Biology subjects

Zhao, D.

Publications and source records attributed to Zhao, D..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

POCALI: Prediction and insight On CAncer LncRNAs by Integrating multi-omics data with machine learning

Long non-coding RNAs (lncRNAs) are receiving increasing attention as biomarkers for cancer diagnosis and therapy. Although there are many computational methods to identify cancer lncRNAs, they do not comprehensively integrate multi-omics features for predictions or systematically evaluate the contribution of each omics to the multifaceted landscape of cancer lncRNAs. In this study, we developed an algorithm, POCALI, to identify cancer lncRNAs by integrating 44 omics features across six categories. We explored the contributions of different omics to identifying cancer lncRNAs and, more specifically, how each feature contributes to a single prediction. We also evaluated our model and benchmarked POCALI with existing methods. Finally, we validated the cancer phenotype and genomics characteristics of the predicted novel cancer lncRNAs. POCALI identified secondary structure and gene expression-related features as strong predictors of cancer lncRNAs, and epigenomic features as moderate predictors. POCALI performed better than other methods, especially in terms of sensitivity, and predicted more candidates. Novel POCALI-predicted cancer lncRNAs had strong relationships with cancer phenotypes, similar to known cancer lncRNAs. Overall, this study facilitated the identification of previously undetected cancer lncRNAs and the comprehensive exploration of the multifaceted feature contributions to cancer lncRNA prediction.

bioinformatics↗

bioRxiv2025

Ustilaginoidea virens suppresses floral immunity through promoting GA biosynthesis by the effector SCRE9

Rice false smut caused by Ustilaginoidea virens has become one of the most important rice diseases. U. virens specifically infects rice florets through stamen filaments. However, it is mysterious how U. virens inhibits floral immunity for successful colonization. Here, we show that a small cysteine-rich effector SCRE9 is a key virulence factor in U. virens. Expression of SCRE9 in rice not only suppresses immune responses and false smut resistance, but also significantly increases expression of expansin and gibberellin (GA) biosynthetic genes and GA content in young rice panicles. We further demonstrate that OsSIP1 (SCRE9-interacting protein 1) in rice is targeted by SCRE9 and interacts with the OsMADS63 and OsMADS68 transcription factors that transcriptionally regulate the GA synthesis gene GA3ox1. Interestingly, the chloroplast-localized OsSIP1 recruits OsMADS63/68 into the chloroplast. SCRE9 causes OsSIP1 to be translocated into the nucleus, thereby releasing OsMADS63/68 from the chloroplast into the nucleus to promote GA3ox1 transcription. Therefore, SCRE9 promotes GA biosynthesis and expression of expansins in rice florets, thus loosening cell walls and overcoming the physical barrier during U. virens infection. The finding uncovers an unidentified floret infection strategy that offsets the deficiency in cell wall degrading ability in the unique floret colonizing fungus.

plant biology↗

bioRxiv2024

Aramchol attenuates fibrosis in mouse models of biliary fibrosis and blocks the TGFβ-induced fibroinflammatory mediators in cholangiocytes

