employed for single and simultaneous multiple-gene RNA interference. Moreover, we successfully applied this system for genome-wide CRISPR interference screening to identify the in-vitro and intracellular growth-regulating genes. This system can be extensively used to explore the functional genomics of M.tb and facilitate the development of new anti-Mycobacterial drugs and vaccines. SummaryTuberculosis caused by Mycobacterium tuberculosis (M.tb) is the current leading infectious disease affecting more than ten million people annually. To dissect the functional genomics and understand its virulence, persistence, and antibiotics resistance, a powerful genome editing tool and high-throughput screening methods are desperately wanted. Our study developed an efficient and a robust tool for genome editing and RNA interference in M.tb using its endogenous CRISPR cas10 system. Moreover, the system has been successfully applied for genome-wide CRISPR interference screening. This tool could be employed to explore the functional genomics of M.tb and facilitate the development of anti-M.tb drugs and vaccines.

microbiology↗

bioRxiv2019

Primed acquisition and microhomology-mediated end-joining cooperate to confer specific CRISPR immunity against invasive genetic elements

CRISPR-Cas systems provide archaea and bacteria with adaptive immunity against invasive genetic elements through acquisition of invader-derived spacers. De novo spacer acquisition generally adapts spacers from both invaders and hosts with a bias towards invaders, while primed acquisition shows higher specificity to adapt spacers from invaders. Here, in Zymomonas mobilis subtype I-F system, primed acquisition showed much higher efficiency than de novo acquisition in this system. However, both routes recognised a large proportion of protospacers with the less conserved 5-end CC PAM. Moreover, primed acquisition showed a preference towards protospacers located at the opposite strand and 3 direction of the priming protospacer sites, differing from the canonical subtype I-F system. Further, self-spacers were adapted at a higher frequency during de novo spacer acquisition, probably leading to self-interference. Importantly, this species employed microhomology-mediated end-joining (MMEJ) for repair of host DNA breaks guided by self-targeted spacers, and overexpression of a host NAD+-dependent Ligase-A significantly increased the repair efficiency. In summary, our findings demonstrate that Z. mobilis uses primed acquisition for higher specific uptake of invader DNA and employs MMEJ to repair host DNA breaks guided by self-targeted spacers, showing specific immunity against invasive genetic elements.

genetics↗

bioRxiv2018

Cas4 nucleases can effect specific integration of CRISPR spacers

CRISPR-Cas systems incorporate short DNA fragments from invasive genetic elements into host CRISPR arrays in order to generate host immunity.Recently, we demonstrated that the Csa3a regulator protein triggers CCN PAM-dependent CRISPR spacer acquisition in the subtype I-A CRISPR-Cas system of Sulfolobus islandicus. However, the mechanisms underlying specific protospacer selection and spacer insertion remained unclear. Here, we demonstrate that two Cas4 family proteins (Cas4 and Csa1) have essential roles (a) in recognizing the 5 PAM and 3 nucleotide motif of protospacers and (b) in determining both the spacer length and its orientation. Furthermore, we identify uncovered amino acid residues of the Cas4 proteins that facilitate these functions. Overexpression of the Cas4 and Csa1 proteins, and also of an archaeal virus-encoded Cas4 protein, resulted in strongly reduced adaptation efficiency and the formers yielded a high incidence of PAM-dependent atypical spacer integration, or of PAM-independent spacer integration. We further demonstrated that, in the plasmid challenging experiments, overexpressed Cas4-mediated defective spacer acquisition, in turn, potentially enabled targeted DNA to escape subtype I-A CRISPR-Cas interference. In summary, these results define the specific involvement of diverse Cas4 proteins in in vivo CRISPR spacer acquisition. Furthermore, we provide support for an anti-CRISPR role for virus-encoded Cas4 proteins that involves compromising CRISPR-Cas interference activity by hindering spacer acquisition.\n\nImportanceThe Cas4 family endonuclease is an essential component of the adaptation module in many variants of CRISPR-Cas adaptive immunity systems. The Crenarchaeota Sulfolobus islandicus REY15A encodes two cas4 genes (cas4 and csa1) linked to the CRISPR arrays. Here, we demonstrate that Cas4 and Csa1 are essential to CRISPR spacer acquisition in this organism. Both proteins specify the upstream and downstream conserved nucleotide motifs of the protospacers and define the spacer length and orientation in the acquisition process. Conserved amino acid residues, in addition to the recently reported, were identified to be important for above functions. More importantly, overexpression of the Sulfolobus viral Cas4 abolished spacer acquisition, providing support for an anti-CRISPR role for virus-encoded Cas4 proteins that inhibit spacer acquisition.

molecular biology↗