bluuebunny/crossref_metadata_2025_split huggingface dataset:
Application of multiple sgRNAs boosts efficiency of CRISPR/Cas9-mediated gene targeting in Arabidopsis
Authors: Li Jing , Kong Dali , Ke Yongping , Zeng Wenjie , Miki Daisuke 
Published: January 2024

Abstract
Background
Precise gene targeting (GT) is a powerful tool for heritable precision genome engineering, enabling knock-in or replacement of the endogenous sequence via homologous recombination. We recently established a CRISPR/Cas9-mediated approach for heritable GT in Arabidopsis thaliana (Arabidopsis) and rice and reported that the double-strand breaks (DSBs) frequency of Cas9 influences the GT efficiency. However, the relationship between DSBs and GT at the same locus was not examined. Furthermore, it has never been investigated whether an increase in the number of copies of sgRNAs or the use of multiple sgRNAs would improve the efficiency of GT.

Results
Here, we achieved precise GT at endogenous loci Embryo Defective 2410 (EMB2410) and Repressor of Silencing 1 (ROS1) using the sequential transformation strategy and the combination of sgRNAs. We show that increasing of sgRNAs copy number elevates both DSBs and GT efficiency. On the other hand, application of multiple sgRNAs does not always enhance GT efficiency. Our results also suggested that some inefficient sgRNAs would play a role as a helper to facilitate other sgRNAs DSBs activity.

Conclusions
The results of this study clearly show that DSB efficiency, rather than mutation pattern, is one of the most important key factors determining GT efficiency. This study provides new insights into the relationship between sgRNAs, DSBs, and GTs and the molecular mechanisms of CRISPR/Cas9-mediated GTs in plants.
The accumulation of modular serine protease mediated by a novel circRNA sponging miRNA increases Aedes aegypti immunity to fungus
Authors: Lu Tengfei , Ji Yannan , Chang Mengmeng , Zhang Xiaoming , Wang Yanhong , Zou Zhen 
Published: January 2024

Abstract
Background
Mosquitoes transmit many infectious diseases that affect human health. The fungus Beauveria bassiana is a biological pesticide that is pathogenic to mosquitoes but harmless to the environment.

Results
We found a microRNA (miRNA) that can modulate the antifungal immunity of Aedes aegypti by inhibiting its cognate serine protease. Fungal infection can induce the expression of modular serine protease (ModSP), and ModSP knockdown mosquitoes were more sensitive to B. bassiana infection. The novel miRNA-novel-53 is linked to antifungal immune response and was greatly diminished in infected mosquitoes. The miRNA-novel-53 could bind to the coding sequences of ModSP and impede its expression. Double fluorescence in situ hybridization (FISH) showed that this inhibition occurred in the cytoplasm. The amount of miRNA-novel-53 increased after miRNA agomir injection. This resulted in a significant decrease in ModSP transcript and a significant increase in mortality after fungal infection. An opposite effect was produced after antagomir injection. The miRNA-novel-53 was also knocked out using CRISPR-Cas9, which increased mosquito resistance to the fungus B. bassiana. Moreover, mosquito novel-circ-930 can affect ModSP mRNA by interacting with miRNA-novel-53 during transfection with siRNA or overexpression plasmid.

Conclusions
Novel-circ-930 affects the expression level of ModSP by a novel-circ-930/miRNA-novel-53/ModSP mechanism to modulate antifungal immunity, revealing new information on innate immunity in insects.
IGT/LAZY genes are differentially influenced by light and required for light-induced change to organ angle
Authors: Waite Jessica Marie , Dardick Christopher 
Published: January 2024

Abstract
Background
Plants adjust their growth orientations primarily in response to light and gravity signals. Considering that the gravity vector is fixed and the angle of light incidence is constantly changing, plants must somehow integrate these signals to establish organ orientation, commonly referred to as gravitropic set-point angle (GSA). The IGT gene family contains known regulators of GSA, including the gene clades LAZY, DEEPER ROOTING (DRO), and TILLER ANGLE CONTROL (TAC).

