Key Genes Research Articles

Indirubin, an Active Component of Indigo Naturalis, Exhibits Inhibitory Effects on Leukemia Cells via Targeting HSP90AA1 and PI3K/Akt Pathway.

This research intended to predict the active ingredients and key target genes of Indigo Naturalis in treating human chronic myeloid leukemia (CML) using network pharmacology and conduct the invitro verification. The active components of Indigo Naturalis and the corresponding targets and leukemia-associated genes were gathered through public databases. The core targets and pathways of Indigo Naturalis were predicted through protein-protein interaction (PPI) network, gene ontology (GO) function, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Next, after intersecting with leukemia-related genes, the direct core target gene of Indigo Naturalis active components was identified. Subsequently, HL-60 cells were stimulated with indirubin (IND) and then examined for cell proliferation using CCK-8 assay and cell cycle, cell apoptosis, and mitochondrial membrane potential using flow cytometry. The content of apoptosis-associated proteins (Cleaved Caspase 9, Cleaved Caspase 7, Cleaved Caspase 3, and Cleaved parp) were detected using Western blot, HSP90AA1 protein, and PI3K/Akt signaling (PI3K, p-PI3K, Akt, and p-Akt) within HL-60 cells. A total of 9 active components of Indigo Naturalis were screened. The top 10 core target genes (TNF, PTGS2, RELA, MAPK14, IFNG, PPARG, NOS2, IKBKB, HSP90AA1, and NOS3) of Indigo Naturalis active components within the PPI network were identified. According to the KEGG enrichment analysis, these targets were associated with leukemia-related pathways (such as acute myeloid leukemia and CML). After intersecting with leukemia-related genes, it was found that IND participated in the most pairs of target information and was at the core of the target network; HSP90AA1 was the direct core gene of IND. Furthermore, the in-vitro cell experiments verified that IND could inhibit the proliferation, elicit G2/M-phase cell cycle arrest, enhance the apoptosis of HL-60 cells, reduce mitochondrial membrane potential, and promote apoptosis-related protein levels. Under IND treatment, HSP90AA1 overexpression notably promoted cell proliferation and inhibited apoptosis. Additionally, IND exerted tumor suppressor effects on leukemia cells by inhibiting HSP90AA1 expression. IND, an active component of Indigo Naturalis, could inhibit CML progression, which may be achieved via inhibiting HSP90AA1 and PI3K/Akt signaling expression levels.

Exploring the ferroptosis-related gene lipocalin 2 as a potential biomarker for sepsis-induced acute respiratory distress syndrome based on machine learning

BackgroundSepsis is a major cause of mortality in patients, and ARDS is one of the most common outcomes. The pathophysiology of acute respiratory distress syndrome (ARDS) caused by sepsis is significantly impacted by genes related to ferroptosis. MethodsIn this study, Weighted gene co-expression network analysis (WGCNA), protein-protein interaction (PPI) networks, functional enrichment analysis, and machine learning were employed to identify characterized genes and to construct receiver operating characteristic (ROC) curves. Additionally, DNA methylation levels were quantified and single-cell analysis was conducted. To validate the alterations in the expression of Lipocalin-2 (LCN2) and ferroptosis-related proteins in the in vitro model, Western blotting was carried out, and the changes in intracellular ROS and Fe2+ levels were detected. ResultsA combination of eight machine learning algorithms, including RFE, LASSO, RandomForest, SVM-RFE, GBDT, Bagging, XGBoost, and Boruta, were used with a machine learning model to highlight the significance of LCN2 as a key gene in sepsis-induced ARDS. Analysis of immune cell infiltration showed a positive correlation between neutrophils and LCN2. In a cell model induced by LPS, it was found that Fer-1, a ferroptosis inhibitor, was able to reverse the expression of LCN2. Knocking down LCN2 in BEAS-2B cells reversed the LPS-induced lipid peroxidation, Fe2+ levels, ACSL4, and GPX4 levels, indicating that LCN2, a FRG, plays a crucial role in mediating ferroptosis. ConclusionUpon establishing an FRG model for individuals with sepsis-induced ARDS, we determined that LCN2 could be a dependable marker for predicting survival in these patients. This finding provides a basis for more accurate ARDS diagnosis and the exploration of innovative treatment options.

[Prediction of prognosis of patients with radical resection of intrahepatic cholangiocarcinoma based on single cell omics].

