Biology subjects

Lin, X.

Publications and source records attributed to Lin, X..

At least 163 records · Page 9Linked to original sources

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↗

bioRxiv2024

A hemagglutinin and neuraminidase biased immunological memory shapes the dynamics of antibody responses to Influenza A virus

Influenza A virus (IAV) infection establishes a more diverse immunological memory to different viral proteins compared to vaccination. We hypothesized that the relative abundance of pre-existing immune memory to different viral antigens could skew post-infection antibody responses. To explore this, we generated mouse models with either an IAV hemagglutinin (HA)- or neuraminidase (NA)-biased immunological memory. We inoculated groups of mice with cocktails of isogenic viruses bearing antigenically-distinct HA (H3v) or NA (N2v) chosen to span the IAV H3N2 human circulation history. We challenged the mice with two H3N2 strains of opposing virulence and antigenic distance (AD) and examined the post-infection antibody landscapes. In both challenges, immune-naive mice seroconverted to both HA and NA whereas in primed mice, antibody response was detected to the antigen for which there is no pre-existing memory. In cases where the homologous antibody response was blunted, there was diversification on the breadth of response to antigenically-related strains with low baseline titers. Our findings clarifies the concept of "original antigenic sin" and demonstrate a mechanism by which the dynamics of antibody responses to HA and NA after infection can be altered by pre-existing immunity.

immunology↗

bioRxiv2024

Inactive Parp2 causes Tp53-dependent lethal anemia by blocking replication-associated nick ligation in erythroblasts

PARP1&2 enzymatic inhibitors (PARPi) are promising cancer treatments. But recently, their use has been hindered by unexplained severe anemia and treatment-related leukemia. In addition to enzymatic inhibition, PARPi also trap PARP1&2 at DNA lesions. Here, we report that unlike Parp2-/- mice, which develop normally, mice expressing catalytically-inactive Parp2 (E534A, Parp2EA/EA) succumb to Tp53- and Chk2-dependent erythropoietic failure in utero, mirroring Lig1-/- mice. While DNA damage mainly activates PARP1, we demonstrate that DNA replication activates PARP2 robustly. PARP2 is selectively recruited and activated by 5-phosphorylated nicks (5p-nicks) between Okazaki fragments, typically resolved by Lig1. Inactive PARP2, but not its active form or absence, impedes Lig1- and Lig3-mediated ligation, causing dose-dependent replication fork collapse, particularly harmful to erythroblasts with ultra-fast forks. This PARylation-dependent structural function of PARP2 at 5p-nicks explains the detrimental effects of PARP2 inhibition on erythropoiesis, revealing the mechanism behind the PARPi-induced anemia and leukemia, especially those with TP53/CHK2 loss. SignificanceThis work shows that the hematological toxicities associated with PARP inhibitors stem not from impaired PARP1 or PARP2 enzymatic activity but rather from the presence of inactive PARP2 protein. Mechanistically, these toxicities reflect a unique role of PARP2 at 5-phosphorylated DNA nicks during DNA replication in erythroblasts.

genetics↗

bioRxiv2024

Cannabinoid CB2 receptors in primary sensory neurons are implicated in CB2 agonist-mediated suppression of paclitaxel-induced neuropathic nociception and sexually-dimorphic sparing of morphine tolerance