BackgroundFibroinflammatory cholangiopathies, such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), are characterized by inflammation and biliary fibrosis, driving disease-related complications. In biliary fibrosis, cholangiocytes activated by transforming growth factor-{beta} (TGF{beta}) release signals that recruit immune cells to drive inflammation and activate hepatic myofibroblasts to deposit the extracellular matrix (ECM). TGF{beta} regulates stearoyl-CoA desaturase (SCD), an enzyme that catalyzes the synthesis of monounsaturated fatty acids, in stimulating fibroinflammatory lipid signaling. However, the role of SCD or its inhibitor, Aramchol, has not been investigated in biliary fibrosis or TGF{beta}-mediated cholangiocyte activation. Method10-16-week-old multi-drug resistance 2 knockout (Mdr2-/-) and 3,5-diethoxycarboncyl-1,4-dihydrocollidine (DDC) diet-fed mice were orally gavaged daily with Aramchol at 12.5 mg/kg/day for 4 and 3 weeks, respectively. Liver and serum were harvested for the assessment of fibrosis and inflammation. Transformed human cholangiocyte cells (H69) and mouse large biliary epithelial cells (MLEs) were used to test the effects of the SCD inhibitor, Aramchol, at varying doses on TGF{beta}-mediated expression of fibroinflammatory signals and were confirmed in PSC-derived cholangiocytes (PSC-Cs) using ELISA, qPCR, and Western blot analyses. ResultsAramchol treatment of Mdr2-/- mice with established biliary fibrosis (treatment) and DDC diet-induced (prevention) models of cholestatic injury and fibrosis demonstrated significant reductions in both measures of ECM synthesis (mRNA expression of ECM components in the liver), collagen content of the liver (picrosirius red staining and hydroxyproline content) and myofibroblast activation (SMA staining). Il6 and Tnfa were also reduced with Aramchol in the liver. RNA-seq analysis of H69 cells showed that Aramchol co-treatment led to significant inhibition of TGF{beta}-induced hepatic fibrosis pathways while upregulating peroxisome proliferator-activated receptor (PPAR) signaling. SCD expression was significantly increased in TGF{beta}-treated H69 cells (2-fold, p<0.05). Aramchol in a dose-dependent manner significantly attenuated the increased expression of the fibrotic marker, plasminogen activator inhibitor-1 (PAI-1/SERPINE1), and hepatic stellate cell-activating genes (VEGFA and PDGFB) in TGF{beta}-activated H69 and MLEs. Aramchol also markedly reduced the expression of the inflammatory cytokine, interleukin 6 (IL6). SCD siRNA knockdown produced similar results in H69 cells. Furthermore, in PSC-Cs, the expressions of SCD, VEGFA and IL6 were significantly reduced with Aramchol. The expression of the anti-fibroinflammatory factors PPAR and -{gamma} were modestly increased in cholangiocyte cell lines with increased expression of PPAR-responsive genes and increased nuclear binding of DNA PPAR response elements with Aramchol co-treatment compared to TGF{beta} only. ConclusionAramchol, an SCD inhibitor, both attenuates and prevents biliary fibrosis in mouse models of cholestatic injury and fibrosis. This effect is partially due to Aramchol inhibiting TGF{beta}-induced fibroinflammatory mediators in cholangiocytes by upregulating PPAR and -{gamma} expression and activity. These findings, along with Aramchols excellent safety profile in clinical trials, provide the rationale for assessing Aramchol in further clinical studies in patients with biliary fibrosis, particularly PSC, where a treatment is desperately needed.

cell biology↗

bioRxiv2024

Initiation of ERAD by the bifunctional complex of Mnl1 mannosidase and protein disulfide isomerase

Misfolded glycoproteins in the endoplasmic reticulum (ER) lumen are translocated into the cytosol and degraded by the proteasome, a conserved process called ER-associated protein degradation (ERAD). In S. cerevisiae, the glycan of these proteins is trimmed by the luminal mannosidase Mnl1 (Htm1) to generate a signal that triggers degradation. Curiously, Mnl1 is permanently associated with protein disulfide isomerase (Pdi1). Here, we have used cryo- electron microscopy, biochemical, and in vivo experiments to clarify how this complex initiates ERAD. The Mnl1-Pdi1 complex first de-mannosylates misfolded, globular proteins that are recognized through a C-terminal domain (CTD) of Mnl1; Pdi1 causes the CTD to ignore completely unfolded polypeptides. The disulfides of these globular proteins are then reduced by the Pdi1 component of the complex, generating unfolded polypeptides that can be translocated across the membrane. Mnl1 blocks the canonical oxidative function of Pdi1, but allows it to function as the elusive disulfide reductase in ERAD.

cell biology↗

bioRxiv2024

An all-in-one AAV vector for cardiac-specific gene silencing by an adenine base editor