Objectives: To analyze the survival benefit of intrahepatic cholangiocarcinoma (ICC) radical resection based on single cell omics. Methods: This is a retrospective case-series study. ICC single-cell sequencing was integrated from four data sets in the Gene Expression Omnibus Database, with a total of 46 patients undergoing radical resection, to explore the characteristics of the microenvironment. Microarray data of 100 ICC cases was analyzed in the EMBI database with survival data. The infiltration abundance of each epithelial cell cluster was calculated in each microarray data sample using the ssGSEA algorithm. The key epithelial cell cluster associated with poor patient outcomes was explored. The clinical value of key marker genes in this subgroup was studied. Prognostic marker genes were selected using the univariate and multivariate Cox proportional hazards(CoxPH) model. The The CoxPH model was constructed by the target genes and a nomogram was drawn. Kaplan-Meier survival analysis was used to verify the relationship between score and prognosis of patients. The predictive power of the model was evaluated by receiver operating characteristic(ROC) curves, calibration curves, and decision curve analysis (DCA). Results: Epithelial cell clusters infiltrated almost exclusively in tumor tissue. The MT2A+ epithelial cell subset was associated with a poorer prognosis for patients with a high invasion abundance and patients characterized by infiltration of this group were defined as antioxidant. After screening marker genes in this cluster using a univariate and multivariate CoxPH model, the following genes were found to be independent prognostic factors: FILPIL, NFKBIA, PEG10, SERPINB5. The CoxPH model was constructed using the four gene expression levels, and the survival rate of patients in the high-risk group was significantly lower than those in the low-risk group (all P<0.05). The constructed nomogram had good discrimination and validity. The ROC curve showed that the predicted area under the curve was 0.779, 0.844 and 0.845 at 1, 3 and 5 years, respectively. Compared to clinical indicators, the model had better predictive power using the calibration curve and the DCA test. Conclusions: The MT2A+ epithelial cell group may be associated with the prognosis of patients with ICC, and the concept of ICC tissue typing of antioxidant and non-antioxidant types is proposed. The type of antioxidant may predict the poor prognosis of the patients, and postoperative adjuvant therapy and other means could be considered to improve the survival of the patients.

Seasonal Gene Profiling in Tuz Lake with Regard to Biogeochemical Cycling

Tuz Lake, a thalassohaline lake with a salt rate of 32%, is a unique habitat for a halophilic microbiome. Culture-dependent and independent methods have been applied to identify prokaryotic microbial diversity in Tuz Lake. However, the key processes and genes involved in biogeochemical cycles in Tuz Lake have not been investigated seasonally. The aim of the study is to investigate seasonal gene profiling in Tuz Lake associated with biogeochemical cycling and thereby reveal more about the ecosystem dynamics of this extreme environment. Therefore, the PICRUSt2 tool was applied to analyze the metabolic function of archaeal and bacterial diversity in Tuz Lake. As a result of metabolic functions based on 16S rDNA amplicon sequencing data, it was observed that methane production by H2 and CO2 by anaerobic archaea in Tuz Lake was the predominant methanogenesis pathway. It was determined that sulfur oxidation was the dominant sulfur metabolism, while the reductive citric acid cycle was the dominant carbon fixation pathway.

Identification of Key Genes in Angiogenesis of Breast and Prostate Cancers in the Context of Different Cell Types.

Angiogenesis involves the development of new blood vessels. Biochemical signals start this process in the body, which is followed by migration, growth, and differentiation of endothelial cells that line the inside wall of blood vessels. This process is vital for the growth of cancer cells and tumors. We started our analysis by composing a list of genes that have a validated impact in humans with respect to angiogenesis-related phenotypes. Here, we have investigated the expression patterns of angiogenesis-related genes in the context of previously published single-cell RNA-Seq data from prostate and breast cancer samples. Using a protein-protein interaction network, we showed how different modules of angiogenesis-related genes are overexpressed in different cell types. In our results, genes, such as ACKR1, AQP1, and EGR1, showed a strong cell type-dependent overexpression pattern in the two investigated cancer types, which can potentially be helpful in the diagnosis and follow-up of patients with prostate and breast cancer. Our work demonstrates how different biological processes in distinct cell types contribute to the angiogenesis process, which can provide clues regarding the potential application of targeted inhibition of the angiogenesis process.