Results
Here, we investigated the influence of light on different aspects of GSA phenotypes in LAZY and DRO mutants, as well as the influence of known light signaling pathways on IGT gene expression. Phenotypic analysis revealed that LAZY and DRO genes are collectively required for changes in the angle of shoot branch tip and root growth in response to light. Single lazy1 mutant branch tips turn upward in the absence of light and in low light, similar to wild-type, and mimic triple and quadruple IGT mutants in constant light and high-light conditions, while triple and quadruple IGT/LAZY mutants show little to no response to changing light regimes. Further, the expression of IGT/LAZY genes is differentially influenced by daylength, circadian clock, and light signaling.

Conclusions
Collectively, the data show that differential expression of LAZY and DRO genes are required to enable plants to alter organ angles in response to light-mediated signals.
A chromosome-level genome for the nudibranch gastropod Berghia stephanieae helps parse clade-specific gene expression in novel and conserved phenotypes
Authors: Goodheart Jessica A. , Rio Robin A. , Taraporevala Neville F. , Fiorenza Rose A. , Barnes Seth R. , Morrill Kevin , Jacob Mark Allan C. , Whitesel Carl , Masterson Park , Batzel Grant O. , Johnston Hereroa T. , Ramirez M. Desmond , Katz Paul S. , Lyons Deirdre C. 
Published: January 2024

Abstract
Background
How novel phenotypes originate from conserved genes, processes, and tissues remains a major question in biology. Research that sets out to answer this question often focuses on the conserved genes and processes involved, an approach that explicitly excludes the impact of genetic elements that may be classified as clade-specific, even though many of these genes are known to be important for many novel, or clade-restricted, phenotypes. This is especially true for understudied phyla such as mollusks, where limited genomic and functional biology resources for members of this phylum have long hindered assessments of genetic homology and function. To address this gap, we constructed a chromosome-level genome for the gastropod Berghia stephanieae (Valdés, 2005) to investigate the expression of clade-specific genes across both novel and conserved tissue types in this species.

Results
The final assembled and filtered Berghia genome is comparable to other high-quality mollusk genomes in terms of size (1.05 Gb) and number of predicted genes (24,960 genes) and is highly contiguous. The proportion of upregulated, clade-specific genes varied across tissues, but with no clear trend between the proportion of clade-specific genes and the novelty of the tissue. However, more complex tissue like the brain had the highest total number of upregulated, clade-specific genes, though the ratio of upregulated clade-specific genes to the total number of upregulated genes was low.

Conclusions
Our results, when combined with previous research on the impact of novel genes on phenotypic evolution, highlight the fact that the complexity of the novel tissue or behavior, the type of novelty, and the developmental timing of evolutionary modifications will all influence how novel and conserved genes interact to generate diversity.
Refining and illuminating acetogenic Eubacterium strains for reclassification and metabolic engineering
Authors: Flaiz Maximilian , Poehlein Anja , Wilhelm Wiebke , Mook Alexander , Daniel Rolf , Dürre Peter , Bengelsdorf Frank R. 
Published: January 2024

Abstract
Background
The genus Eubacterium is quite diverse and includes several acetogenic strains capable of fermenting C1-substrates into valuable products. Especially, Eubacterium limosum and closely related strains attract attention not only for their capability to ferment C1 gases and liquids, but also due to their ability to produce butyrate. Apart from its well-elucidated metabolism, E. limosum is also genetically accessible, which makes it an interesting candidate to be an industrial biocatalyst.

Results
In this study, we examined genomic, phylogenetic, and physiologic features of E. limosum and the closest related species E. callanderi as well as E. maltosivorans. We sequenced the genomes of the six Eubacterium strains ‘FD’ (DSM 3662T), ‘Marburg’ (DSM 3468), ‘2A’ (DSM 2593), ‘11A’ (DSM 2594), ‘G14’ (DSM 107592), and ‘32’ (DSM 20517) and subsequently compared these with previously available genomes of the E. limosum type strain (DSM 20543T) as well as the strains ‘B2’, ‘KIST612’, ‘YI’ (DSM 105863T), and ‘SA11’. This comparison revealed a close relationship between all eleven Eubacterium strains, forming three distinct clades: E. limosum, E. callanderi, and E. maltosivorans. Moreover, we identified the gene clusters responsible for methanol utilization as well as genes mediating chain elongation in all analyzed strains. Subsequent growth experiments revealed that strains of all three clades can convert methanol and produce acetate, butyrate, and hexanoate via reverse β-oxidation. Additionally, we used a harmonized electroporation protocol and successfully transformed eight of these Eubacterium strains to enable recombinant plasmid-based expression of the gene encoding the fluorescence-activating and absorption shifting tag (FAST). Engineered Eubacterium strains were verified regarding their FAST-mediated fluorescence at a single-cell level using a flow cytometry approach. Eventually, strains ‘FD’ (DSM 3662T), ‘2A’ (DSM 2593), ‘11A’ (DSM 2594), and ‘32’ (DSM 20517) were genetically engineered for the first time.