Cannabinoid CB2 agonists show therapeutic efficacy without the unwanted side effects commonly associated with direct activation of CB1 receptors. The G protein-biased CB2 receptor agonist LY2828360 attenuates the maintenance of chemotherapy-induced neuropathic nociception in male mice and blocks the development of morphine tolerance in this model. However, the specific cell types involved in this phenomenon have never been investigated and whether this therapeutic profile is observed in female mice remains poorly understood. We used conditional deletion of CB2 receptors from specific cell populations to determine the population(s) mediating the anti-allodynic and morphine-sparing effects of CB2 agonists. Anti-allodynic effects of structurally distinct CB2 agonists (LY2828360 and AM1710) were present in paclitaxel-treated CB2f/f mice of either sex. The anti-allodynic effect of the CB2 agonists were absent in conditional knockout (KO) mice lacking CB2 receptors in peripheral sensory neurons (AdvillinCRE/+; CB2f/f) but preserved in mice lacking CB2 receptors in CX3CR1 expressing microglia/macrophages (CX3CR1CRE/+; CB2f/f). The morphine-sparing effect of LY28282360 occurred in a sexually-dimorphic manner, being present in male mice but absent in female mice of any genotype. In mice with established paclitaxel-induced neuropathy, prior LY2828360 treatment (3 mg/kg per day i.p. x 12 days) blocked the subsequent development of morphine tolerance in male CB2f/f mice but was absent in male (or female) AdvillinCRE/+; CB2f/f mice. LY2828360-induced sparing of morphine tolerance was preserved in male CX3CR1CRE/+; CB2f/f mice, but this effect was not observed in female CX3CR1CRE/+; CB2f/f mice. Similarly, co-administration of morphine with a low dose of LY2828360 (0.1 mg/kg per day i.p. x 6 days) reversed tolerance to the anti-allodynic efficacy of morphine in paclitaxel-treated male CB2f/f mice, but this effect was absent in female CB2f/f mice and AdvillinCRE/+; CB2f/f mice of either sex. Additionally, LY2828360 (3 mg/kg per day i.p. x 8 days) delayed, but did not prevent, the development of paclitaxel-induced mechanical and cold allodynia in either CB2f/f or CX3CR1CRE/+; CB2f/f mice of either sex. Our studies reveal that CB2 receptors in primary sensory neurons are required for the anti-allodynic effects of CB2 agonists in a mouse model of paclitaxel-induced neuropathic nociception. We also find that CB2 agonists acting on primary sensory neurons produce a sexually-dimorphic sparing of morphine tolerance in males, but not female, paclitaxel-treated mice.

neuroscience↗

bioRxiv2024

TabHLH27 orchestrates root growth and drought tolerance to enhance water use efficiency in wheat

Cultivating high-yield wheat under limited water resources is essential for sustainable agriculture in semiarid regions. Amid water scarcity, plants activate drought response signaling, yet the delicate balance between drought tolerance and development remains unclear. Through genome-wide-association study (GWAS) and transcriptome profiling, we identified a wheat atypical basic helix-loop-helix (bHLH) transcription factor (TF), TabHLH27-A1, as a promising quantitative trait locus (QTL) candidate for both relative root dry weight (DW.R%) and spikelet number per spike (SPS) in wheat. TabHLH27-A1/B1/D1 knockout reduced wheat drought tolerance, yield, and water use efficiency (WUE). TabHLH27-A1 exhibited rapid induction with PEG treatment, gradually declining over days. It activated stress response genes such as TaCBL8-B1 and TaCPI2-A1 while inhibiting root growth genes like TaSH15-B1 and TaWRKY70-B1 under short-term PEG stimulus. The distinct transcriptional regulation of TabHLH27-A1 involved diverse interacting factors such as TaABI3-D1 and TabZIP62-D1. Natural variations of TabHLH27-A1 influences its transcriptional responses to drought stress, with TabHLH27-A1Hap-II associated with stronger drought tolerance, larger root system, more spikelets, and higher WUE in wheat. Significantly, the elite TabHLH27-A1Hap-II was selected during the breeding process in China, and introgression of TabHLH27-A1Hap-II allele improves drought tolerance and grain yield, especially under water-limited conditions. Our study highlights TabHLH27-A1s role in balancing root growth and drought tolerance, providing a genetic manipulation locus for enhancing WUE in wheat.