Comparative transcriptome and coexpression network analysis reveals key pathways and hub candidate genes associated with sunflower (Helianthus annuus L.) drought tolerance

BackgroundDrought severely limits sunflower production especially at the seedling stage. To investigate the response mechanism of sunflowers to drought stress, we utilized two genotypes of sunflower materials with different drought resistances as test materials. The physiological responses were investigated under well-watered (0 h) and drought-stressed conditions (24 h, 48 h, and 72 h).ResultsANOVA revealed the greatest differences in physiological indices between 72 h of drought stress and 0 h of drought stress. Transcriptome analysis was performed after 72 h of drought stress. At 0 h, there were 7482 and 5627 differentially expressed genes (DEGs) in the leaves of K55 and K58, respectively, and 2150 and 2527 DEGs in the roots of K55 and K58, respectively. A total of 870 transcription factors (TFs) were identified among theDEGs, among which the high-abundance TF families included AP2/ERF, MYB, bHLH,and WRKY. Five modules were screened using weighted gene coexpressionnetwork analysis (WGCNA), three and two of which were positively and negatively, respectively, related to physiological traits. KEGG analysis revealedthat under drought stress, “photosynthesis”, “carotenoid biosynthesis”, “starch and sucrose metabolism”, “ribosome”, “carotenoid biosynthesis”, “starch and sucrose metabolism”, “protein phosphorylation” and “phytohormone signaling” are six important metabolic pathways involved in the response of sunflower to drought stress. Cytoscape software was used to visualize the three key modules, and the hub genes were screened. Finally, a total of 99 important candidate genes that may be associated with the drought response in sunflower plants were obtained, and the homology of these genes was compared with that in Arabidopsis thaliana.ConclusionsTaken together, our findings could lead to a better understanding of drought tolerance in sunflowers and facilitate the selection of drought-tolerant sunflower varieties.

Prognostic value and potential function of a novel heme-related LncRNAs signature in gastric cancer

Heme is a coordination complex formed by the binding of iron ions and porphyrin rings. Its metabolic processes are associated with various cancers, including gastric cancer (GC). In recent years, long non-coding RNAs (LncRNAs) have been identified as key regulatory factors in GC. However, the role of LncRNAs associated with heme metabolism in GC and their relationship with prognosis have not been reported. In this study, we constructed a novel LncRNAs signature related to heme metabolism (HMlncSig) and validated its prognostic value for predicting the survival of GC patients through training, test, and entire cohorts. Kaplan-Meier analysis demonstrated that patients in the high-risk group had shorter survival times. Univariate and multivariate Cox regression analysis showed that HMlncSig was an independent prognostic indicator for GC patients, regardless of other clinical pathological features. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis and gene set variation analysis pathways showed that the activation of these markers may be involved in tumor progression, influencing the survival of GC patients. The nomogram, based on HMlncSig score and clinical features, demonstrated the strong predictive ability of this signature. Additionally, significant differences were observed between the high-risk and low-risk groups in terms of immune cell subtypes, expression of immune checkpoint genes, and response to chemotherapy and immunotherapy. Through clinical validation, we found that the risk score and heme levels of GC patients were both significantly elevated and correlated with the degree of malignancy. Furthermore, we found that AP000692.1, a key gene in this signature, promoted the proliferation, migration, and invasion of GC cells. In conclusion, our HMlncSig model has significant predictive value for the prognosis of GC patients and can provide clinical guidance for personalized immunotherapy.

The accessible chromatin landscape of lipopolysaccharide-induced systemic inflammatory response identifying epigenome signatures and transcription regulatory networks in chickens

Lipopolysaccharide (LPS) can induce systemic inflammatory response (SIR) in animals. Understanding the regulatory mechanism of SIR and therapies to ensure healthy growth is urgently needed. Chromatin remodeling plays a crucial role in the expression of genes involved in immune diseases. In the present study, the ATAC-seq analysis revealed 3491 differential open chromatin sites in the spleen of chicks with SIR induced by LPS challenge, and we presented the motifs on these sites and the associated transcription factors. The regulatory network was presented by combining the differential open chromatin data with the mRNAs and exploded cytokines. Interestingly, the LPS challenge could regulate the mRNA expression of 202 genes through chromatin reprogramming, including critical genes such as TLE1 and JUN, which regulate signaling pathways such as I-κB kinase/NF-κB, Toll-like receptor, and downstream cytokine genes. Furthermore, dietary daidzein could inhibit DNA topoisomerase II, which reprograms the spatial conformation of chromatin in the inflammatory response and attenuates SIR. In conclusion, we successfully identified key genes directly regulated by chromatin reprogramming in SIR and demonstrated the chromatin epigenome signatures and transcriptional regulatory network, which provides an important reference for further research on avian epigenetics. There is great potential for alleviating SIR using dietary daidzein.