ABE editing outcomes are highly variable and unpredictable, depending on various factors. Therefore, the success rate of creating targeted A*T-to-G*C conversions using ABE is not high. But utilizing ABE to target RNA splicing sites for gene silencing has a higher success rate. Another challenge for base editing in the heart is that traditional ABE is too large in size, necessitating dual AAV delivery. Whether single AAV delivery can be achieved remains to be explored. In this study, we demonstrated how the diversity of Cas9 homologs and screening of sgRNAs can facilitate cardiac base editing. Single-AAV base editing outperformed dual-AAV systems in the heart, potentially benefiting from chromatin accessibility-guided sgRNA selection. These findings have important implications for the development of more effective and predictable base editing tools for cardiac gene therapy.

molecular biology↗

bioRxiv2024

The transcription factor ZmbZIP75 promotes both grain filling and kernel dehydration in maize

Selecting both high-yield and low-kernel-moisture varieties is essential for modern maize production, but relevant breeding efforts are hindered by a lack of valuable regulatory genes. Here, we demonstrate that the transcription factor (TF) basic leucine zipper 75 (ZmbZIP75) promotes grain yield and reduces kernel moisture in maize. Knockout of ZmbZIP75 results in defective grain filling and kernel dehydration, whereas ZmbZIP75 overexpression confers increased grain yield per plant and decreased kernel moisture without altering plant architecture. Mechanistically, during the grain filling stage, ZmbZIP75 is transcriptionally induced by maternal-derived basal abscisic acid (ABA) and directly activates multiple core starch synthesis-related genes and key TFs, thereby promoting grain filling and final yield. In the late stage of kernel development, high concentrations of zygotic ABA enhance ZmbZIP75 phosphorylation through SnRK2.10. The phosphorylated ZmbZIP75 subsequently transactivates and interacts with TF VP1 to synergistically promote kernel dehydration. This study thus highlights the potential of ZmbZIP75 for engineering both high-yield and low-kernel-moisture varieties to meet the demands of high-efficient maize production. IN A NUTSHELLO_ST_ABSBackgroundC_ST_ABSHigh grain yield in maize is generally associated with elevated kernel moisture at harvest, which is the main limiting factor for modern maize production. Therefore, it is of importance to select varieties with both high-yield and low-kernel-moisture. These traits are largely controlled by grain filling and kernel dehydration, two tightly connected processes during maize kernel development. Abscisic acid (ABA) is well-documented for its vital role in grain filling and dehydration maturation. However, the molecular mechanisms by which ABA coordinates these two processes remain unclear. QuestionWhat are the regulatory factors involved, and how do they mediate ABA signaling to coordinate grain filling and dehydration maturation in maize? FindingsZmbZIP75 is transcriptionally induced by ABA and directly activates multiple core starch synthesis-related genes and key TFs in developing maize endosperm, thereby promoting grain filling. In developing embryo, ABA enhances ZmbZIP75 phosphorylation via SnRK2.10. The phosphorylated ZmbZIP75 then transactivates and interacts with VP1 to synergistically promote kernel dehydration. Moreover, ZmbZIP75 overexpression confers increased grain yield and reduced kernel moisture in maize. Next stepsWhile ZmbZIP75 is directly phosphorylated to mediate ABA signaling during dehydration maturation, ZmbZIP75 is transcriptionally induced by ABA during grain filling. We plan to identify upstream factors that mediate ABA signaling to regulate ZmbZIP75 expression, thereby enhancing our understanding of ABA-promoted grain filling in maize.

plant biology↗

bioRxiv2024

Quantitative site-specific glycoproteomics by ZenoTOF reveals glyco-signatures for breast cancer diagnosis