Conclusion
Strains of E. limosum, E. callanderi, and E. maltosivorans are outstanding candidates as biocatalysts for anaerobic C1-substrate conversion into valuable biocommodities. A large variety of strains is genetically accessible using a harmonized electroporation protocol, and FAST can serve as a reliable fluorescent reporter protein to characterize genetically engineered cells. In total eleven strains have been assigned to distinct clades, providing a clear and updated classification. Thus, the description of respective Eubacterium species has been emended, improved, aligned, and is requested to be implemented in respective databases.
Asymptomatic Plasmodium falciparum carriage at the end of the dry season is associated with subsequent infection and clinical malaria in Eastern Gambia
Authors: Fogang Balotin , Lellouche Lionel , Ceesay Sukai , Drammeh Sainabou , Jaiteh Fatou K. , Guery Marc-Antoine , Landier Jordi , Haanappel Cynthia P. , Froberg Janeri , Conway David , D’Alessandro Umberto , Bousema Teun , Claessens Antoine 
Published: January 2024

Abstract
Background
Chronic carriage of asymptomatic low-density Plasmodium falciparum parasitaemia in the dry season may support maintenance of acquired immunity that protects against clinical malaria. However, the relationship between chronic low-density infections and subsequent risk of clinical malaria episodes remains unclear.

Methods
In a 2-years study (December 2014 to December 2016) in eastern Gambia, nine cross-sectional surveys using molecular parasite detection were performed in the dry and wet season. During the 2016 malaria transmission season, passive case detection identified episodes of clinical malaria.

Results
Among the 5256 samples collected, 444 (8.4%) were positive for P. falciparum. A multivariate model identified village of residence, male sex, age ≥ 5 years old, anaemia, and fever as independent factors associated with P. falciparum parasite carriage. Infections did not cluster over time within the same households or recurred among neighbouring households. Asymptomatic parasite carriage at the end of dry season was associated with a higher risk of infection (Hazard Ratio, HR = 3.0, p < 0.0001) and clinical malaria (HR = 1.561, p = 0.057) during the following transmission season. Age and village of residence were additional predictors of infection and clinical malaria during the transmission season.

Conclusion
Chronic parasite carriage during the dry season is associated with an increased risk of malaria infection and clinical malaria. It is unclear whether this is due to environmental exposure or to other factors.
Employment conditions and mental health of overseas female migrant domestic workers in Hong Kong: a parallel mediation analysis
Authors: Sumerlin Timothy S. , Kim Jean H. , Hui Alvin Yik-Kiu , Chan Dicken , Liao Tim , Padmadas Sabu , Fong Eric , Chung Roger Y. 
Published: January 2024

Abstract
Background
Female migrant domestic workers (MDW), approximately 8.5 million globally, often live in their employer’s home under vulnerable conditions. In Hong Kong, MDWs currently comprise 5% of the population. This study was conducted to assess the association between employment conditions and mental health, and the mediating roles stress and job satisfaction have, among female MDWs in Hong Kong.

Methods
Participants completed an online cross-sectional survey. A total of 1,965 survey were collected between August 2020 and August 2021. Questions in the survey were related to MDWs background information, employment conditions, stress, job satisfaction, and two mental health outcomes: anxiety and depression. An employment conditions score was created to assess the cumulative effect poor employment conditions had on mental health. A multicategorical parallel mediation analysis was used to assess the direct effect employment conditions have on mental health and the indirect effects through stress and job satisfaction.