genetics↗

bioRxiv2024

Gustatory-neuron-supplied R-spondin-2 is required for taste bud replenishment

Taste buds undergo continuous cell turnover throughout life, and taste cell replenishment relies strictly on innervation, a phenomenon first described almost 150 years ago. Recently, we provided evidence that R-spondin 2 (Rspo2) may be the long-sought gustatory neuron-supplied factor that regulates taste stem cell activity, via its interaction with taste stem/progenitor cell-expressed receptor Rnf43/Znrf3. Yet, whether gustatory-neuron-supplied Rspo2 is strictly required for taste tissue maintenance has not been resolved. Here, we set out to determine the necessity of gustatory-neuron-supplied Rspo2 in taste tissue homeostasis using genetic approaches. We used a mouse line that harbors the neomycin-resistance gene (NeoR) in one of the intron regions of the Rspo2 gene, which results in reduced expression of Rspo2. The number of taste buds is significantly reduced in these mice, compared to wild-type mice, in both anterior and posterior tongue. This phenotypic change was completely reversed by removing NeoR from the Rspo2 gene, thus making it normal. We also combined adeno-associated virus (AAV)-based delivery of Cre recombinase with a mouse line amenable to Cre-based ablation of the Rspo2 exons encoding the receptor-binding domains. Such deletion of Rspo2 in the nodose-petrosal-jugular ganglion complex led to nearly complete loss of taste buds in the circumvallate papilla. Thus, we demonstrate that Rspo2 is the long-sought gustatory-neuron-supplied factor that acts on taste stem cells to maintain taste tissue homeostasis. SignificanceWe have known for 150 years that innervation is required to induce and maintain cell replacement in taste buds. Until recently, the identity of the inducing factor produced by neurons was unknown. We have shown that R-spondin alone is sufficient to substitute for neuronal input to induce taste bud regeneration. Using a genetic loss-of-function approach, we now demonstrate that gustatory-neuron-expressed Rspo2 is required to maintain taste tissue homeostasis. Altogether, our work reveals that Rspo2 is the long-sought neuron-supplied factor that regulates the activity of taste stem/progenitor cells.

neuroscience↗

bioRxiv2024

Chronic alcohol exposure induces cerebral microbleeds by TGF-β1/Smad signaling pathway mediated remodeling of cerebral small vessels

BackgroundLong-term heavy drinking is a major risk factor for cerebral microbleeds(CMBs), which are increasingly gaining attention as a pathological phenotype of cerebral small vessel diseases(CSVD). Under pathological conditions, remodeling of the extracellular matrix(ECM) on the walls of small vessels causes disarray in the structure and function of these vessels, leading to cerebral small vessel sclerosis and consequent rupture and bleeding. This can result in cognitive and emotional disorders, abnormal gait and increased risk of falling. However, the mechanisms underlying how long-term alcohol consumption leads to CMBs and decline in motor function remain unknown. MethodsWe constructed a chronic alcohol exposure mouse model and measured the deposition of ECMs on the small vessels in motor-related brain regions. The presence of microbleeds was confirmed through Prussian blue staining and Magnetic Resonance Imaging. We also extracted primary cerebral microvascular smooth muscle cells (CMVSMCs) from the newborn mice and explored the effects of alcohol on the phenotypic transformation and substance synthesis function. Additionally, we conducted interventional experiments on the cell and animal models with an anti-fibrotic drugs Pirfenidone(PFD). ResultsWe found that mice with long-term alcohol exposure showed decreased motor function. In their motor-related brain regions, such as the motor cortex(MC), thalamus/basal ganglia(Tha/BG), and cerebellum(CB), we observed microbleeds. On the small vessels in these areas, we detected excessive deposited ECM proteins. In vitro experiments with primary CMVSMCs revealed that after alcohol treatment, the cells underwent a transformation into fibroblast-like phenotypes, and excessive production of the aforementioned ECM proteins, which is regulated by upstream TGF{beta}1/Smad signaling pathway. Additionally, PFD applied on cell and animal models could reverse the above processes to some extent. ConclusionsOur study found that the remodeling of ECM accompanied by activation of TGF-{beta}1/Smad signaling pathway may be involved in alcohol-induced CMBs. It could be a potential therapeutic target for CMBs or CSVD.