SH3GL2 and MMP17 as lung adenocarcinoma biomarkers: a machine-learning based approach

ObjectiveUsing bioinformatics machine learning methods, our research aims to identify the potential key genes associated with Lung adenocarcinoma (LUAD). MethodsWe obtained two gene expression profiling microarrays (GSE68571 and GSE74706) from the public Gene Expression Omnibus (GEO) database at the National Centre for Biotechnology Information (NCBI). The purpose was to identify Differentially Expressed Genes (DEGs) between the lung adenocarcinoma group and the healthy control group. The limma R package in R was utilized for this analysis. For the differential gene diagnosis of lung adenocarcinoma, we employed the least absolute shrinkage and selection operator (LASSO) regression and SVM-RFE screening crossover. To evaluate the performance, ROC curves were plotted. We performed immuno-infiltration analysis using CIBERSORT. Finally, we validated the key genes through qRT-PCR and Western-blot verification, then downregulated MMP17 gene expression, upregulated SH3GL2 gene expression, and performed CCK8 experiments. ResultsA total of 32 Differentially Expressed Genes (DEGs) were identified. Two diagnostic marker genes, SH3GL2 and MMP17, were selected by employing LASSO and SVM-RFE machine learning methods. In Lung adenocarcinoma cells, the expression of MMP17 was observed to be elevated compared to normal lung epithelial cells in the control group (P < 0.05). In contrast, a down-regulation of SH3GL2 was found in Lung adenocarcinoma cells (P < 0.05). Finally, we downregulated MMP17 and upregulated SH3GL2 gene expression, then the CCK8 showed that the proliferation of both lung cancer cells was inhibited. ConclusionSH3GL2 and MMP17 are expected to be potential biomarkers for Lung adenocarcinoma.

Three-Dimensional Genomics: A New Perspective and Therapeutic Strategies in Prostate Cancer Research

Prostate cancer (PC) poses significant health risks to men globally. Enhancing our understanding of prostate cancer biology is crucial for facilitating early diagnosis and effective treatment strategies. A multitude of high-throughput sequencing studies, encompassing whole genome resequencing, transcriptome sequencing, and genome-wide association studies, have unearthed vital point mutations, structural variations, and epigenomic alterations linked to prostate cancer. These findings have significantly enriched our knowledge of the genomic framework for prostate cancer. Still, these investigations have predominantly centered on the one- or two-dimensional landscape of the genome. Research in three-dimensional genomics underscores the critical role of the genome's three-dimensional spatial structure in maintaining normal cellular functions. Additionally, it demonstrates that dysregulation of key genes in numerous cancers relates to the chromatin's spatial organization across various levels. This article explores the intricate three-dimensional architecture of chromosomes. It outlines the progressive development of techniques used in three-dimensional genomic research and synthesizes the application of these techniques in the study of prostate cancer biology. Furthermore, it proposes potential therapeutic strategies for prostate cancer.

Identification of Anoikis-Related Genes in Spinal Cord Injury: Bioinformatics and Experimental Validation.

Spinal cord injury (SCI) is a serious disease without effective therapeutic strategies. To identify the potential treatments for SCI, it is extremely important to explore the underlying mechanism. Current studies demonstrate that anoikis might play an important role in SCI. In this study, we aimed to identify the key anoikis-related genes (ARGs) providing therapeutic targets for SCI. The mRNA expression matrix of GSE45006 was downloaded from the Gene Expression Omnibus (GEO) database, and the ARGs were downloaded from the Molecular Signatures Database (MSigDB database). Then, the potential differentially expressed ARGs were identified. Next, correlation analysis, gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and protein-protein interaction (PPI) analysis were employed for the differentially expressed ARGs. Moreover, miRNA-gene networks were constructed by the hub ARGs. Finally, RNA expression of the top ten hub ARGs was validated in the SCI cell model and rat SCI model. A total of 27 common differentially expressed ARGs were identified at different time points (1, 3, 7, and 14days) following SCI. The GO and KEGG enrichment analysis of these ARGs indicated several enriched terms related to proliferation, cell cycle, and apoptotic process. The PPI results revealed that most of the ARGs interacted with each other. Ten hub ARGs were further screened, and all the 10 genes were validated in the SCI cell model. In the rat model, only seven genes were validated eventually. We identified 27 differentially expressed ARGs of the SCI through bioinformatic analysis. Seven real hub ARGs (CCND1, FN1, IGF1, MYC, STAT3, TGFB1, and TP53) were identified eventually. These results may expand our understanding of SCI and contribute to the exploration of potential SCI targets.