Intact glycopeptide characterization by mass spectrometry has proven a versatile tool for site-specific glycoproteomics analysis and biomarker screening. Here, we present a method using the ZenoTOF instrument with optimized fragmentation for intact glycopeptide identification and demonstrate its ability to analyze large-cohort glycoproteomes. From 124 clinical serum samples of breast cancer, non-cancerous diseases, and non-disease controls, a total of 6901 unique site-specific glycans on 807 gly-cosites of proteins were detected. Much more differences of glycoproteome were observed in breast diseases than the proteome. By employing machine learning, 15 site-specific glycans were determined as potential glyco-signatures in detecting breast cancer. The results demonstrate that our method provides a powerful tool in glycoproteomic analyses for biomarker discovery studies.

cancer biology↗

bioRxiv2024

Extracellular Vesicle-Associated Neutrophil Elastase Activates Hepatic Stellate Cells and Promotes Liver Fibrogenesis via ERK1/2 Pathway

Liver fibrosis associated with increased mortality is caused by activation of hepatic stellate cells and excessive production and accumulation of extracellular matrix in response to fibrotic insults. It has been shown that in addition to liver inflammation, systemic inflammation also contributes to liver fibrogenesis. A deeper understanding of mechanisms that control liver fibrotic response to intra-and extra-hepatic inflammation is essential to develop novel clinical strategies against this disease. Extracellular vesicles (EV) have been recognized as immune mediators that facilitate activation of hepatic stellate cells. In inflammatory diseases, activated neutrophils release neutrophil elastase (NE) bound to EV, which has been identified as a significant contributor to inflammation by promoting immune cell activation. Here, we aimed to explore the role of inflammation derived plasma EV-associated NE in liver fibrogenesis and its potential mechanisms. We show EV-associated NE induces activation, proliferation and migration of hepatic stellate cells by promoting activation of the ERK1/2 signaling pathway. This effect did not occur through EV without surface NE, and Sivelestat, a NE inhibitor, inhibited activation of the ERK1/2 signaling pathway mediated by EV-associated NE. Moreover, we found plasma EV-associated NE increases deposition of collagen1 and -smooth muscle actin in the liver of a mouse model of liver fibrosis (Mdr2-/-). Notably, this effect does not occur in control mice without preexisting liver disease. These data suggest that EV-associated NE is a pro-fibrogenic factor for hepatic stellate cell activation via the ERK1/2 signaling pathway in pre-existing liver injuries. Inhibition of the plasma EV-associated NE in inflammatory conditions may be a therapeutic target for liver fibrosis in patients with inflammatory diseases.

cell biology↗

bioRxiv2024

Unbiased functional genetic screens reveal essential RNA modifications in human cancer and drug resistance

RNA modification pathways are mis-regulated in multiple types of human cancer. To comprehensively identify cancer-relevant RNA modifications and their regulators, we screened all 150 annotated human RNA modifying proteins across 18 different normal and cancer cell lines using a CRISPR-based genetic knockout system. Fifty RNA modifying proteins were essential for survival of at least one cell type. A third of these essential genes were amplified in 38 different human primary cancer types and potentially drive cancer growth. Unexpectedly, the number of essential genes per cell line varied considerably, and this variation was not due to tissue of origin. Instead, we found that cancer cell-specific mitochondrial metabolic plasticity was responsible for the unique requirement of certain RNA modifications. For example, leukemia cells with high intrinsic drug tolerance required mitochondrial flexibility to survive treatment with the anti-leukemic drugs cytarabine and venetoclax. Synthetic lethality screens revealed that drug-resistance is abolished by deleting the mitochondrial methyltransferase TRMT5, which is responsible for the formation of N1-methylguanosine (m1G) in the tRNA anticodon loop. In summary, our study identifies cancer-relevant RNA modifying enzymes, and reveals a novel promising drug target for therapy-resistant acute myeloid leukemia.

cell biology↗

bioRxiv2024

A Drug-Elicitable Alternative-Splicing Module (DreAM) for Tunable AAV Expression and Controlled Myocardial Regeneration