Results
Overall, 17.7% of MDWs were reported to be suffering from anxiety and 30.8% from depression. An increase in poor employment conditions was statistically associated with an increase in both outcomes, while stress levels and job satisfaction mediated this association.

Conclusions
The findings call for increased scrutiny of employment conditions and mental well-being of MDWs.
Clinical evaluation of augmented reality-based 3D navigation system for brachial plexus tumor surgery
Authors: Zhao Xuanyu , Zhao Huali , Zheng Wanling , Gohritz Andreas , Shen Yundong , Xu Wendong 
Published: January 2024

AbstractBackgroundAugmented reality (AR), a form of 3D imaging technology, has been preliminarily applied in tumor surgery of the head and spine, both are rigid bodies. However, there is a lack of research evaluating the clinical value of AR in tumor surgery of the brachial plexus, a non-rigid body, where the anatomical position varies with patient posture.MethodsPrior to surgery in 8 patients diagnosed with brachial plexus tumors, conventional MRI scans were performed to obtain conventional 2D MRI images. The MRI data were then differentiated automatically and converted into AR-based 3D models. After point-to-point relocation and registration, the 3D models were projected onto the patient’s body using a head-mounted display for navigation. To evaluate the clinical value of AR-based 3D models compared to the conventional 2D MRI images, 2 senior hand surgeons completed questionnaires on the evaluation of anatomical structures (tumor, arteries, veins, nerves, bones, and muscles), ranging from 1 (strongly disagree) to 5 (strongly agree).ResultsSurgeons rated AR-based 3D models as superior to conventional MRI images for all anatomical structures, including tumors. Furthermore, AR-based 3D models were preferred for preoperative planning and intraoperative navigation, demonstrating their added value. The mean positional error between the 3D models and intraoperative findings was approximately 1 cm.ConclusionsThis study evaluated, for the first time, the clinical value of an AR-based 3D navigation system in preoperative planning and intraoperative navigation for brachial plexus tumor surgery. By providing more direct spatial visualization, compared with conventional 2D MRI images, this 3D navigation system significantly improved the clinical accuracy and safety of tumor surgery in non-rigid bodies.
Transformation to small cell lung cancer is irrespective of EGFR and accelerated by SMAD4-mediated ASCL1 transcription independently of RB1 in non-small cell lung cancer
Authors: Ding Xi , Shi Min-xing , Liu Di , Cao Jing-xue , Zhang Kai-xuan , Zhang Run-dong , Zhang Li-ping , Ai Kai-xing , Su Bo , Zhang Jie 
Published: January 2024

Abstract
Objectives
Histological transformation to small cell lung cancer (SCLC) has been identified as a mechanism of TKIs resistance in EGFR-mutant non-small cell lung cancer (NSCLC). We aim to explore the prevalence of transformation in EGFR-wildtype NSCLC and the mechanism of SCLC transformation, which are rarely understood.

Methods
We reviewed 1474 NSCLC patients to investigate the NSCLC-to-SCLC transformed cases and the basic clinical characteristics, driver gene status and disease course of them. To explore the potential functional genes in SCLC transformation, we obtained pre- and post-transformation specimens and subjected them to a multigene NGS panel involving 416 cancer-related genes. To validate the putative gene function, we established knocked-out models by CRISPR-Cas 9 in HCC827 and A549-TP53-/- cells and investigated the effects on tumor growth, drug sensitivity and neuroendocrine phenotype in vitro and in vivo. We also detected the expression level of protein and mRNA to explore the molecular mechanism involved.

Results
We firstly reported an incidence rate of 9.73% (11/113) of SCLC transformation in EGFR-wildtype NSCLC and demonstrated that SCLC transformation is irrespective of EGFR mutation status (P = 0.16). We sequenced 8 paired tumors and identified a series of mutant genes specially in transformed SCLC such as SMAD4, RICTOR and RET. We firstly demonstrated that SMAD4 deficiency can accelerate SCLC transition by inducing neuroendocrine phenotype regardless of RB1 status in TP53-deficient NSCLC cells. Further mechanical experiments identified the SMAD4 can regulate ASCL1 transcription competitively with Myc in NSCLC cells and Myc inhibitor acts as a potential subsequent treatment agent.