Abstract 5589: CBFB::MYH11 expression activates Trp53 in myeloid cells, which inhibits cell growth and sensitizes cells to cytarabine

Abstract CBFB::MYH11 acute myeloid leukemia (AML) is a subset of Core Binding Factor AML initiated by an oncofusion in which Core Binding Factor Beta (CBFB) is fused to the C-terminus of smooth muscle myosin heavy chain 11 (MYH11). CBFB::MYH11 AML has a high cure rate with conventional anthracycline/cytarabine induction and high-dose cytarabine consolidation in patients under the age of 60, and remains chemosensitive after multiple relapses; the mechanisms underlying chemosensitivity are unknown. TP53 is a key regulator of DNA damage response and apoptosis, and TP53-mutated AML is often chemoresistant. Large sequencing studies have revealed only rare co-occurrence of TP53 mutations with CBFB::MYH11. We initially used CBFB::MYH11 as a negative control for an unrelated study of TP53 function and unexpectedly found striking synergy. While expression of CBFB::MYH11 in WT murine hematopoietic stem and progenitor cells (HSPCs) induced a low-penetrance (~20%), long latency (~6 months) myelomonocytic AML, expression of CBFB::MYH11 in Trp53-/- HSPCs induced AML with 100% penetrance and a median latency of 8 weeks. CBFB::MYH11-induced AMLs arising in Trp53-/- cells were similar by morphology and flow cytometry phenotype. We validated these findings in a second model, using a conditional Trp53flox/flox x Vav1-Cre model in which Trp53 is inactivated in HSPCs, with nearly identical results. Trp53-/- cells transduced with Empty Vector did not develop AML, but did develop T cell lymphoma with a median latency of 16 weeks, similar to prior studies. Trp53 deficiency also cooperated with RUNX1::RUNX1T1, the other Core Binding Factor oncofusion, to induce a high-penetrance (100%) and short latency AML (median latency 10 weeks); consistent with prior data, RUNX1::RUNX1T1 expression alone in Trp53 WT cells was insufficient to induce AML. To determine whether Trp53 influences the chemosensitivity of CBFB::MYH11 expressing cells, we performed cytarabine dose-response experiments. While CBFB::MYH11-transduced WT HSPCs were very sensitive to cytarabine (relative to EV), sensitivity was reduced in CBFB::MYH11-transduced Trp53-/- cells, suggesting that CBFB::MYH11-induced chemosensitivity is, at least in part, mediated by the TP53 pathway. To determine whether CBFB::MYH11 expression results in upregulation of key TP53 pathway genes, we transduced WT HSPCs with EV or CBFB::MYH11, transplanted cells into WT mice, harvested bone marrow at 3 months, and performed RNA-Seq. Key TP53 pathway genes were upregulated, including Bbc3/Puma, Bax, Cdkn1a/p21, and Trp53 (all 2.6-4.6-fold, p≤0.05). These data suggest that the chemosensitivity and the favorable prognosis of CBFB::MYH11 AML may be due to the direct activation of the TP53 pathway by CBFB::MYH11. Similar studies are underway with RUNX1::RUNX1T1-transduced cells to determine whether TP53 is relevant for the chemosensitivity of that CBF AML as well. Citation Format: Ryan B. Day, Julia A. Hickman, Casey D. Katerndahl, Sai Mukund Ramakrishnan, Christopher A. Miller, Timothy J. Ley. CBFB::MYH11 expression activates Trp53 in myeloid cells, which inhibits cell growth and sensitizes cells to cytarabine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5589.

Expanded phylogeny of extremely halophilic archaea shows multiple independent adaptations to hypersaline environments.

Extremely halophilic archaea (Haloarchaea, Nanohaloarchaeota, Methanonatronarchaeia and Halarchaeoplasmatales) thrive in saturating salt concentrations where they must maintain osmotic equilibrium with their environment. The evolutionary history of adaptations enabling salt tolerance remains poorly understood, in particular because the phylogeny of several lineages is conflicting. Here we present a resolved phylogeny of extremely halophilic archaea obtained using improved taxon sampling and state-of-the-art phylogenetic approaches designed to cope with the strong compositional biases of their proteomes. We describe two uncultured lineages, Afararchaeaceae and Asbonarchaeaceae, which break the long branches at the base of Haloarchaea and Nanohaloarchaeota, respectively. We obtained 13 metagenome-assembled genomes (MAGs) of these archaea from metagenomes of hypersaline aquatic systems of the Danakil Depression (Ethiopia). Our phylogenomic analyses including these taxa show that at least four independent adaptations to extreme halophily occurred during archaeal evolution. Gene-tree/species-tree reconciliation suggests that gene duplication and horizontal gene transfer played an important role in this process, for example, by spreading key genes (such as those encoding potassium transporters) across extremely halophilic lineages.