Adeno-associated virus (AAV) is a major vector for gene therapy. A technique to fine-tune the time and level of AAV expression is lacking, which greatly restricts indication choice, therapeutic efficacy and safety. To solve this problem, here we developed a drug-elicitable alternative-splicing module (DreAM) responsive to risdiplam, an FDA-approved alternative splicing modulator. Risdiplam activates DreAM-regulated AAV expression in a dose-dependent manner with an over 2000-fold inducible change depending on the inducer dosage and the organ of interests. With a temporally resolution of a couple of days, DreAM could repeatedly activate AAV expression, determined by the frequency and duration of risdiplam administration. As an example of DreAM-realized gene therapy, this technique was used to control AAV-based cardiac delivery of YAP5SA, a potent cardiomyocyte regeneration factor. DreAM transiently activated AAV-YAP5SA, establishing the dedifferentiation-proliferation-redifferentiation cycle in cardiomyocytes. Consequently, DreAM fulfilled the regenerative capacity of AAV-YAP5SA in myocardial infarction while circumventing toxicity associated with prolonged AAV-YAP5SA expression. Together, these data demonstrated a tremendous potential of DreAM to enhance the efficacy, safety and applicable scope of gene therapy.

bioengineering↗

bioRxiv2024

Chromosome-scale Salvia hispanica L. (Chia) genome assembly reveals rampant Salvia interspecies introgression

Salvia hispanica L. (Chia), a member of the Lamiaceae, is an economically important crop in Mesoamerica, with health benefits associated with its seed fatty acid composition. Chia varieties are distinguished based on seed color including mixed white and black (Chia pinta) and black (Chia negra). To facilitate research on Chia and expand on comparative analyses within the Lamiaceae, we generated a chromosome-scale assembly of a Chia pinta accession and performed comparative genome analyses with a previously published Chia negra genome assembly. The Chia pinta and negra genome sequences were highly similar as shown by a limited number of single nucleotide polymorphisms and extensive shared orthologous gene membership. There is an enrichment of terpene synthases in the Chia pinta genome relative to the Chia negra genome. We sequenced and analyzed the genomes of 20 Chia accessions with differing seed color and geographic origin revealing population structure within S. hispanica and interspecific introgressions of Salvia species. As the genus Salvia is polyphyletic, its evolutionary history remains unclear. Using large-scale synteny analysis within the Lamiaceae and orthologous group membership, we resolved the phylogeny of Salvia species. This study and its collective resources further our understanding of genomic diversity in this food crop and the extent of inter-species hybridizations in Salvia. PLAIN LANGUAGE SUMMARYChia pinta is an economically important crop due to the high fatty acid present in the seeds. There are multiple types of Chia based on the seeds color including mixed which and black (Chia pinta), black (Chia negra), and white (Chia blanca). We generated a genome assembly of Chia pinta and compared it to existing genome assemblies. While the assemblies are highly similar there are key differences in terpene synthase composition between Chia pinta and Chia negra. We also sequenced 20 other Chia accessions with different seed color and geographic origin to determine a population structure within Chia. We generated genomic resources to further our understanding of this food crop. ABBREVIATIONSBGC Biosynthetic gene cluster BUSCO Benchmarking Universal Single Copy Orthologs GO Gene ontology SNP Single nucleotide polymorphism TIR Terminal inverted repeat TPS Terpene synthase WGS Whole genome shotgun

genomics↗

bioRxiv2024

The Prefrontal Cortex to the Hippocampus Glutamatergic Pathway Regulates Reward Effects of Methamphetamine

Methamphetamine (METH) induces reward effects. The prefrontal cortex (PFC) plays a role in regulating top-down. The hippocampus is closely involved reward memory. However, the direct projects from the PFC to the hippocampus have not been studied well. Here, we use conditioned place preference (CPP), a recombinant adeno-associated virus 2/9 (rAAV2/9) and optogenetic approaches to explore a role in METH-CPP for a novel pathway projecting from the medial prefrontal prelimbic cortex (PrL) directly to the dorsal hippocampus CA1 (dCA1). Expressing CaMKII-ChR2 or CaMKII-eNpHR3.0 in the PrL and optically stimulating in the dCA1 blocks or enhances METH-CPP, respectively. Moreover, silencing the PrL to dCA1 glutamatergic pathway with tetanus neurotoxin (TeNT) enhances METH-CPP. These results identify a novel PrL-to-dCA1 glutamatergic pathway regulating METH-CPP.