Conclusions
Transformation to SCLC is irrespective of EFGR status and can be accelerated by SMAD4 in non-small cell lung cancer. Myc inhibitor acts as a potential therapeutic drug for SMAD4-mediated resistant lung cancer.
NAT10 mediated ac4C acetylation driven m6A modification via involvement of YTHDC1-LDHA/PFKM regulates glycolysis and promotes osteosarcoma
Authors: Mei Zhongting , Shen Zhihua , Pu Jiaying , Liu Qian , Liu Guoxin , He Xuting , Wang Yang , Yue Jinrui , Ge Shiyu , Li Tao , Yuan Ye , Yang Lei 
Published: January 2024

AbstractThe dynamic changes of RNA N6-methyladenosine (m6A) during cancer progression participate in various cellular processes. However, less is known about a possible direct connection between upstream regulator and m6A modification, and therefore affects oncogenic progression. Here, we have identified that a key enzyme in N4-acetylcytidine (ac4C) acetylation NAT10 is highly expressed in human osteosarcoma tissues, and its knockdown enhanced m6A contents and significantly suppressed osteosarcoma cell growth, migration and invasion. Further results revealed that NAT10 silence inhibits mRNA stability and translation of m6A reader protein YTHDC1, and displayed an increase in glucose uptake, a decrease in lactate production and pyruvate content. YTHDC1 recognizes differential m6A sites on key enzymes of glycolysis phosphofructokinase (PFKM) and lactate dehydrogenase A (LDHA) mRNAs, which suppress glycolysis pathway by increasing mRNA stability of them in an m6A methylation-dependent manner. YTHDC1 partially abrogated the inhibitory effect caused by NAT10 knockdown in tumor models in vivo, lentiviral overexpression of YTHDC1 partially restored the reduced stability of YTHDC1 caused by lentiviral depleting NAT10 at the cellular level. Altogether, we found ac4C driven RNA m6A modification can positively regulate the glycolysis of cancer cells and reveals a previously unrecognized signaling axis of NAT10/ac4C-YTHDC1/m6A-LDHA/PFKM in osteosarcoma.
ZDHHC5-mediated S-palmitoylation of FAK promotes its membrane localization and epithelial-mesenchymal transition in glioma
Authors: Wang Yang , Shen Na , Yang Yang , Xia Yuan , Zhang Wenhao , Lu Yu , Wang Zhicheng , Yang Ze , Wang Zhangjie 
Published: January 2024

Abstract
Background
Abnormal activation of FAK is associated with tumor development and metastasis. Through interactions with other intracellular signalling molecules, FAK influences cytoskeletal remodelling, modulation of adhesion signalling, and activation of transcription factors, promoting migration and invasion of tumor cells. However, the exact mechanism that regulates these processes remains unresolved. Herein, our findings indicate that the S-palmitoylation of FAK is crucial for both its membrane localization and activation.

Methods
The palmitoylation of FAK in U251 and T98G cells was assessed by an acyl-PEG exchange (APE) assay and a metabolic incorporation assay. Cellular palmitoylation was inhibited using 2-bromopalmitate, and the palmitoylation status and cellular localization of FAK were determined. A metabolic incorporation assay was used to identify the potential palmitoyl acyltransferase and the palmitoylation site of FAK. Cell Counting Kit-8 (CCK8) assays, colony formation assays, and Transwell assays were conducted to assess the impact of ZDHHC5 in GBM. Additionally, intracranial GBM xenografts were utilized to investigate the effects of genetically silencing ZDHHC5 on tumor growth.

Results
Inhibiting FAK palmitoylation leads to its redistribution from the membrane to the cytoplasm and a decrease in its phosphorylation. Moreover, ZDHHC5, a protein-acyl-transferase (PAT), catalyzes this key modification of FAK at C456. Knockdown of ZDHHC5 abrogates the S-palmitoylation and membrane distribution of FAK and impairs cell proliferation, invasion, and epithelial-mesenchymal transition (EMT). Taken together, our research reveals the crucial role of ZDHHC5 as a PAT responsible for FAK S-palmitoylation, membrane localization, and activation.

Conclusions
These results imply that targeting the ZDHHC5/FAK axis has the potential to be a promising strategy for therapeutic interventions for glioblastoma (GBM).