Impact of visceral adipose tissue on longevity and metabolic health: a comparative study of gene expression in perirenal and epididymal fat of Ames dwarf mice.

Emerging research underscores the pivotal role of adipose tissue in regulating systemic aging processes, particularly when viewed through the lens of the endocrine hypotheses of aging. This study delves into the unique adipose characteristics in an important animal model of aging - the long-lived Ames dwarf (df/df) mice. Characterized by a Prop1df gene mutation, these mice exhibit a deficiency in growth hormone (GH), prolactin, and TSH, alongside extremely low circulating IGF-1 levels. Intriguingly, while surgical removal of visceral fat (VFR) enhances insulin sensitivity in normal mice, it paradoxically increases insulin resistance in Ames dwarfs. This suggests an altered profile of factors produced in visceral fat in the absence of GH, indicating a unique interplay between adipose tissue function and hormonal influences in these models. Our aim was to analyze the gene expression related to lipid and glucose metabolism, insulin pathways, inflammation, thermoregulation, mitochondrial biogenesis, and epigenetic regulation in the visceral (perirenal and epididymal) adipose tissue of Ames dwarf and normal mice. Our findings reveal an upregulation in the expression of key genes such as Lpl, Adrβ3, Rstn, Foxo1, Foxo3a, Irs1, Cfd, Aldh2, Il6, Tnfα, Pgc1α, Ucp2, and Ezh2 in perirenal and Akt1, Foxo3a, PI3k, Ir, Acly, Il6, Ring1a, and Ring 1b in epididymal fat in df/df mice. These results suggest that the longevity phenotype in Ames dwarfs, which is determined by peripubertal GH/IGF-1 levels, may also involve epigenetic reprogramming of adipose tissue influenced by hormonal changes. The increased expression of genes involved in metabolic regulation, tumor suppression, mitochondrial biogenesis, and insulin pathways in Ames dwarf mice highlights potentially beneficial aspects of this model, opening new avenues for understanding the molecular underpinnings of longevity and aging.

Abstract 4665: USP18 promotes anlotinib resistance in medullary thyroid carcinoma by stabilizing aurora B kinase

Abstract Targeted therapy resistance is the main cause of disease progression in advanced medullary thyroid carcinoma (MTC). However, the key molecules and mechanisms remain unclear. We previously performed a genome-wide in vitro screen of Anlotinib resistance-related genes in human MTC cell line TT using CRISPR-dCas9-SAM system and identified ubiquitin-specific protease 18 (USP18) as a key molecule mediating Anlotinib resistance in MTC. However, its downstream mechanism needs further clarification. Based on the high-throughput DNA sequencing results of the control group and the Anlotinib screening group, a group of key genes related to Anlotinib resistance in MTC was screened through the MAGeCK algorithm and the enrichment ratio of sgRNA in the Anlotinib screening group. Among them, USP18 ranked first in the enrichment results. Analysis of the correlation between the expression level of USP18 in MTC patient specimens and survival revealed that patients with high USP18 expression in tumor cells had poorer prognosis. Overexpression of USP18 in MTC tumor cells significantly promoted the growth of subcutaneous xenograft tumors in NOD/SCID mice treated with Anlotinib. In vitro cell experiments also showed that overexpression of USP18 promoted the proliferation of MTC tumor cells and significantly inhibited the proportion of cell apoptosis after Anlotinib treatment. Further research found that USP18, as a deubiquitinating enzyme that can specifically remove ISG15-like ubiquitin-like proteins from substrate proteins, can improve the kinase stability of Aurora B by removing ISG15-like ubiquitin modification and promote MTC tumor cell proliferation through the activation of PI3K-Akt signaling regulated by Aurora B kinase. Application of Aurora B kinase inhibitor in Anlotinib-resistant MTC cell lines significantly inhibited tumor cell proliferation, and Aurora B kinase inhibitor significantly increased the proportion of cell apoptosis in MTC tumor cells overexpressing USP18. In the subcutaneous xenograft tumor model of MTC in NOD/SCID mice, the growth of xenograft tumors in the Anlotinib combined with Aurora B kinase inhibitor treatment group was also significantly inhibited. In summary, overexpressed USP18 can activate PI3K-Akt signaling and promote tumor cell proliferation by increasing the stability of Aurora B kinase, ultimately leading to the formation of Anlotinib resistance in MTC patients. Combined use of Aurora B kinase inhibitor is a potentially effective combined treatment strategy for Anlotinib-resistant MTC patients. Citation Format: Xuan Qin, Xing Wan, Dapeng Li, Yimeng Liu, Xianhui Ruan, Fangkun Luan, Kejia Xu, Jia Li, Rong Xiang, Yongjun Piao, Yi Shi, Xiangqian Zheng. USP18 promotes anlotinib resistance in medullary thyroid carcinoma by stabilizing aurora B kinase [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4665.