neuroscience↗

bioRxiv2024

Genome-Wide Identification and Expression Pattern of the N-acetylserotonin deacetylase (ASDAC) Gene Family in Orchidaceae

Orchids are a kind of horticultural plant with highly ornamental and medical value. N-acetylserotonin deacetylase (ASDAC) is the only reverse enzyme of the melatonin biosynthesis pathway, and plays an important role in regulating the balance of melatonin. Melatonin as a multifunctional molecule, is typically involved in plant growth and development regulation, as well as abiotic stress tolerance. Here, we aimed at identifying ASDAC genes from the orchid genome to provide valuable information for further study of the role of melatonin in orchids. In this study, a total of 7 ASDAC genes were identified from the 7 orchid genome with one member in each species. The 7 orchid ASDACs have an HDAC functional domain, and cluster together with functionally confirmed OsHDAC10 and AtHDAC14, it shows that these members may have function of N-acetylserotonin deacetylase. Furthermore, based on the phylogenetic, motif, and gene structure analysis, the same clusters orchid ASDAC or ASDAC-like genes generally contained similar introns and motifs, suggesting the distribution pattern of exons/introns and motifs were strongly related to phylogeny on an evolutionary basis. Interestingly, homologous genes of OsHDAC10 and AtHDAC14 in Gastrodia elata have low homology and not cluster together with rice and A.thaliana ASDACs, showing that ASDAC gene family may lost in the holomycoheterotrophic orchids. The Ka/Ks ratios of ASDAC gene pairs from lower plant to higher plant were less than one, suggested that ASDAC genes have undergone purifying selection during the evolution process. Cis-acting element analysis results showed that the promoter regions of orchid ASDAC genes contained plant growth and development, phytohormone, and stress-responsive elements. Moreover, most orchid ASDACs were expressed in the roots, stems, leaves, flowers, and seeds. Combined cis-acting element and tissue expression analysis, indicating orchid ASDAC genes are involved in melatonin regulation of growth and development, as well as melatonin responding to various stresses in orchids. These findings of orchid ASDAC genes may provide valuable information for further study of the role of melatonin in orchids.

bioinformatics↗

bioRxiv2024

Multi-trait modeling and machine learning discover new markers associated with stem traits in alfalfa

Alfalfa biomass can be fractionated into leaf and stem components. Leaves comprise a protein-rich and highly digestible portion of biomass for ruminant animals, while stems constitute a high fiber and less digestible fraction, representing 50 to 70% of the biomass. However, little attention has focused on stem-related traits, which are a key aspect in improving the nutritional value and intake potential of alfalfa. This study aimed to identify molecular markers associated with four morphological traits in a panel of five populations of alfalfa generated over two cycles of divergent selection based on 16-h and 96-h in vitro neutral detergent fiber digestibility in stems. Phenotypic traits of stem color, presence of stem pith cells, winter standability, and winter injury were modeled using univariate and multivariate spatial mixed linear models (MLM), and the predicted values were used as response variables in genome-wide association studies (GWAS). The alfalfa panel was genotyped using a 3K DArTag SNP markers for the evaluation of the genetic structure and GWAS. Principal component and population structure analyses revealed differentiations between populations selected for high- and low-digestibility. Thirteen molecular markers were significantly associated with stem traits using either univariate or multivariate MLM. Additionally, support vector machine (SVM) and random forest (RF) algorithms were implemented to determine marker importance scores for stem traits and validate the GWAS results. The top-ranked markers from SVM and RF aligned with GWAS findings for solid stem pith, winter standability, and winter injury. Additionally, SVM identified additional markers with high variable importance for solid stem pith and winter injury. Most molecular markers were located in coding regions. These markers can facilitate marker-assisted selection to expedite breeding programs to increase winter hardiness or stem palatability. Author SummaryThe alfalfa stem constitutes a significant portion of forage yield, accounting for 50 to 70% of biomass yield, and influences various traits including plant height, standability, and digestibility. In our study, we identified thirteen molecular markers associated with stem color, presence of stem pith parenchyma, winter standability, and winter injury in a panel of five divergent stem digestibility populations. Multivariate trait modeling enhances the correlation among traits, thereby expanding the number of markers associated via GWAS. Similarly, machine learning algorithms increase the confidence of markers initially identified by GWAS and uncover new candidate regions that could serve as associated markers. Genes harboring markers associated to the four stem traits play roles in plant growth, response to plant injury, or tolerance to cold, underscoring their potential utility in enhancing traits such as cold tolerance and forage quality in alfalfa.

genomics↗

bioRxiv2024

Tap into non-symbiotic carbon? Exogenous myristate fuels the growth of symbiotic arbuscular mycorrhizal fungi but disrupts their carbon-phosphorus exchange with host plants

Arbuscular mycorrhizal fungi (AMF) are obligate biotrophs that rely on symbiotic carbohydrates and, in particular, lipids derived from their host plants. However, it remains unclear whether symbiotic AMF can access exogenous non-symbiotic lipids in the presence of plant-derived carbon, complicating our understanding of their relationship with host plants. Here, we investigated the direct uptake of exogenous 13C1-labeled myristate by three symbiotic AMF species (Rhizophagus irregularis, R. intraradices, and R. diaphanous) and assessed their growth responses using AMF-carrot hairy root co-culture systems. Furthermore, we explored the environmental distribution of myristate, and evaluated the impact of exogenous myristate on the carbon-phosphorus exchange between R. irregularis and alfalfa or rice in a greenhouse experiment. Our results showed that symbiotic AMF can absorb exogenous myristate, as evidenced by 13C enrichment and transcriptional activation of fatty acid transport and metabolism genes in AMF extraradical hyphae. Myristate is commonly present in various soil and plant environments, and its application increased both intraradical and extraradical fungal biomass, possibly linked to suppressed mycorrhizal-activated defense responses in host roots. Unexpectedly, exogenous myristate reduced the mycorrhizal phosphorus benefits for both alfalfa and rice and decreased their symbiotic carbon allocation to root-colonizing AMF, suggesting that the application of exogenous myristate may not be a promising strategy to enhance AM symbiosis. These findings provide new insights into understanding and manipulating the nutritional interactions between AMF and host plants.

ecology↗

bioRxiv2024

Transcriptome-wide identification of 5-methylcytosine by deaminase and reader protein-assisted sequencing

5-Methylcytosine (m5C) is one of the post-transcriptional modifications in mRNA and is involved in the pathogenesis of various diseases. However, the capacity of existing assays for accurately and comprehensively transcriptome-wide m5C mapping still needs improvement. Here, we develop a detection method named DRAM (deaminase and reader protein assisted RNA methylation analysis), in which deaminases (APOBEC1 and TadA-8e) are fused with m5C reader proteins (ALYREF and YBX1) to identify the m5C sites through deamination events neighboring the methylation sites. This antibody-free and bisulfite-free approach provides transcriptome-wide editing regions which are highly overlapped with the publicly available BS-seq datasets and allows for a more stable and comprehensive identification of the m5C loci. In addition, DRAM system even supports ultra-low input RNA (10ng). We anticipate that the DRAM system could pave the way for uncovering further biological functions of m5C modifications.

biochemistry↗

bioRxiv2024

ScRNA-seq and scATAC-seq reveal that sertoli cell mediate spermatogenesis disorders through stage-specific communications in non-obstructive azoospermia