Metabolic engineering of Escherichia coli for high-level production of the biodegradable polyester monomer 2-pyrone-4,6-dicarboxylic acid

2-Pyrone-4,6-dicarboxylic acid (PDC), a chemically stable pseudo-aromatic dicarboxylic acid, is a promising building block compound for manufacturing biodegradable polyesters. This study aimed to construct high-performance cell factories enabling the efficient production of PDC from glucose. Firstly, the effective enzymes of the PDC biosynthetic pathway were overexpressed on the chromosome of the 3-dehydroshikimate overproducing strain. Consequently, the one-step biosynthesis of PDC from glucose was achieved. Further, the PDC production was enhanced by multi-copy integration of the key gene PsligC encoding 4-carboxy-2-hydroxymuconate-6-semialdehyde dehydrogenase and co-expression of Vitreoscilla hemoglobin. Subsequently, the PDC production was substantially improved by redistributing the metabolic flux for cell growth and PDC biosynthesis based on dynamically downregulating the expression of pyruvate kinase. The resultant strain PDC50 produced 129.37 g/L PDC from glucose within 78 h under fed-batch fermentation conditions, with a yield of 0.528 mol/mol and an average productivity of 1.65 g/L/h. The findings of this study lay the foundation for the potential industrial production of PDC.

MORE FLORET1 controls anther development by negatively regulating key tapetal genes in both diploid and tetraploid rice.

Polyploid hybrid rice (Oryza sativa) has great potential for increasing yields. However, hybrid rice depends on male fertility and its regulation, which is less well studied in polyploid rice than in diploid rice. We previously identified a MYB transcription factor, MORE FLORET1 (MOF1), whose mutation causes male sterility in neo-tetraploid rice. MOF1 expression in anthers peaks at anther Stage 7 (S7) and progressively decreases to low levels at S10. However, it remains unclear how the dynamics of MOF1 expression contribute to male fertility. Here, we carefully examined anther development in both diploid and tetraploid mof1 rice mutants, as well as lines ectopically expressing MOF1 in a temporal manner. MOF1 mutations caused delayed degeneration of the tapetum and middle layer of anthers and aberrant pollen wall organization. Ectopic MOF1 expression at later stages of anther development led to retarded cytoplasmic reorganization of tapetal cells. In both cases, pollen grains were aborted and seed production was abolished, indicating that precise control of MOF1 expression is essential for male reproduction. We demonstrated that five key tapetal genes, CYP703A3 (CYTOCHROME P450 HYDROXYLASE 703A3), OsABCG26 (Oryza sativa ATP BINDING CASSETTE G26), PTC1 (PERSISTENT TAPETAL CELL1), PKS2 (POLYKETIDE SYNTHASE 2), and OsABCG15 (Oryza sativa ATP BINDING CASSETTE G15), exhibit expression patterns opposite to those of MOF1 and are negatively regulated by MOF1. Moreover, DNA affinity purification sequencing (DAP-seq), luciferase activity assays, and electrophoretic mobility shift assays indicated that MOF1 binds directly to the PKS2 promoter for transcriptional repression. Our results provide a mechanistic basis for the regulation of male reproduction by MOF1 in both diploid and tetraploid rice. This study will facilitate the development of polyploid male sterile lines, which are useful for breeding of polyploid hybrid rice.

Transcriptomics and metabolomics analysis revealed the regulatory network of apical buds development in lotus (Nelumbo nucifera Gaertn.)