Non-obstructive azoospermia (NOA) belongs to male infertility due to spermatogenesis failure. However, evidence for cell type-specific abnormalities of spermatogenesis disorders in NOA remains lacking. We performed single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) on testicular tissues from patients with obstructive azoospermia(OA) and NOA. HE staining confirmed the structural abnormalities of the seminiferous tubules in NOA patients. We identified 12 germ cell subtypes (spermatogonial stem cell-0 (SSC0), SSC1, SSC2, diffing-spermatogonia (Diffing-SPG), diffed-spermatogonia (Diffed-SPG), pre-leptotene (Pre-Lep), leptotene-zygotene (L-Z), pachytene (Pa), diplotene (Di), spermatids-1 (SPT1), SPT2, and SPT3) and 8 Sertoli cell subtypes (SC1-SC8). Among them, three novel Sertoli cell subtypes phenotypes were identified, namely SC4/immature, SC7/mature, and SC8/further mature Sertoli cells. For each germ or Sertoli cell subtype, we identified unique new markers, among which immunofluorescence confirmed co-localization of ST3GAL4, A2M, ASB9, and TEX19 and DDX4 (classical marker of germ cell). PRAP1, BST2, and CCDC62 were co-expressed with SOX9 (classical marker of Sertoli cell) in testes tissues also confirmed by immunofluorescence. The interaction between germ cell subtypes and Sertoli cell subtypes exhibits stage-specific-matching pattern, as evidenced by SC1/2/5/7 involving in SSC0-2 development, SC3 participating in the whole process of spermiogenesis, SC4/6 participating in Diffing and Diffed-SPG development, and SC8 involving in the final stage of SPT3. This pattern of specific interactions between subtypes of germ cell and Sertoli cell was confirmed by immunofluorescence of novel markers in testes tissues. The interaction was mainly contributed by Notch1/2/3 signaling. Our study profiled the single-cell transcriptome of human spermatogenesis and provided many potentials molecular markers for developing testicular puncture specific marker kits for NOA patients. Impact statementThe specific interactions between Sertoli cell subtypes and different stages of germ subgroups maintain normal development of germ cells. SC3 subtype absence may be the key factor leading to NOA.

cell biology↗

bioRxiv2024

CADTAD: CAncer Driver Topologically Associated Domains identify oncogenic and tumor suppressive lncRNAs

Cancer long noncoding RNAs (lncRNAs) have been identified by experimental and in silico methods. However, current approaches for identifying cancer lncRNAs are not sufficient and effective. To uncover them, we focused on the core cancer driver lncRNAs, which directly interact with cancer driver protein-coding genes (PCGs). We investigated various aspects of cancer lncRNAs, including their expression patterns, genomic locations, and direct interactions with cancer driver PCGs, and developed a pipeline to unearth candidate cancer driver lncRNAs. Finally, we validated the reliability of potential cancer driver lncRNAs through functional analysis of bioinformatics data and CRISPR-Cas9 knockout experiments. We found that cancer lncRNAs were more concentrated in cancer driver topologically associated domains (CDTs), and CDT is an important feature in identifying cancer lncRNAs. Moreover, cancer lncRNAs showed a high tendency to co-express with and bind to cancer driver PCGs. Utilizing these distinctive characteristics, we developed a pipeline CADTAD to unearth candidate cancer driver lncRNAs in pan-cancer, including 256 oncogenic lncRNAs, 177 tumor suppressive lncRNAs, and 75 dual-function lncRNAs, as well as in three individual cancer types, and validated their cancer-related function. More importantly, the function of 10 putative cancer driver lncRNAs in prostate cancer was subsequently validated to influence cancer phenotype through cell studies. In light of these findings, our study offers a new perspective from the 3D genome to study the roles of lncRNAs in cancer. Furthermore, we provide a valuable set of potential lncRNAs that could deepen our understanding of the oncogenic mechanism of cancer driver lncRNAs.

genomics↗