As a perennial ornamental plant, lotus will begin to become dormant in the face of adverse environmental conditions. The dormancy and apical buds development of lotus are of great significance in regulating the flowering period, storage and reproduction of the lotus rhizome, but the related regulatory mechanisms have not been reported. In this study, we sampled the lotus apical buds at 0 d, 12 d and 24 d after planting, and determined that the lotus could complete the dormancy releaseprocess at 12 days after planting. The monitoring of physiological indexes, hormone levels and transcriptome sequencing of 9 samples in three periods were carried out. It was found that the contents of carbohydrates and hormones changed significantly during apical buds development. Transcriptome analysis also showed that a large number of differentially expressed genes were enriched in hormone synthesis and signal transduction pathways, starch and sucrose metabolism pathways, and lipid metabolism pathways, these genes may be involved in the development of lotus apical buds. Subsequently, we analyzed the differential transcription factors during apical buds development and screened a key differential gene NnHSP18.2. NnHSP18.2 is a heat shock protein (HSP) family gene specifically expressed on apical buds, encoding a protein of 160 amino acids with a conserved α-crystallin domain (ACD) domain. Overexpression of NnHSP18.2 can inhibit Arabidopsis seed germination. These results are of great significance for exploring the molecular mechanism of lotus apical buds development and actively promoting the development of the lotus industry.

Identification of key genes for atherosclerosis in different arterial beds

Atherosclerosis (AS) is the pathologic basis of various cardiovascular and cerebrovascular events, with a high degree of heterogeneity among different arterial beds. However, mechanistic differences between arterial beds remain unexplored. The aim of this study was to explore key genes and potential mechanistic differences between AS in different arterial beds through bioinformatics analysis. Carotid atherosclerosis (CAS), femoral atherosclerosis (FAS), infrapopliteal atherosclerosis (IPAS), abdominal aortic atherosclerosis (AAS), and AS-specific differentially expressed genes (DEGs) were screened from the GSE100927 and GSE57691 datasets. Immune infiltration analysis was used to identify AS immune cell infiltration differences. Unsupervised cluster analysis of AS samples from different regions based on macrophage polarization gene expression profiles. Weighted gene co-expression network analysis (WGCNA) was performed to identify the most relevant module genes with AS. Hub genes were then screened by LASSO regression, SVM-REF, and single-gene differential analysis, and a nomogram was constructed to predict the risk of AS development. The results showed that differential expression analysis identified 5, 4, 121, and 62 CAS, FAS, IPAS, AAS-specific DEGs, and 42 AS-common DEGs, respectively. Immune infiltration analysis demonstrated that the degree of macrophage and mast cell enrichment differed significantly in different regions of AS. The CAS, FAS, IPAS, and AAS could be distinguished into two different biologically functional and stable molecular clusters based on macrophage polarization gene expression profiles, especially for cardiomyopathy and glycolipid metabolic processes. Hub genes for 6 AS (ADAP2, CSF3R, FABP5, ITGAX, MYOC, and SPP1), 4 IPAS (CLECL1, DIO2, F2RL2, and GUCY1A2), and 3 AAS (RPL21, RPL26, and RPL10A) were obtained based on module gene, gender stratification, machine learning algorithms, and single-gene difference analysis, respectively, and these genes were effective in differentiating between different regions of AS. This study demonstrates that there are similarities and heterogeneities in the pathogenesis of AS between different arterial beds.

MiR-181a protects the heart against myocardial infarction by regulating mitochondrial fission via targeting programmed cell death protein 4

Worldwide, myocardial infarction (MI) is the leading cause of death and disability-adjusted life years lost. Recent researches explored new methods of detecting biomarkers that can predict the risk of developing myocardial infarction, which includes identifying genetic markers associated with increased risk. We induced myocardial infarction in mice by occluding the left anterior descending coronary artery and performed TTC staining to assess cell death. Next, we performed ChIP assays to measure the enrichment of histone modifications at the promoter regions of key genes involved in mitochondrial fission. We used qPCR and western blot to measure expression levels of relative apoptotic indicators. We report that miR-181a inhibits myocardial ischemia-induced apoptosis and preserves left ventricular function after MI. We show that programmed cell death protein 4 (PDCD4) is the target gene involved in miR-181a-mediated anti-ischemic injury, which enhanced BID recruitment to the mitochondria. In addition, we discovered that p53 inhibits the expression of miR-181a via transcriptional regulation. Here, we discovered for the first time a mitochondrial fission and apoptosis pathway which is controlled by miR-181a and involves PDCD4 and BID. This pathway may be controlled by p53 transcriptionally, and we presume that miR-181a may lead to the discovery of new therapeutic and preventive targets for ischemic heart diseases.