molecular biology↗

bioRxiv2024

FABP4 as a Therapeutic Host Target Controlling SARS-CoV2 Infection

Host metabolic fitness is a critical determinant of infectious disease outcomes. Obesity, aging, and other related metabolic disorders are recognized as high-risk disease modifiers for respiratory infections, including coronavirus infections, though the underlying mechanisms remain unknown. Our study highlights fatty acid-binding protein 4 (FABP4), a key regulator of metabolic dysfunction and inflammation, as a modulator of SARS-CoV-2 pathogenesis, correlating strongly with disease severity in COVID-19 patients. We demonstrate that loss of FABP4 function, by genetic or pharmacological means, reduces SARS-CoV2 replication and disrupts the formation of viral replication organelles in adipocytes and airway epithelial cells. Importantly, FABP4 inhibitor treatment of infected hamsters diminished lung viral titers, alleviated lung damage and reduced collagen deposition. These findings highlight the therapeutic potential of targeting host metabolism in limiting coronavirus replication and mitigating the pathogenesis of infection.

microbiology↗

bioRxiv2024

Stimulated Prorocentrum donghaiense cell growth by in-situ mariculture dissolved organic matter

Mixotrophic dinoflagellates frequently cause harmful algal blooms (HAB) in eutrophic mariculture waters that contain diverse excreted dissolved organic matter (DOM). The phagotrophy and the utilization of single organic small molecules have been extensively investigated in the bloom-forming mixotrophic dinoflagellates. However, their ability to utilize the in-situ DOM via absorbtrophy still remains unexplored. Here we examined the growth promotion effect of the in-situ mariculture DOM on Prorocentrum donghaiense, a representative HAB-forming species in coastal waters. Our results showed that the cell growth and photosynthesis of P. donghaiense were significantly promoted under in-situ DOM culture conditions. Additionally, parallel cultures were set up to disclose the potential role of the bacterioplankton in the free-living community (helper), where they aid in the remineralization of the in-situ DOM, and the phycosphere community (competitor), where they compete against the algal host to acquire nutrients from the in-situ DOM. Meanwhile, we determined the cellular stoichiometry under different culture conditions, showing that mariculture DOM can shape cellular stoichiometry significantly. Elevated cellular N (84.96%) and P (48.3%) were observed in spring DOM groups compared with the control groups. For the first time, this study quantifies the efficient utilization of the in-situ DOM via absorbtrophy, indicating the vital role in the outbreak and maintenance of HAB events.

ecology↗

bioRxiv2024

A single-cell 3D spatiotemporal multi-omics atlas from Drosophila embryogenesis to metamorphosis

The development of a multicellular organism is a highly intricate process tightly regulated by numerous genes and pathways in both spatial and temporal manners. Here, we present Flysta3D, a comprehensive multi-omics atlas of the model organism Drosophila, spanning its developmental lifespan from embryo to pupa. Our datasets encompass 3D single-cell spatial transcriptomic, single-cell transcriptomic, and single-cell chromatin accessibility information. By integrating these multi-dimensional data, we constructed cell state trajectories that uncover the detailed profiles of tissue development. With a focus on the central nervous system (CNS) and midgut, we dissected the spatiotemporal dynamics of gene regulatory networks, cell type diversity, and morphological changes from a multi-omics perspective. This extensive atlas provides an unprecedentedly rich resource and serves as a systematic platform for studying Drosophila development with integrated single-cell data at an ultra-high spatiotemporal resolution.

developmental biology↗

bioRxiv2024

Cortico-spinal Mechanisms of Periphery Neuromodulation induced Analgesia

BackgroundTranscutaneous Electrical Nerve Stimulation (TENS) is widely used for pain management, yet how different parameters--Conventional (high frequency, low intensity) versus Acupuncture-Like (low frequency, high intensity)--modulate pain remains controversial. These modes produce variable analgesic effects, but their underlying brain-spinal mechanisms are unclear. Clarifying these pathways could refine TENS protocols, optimize clinical outcomes, and reduce opioid reliance, highlighting the need for more precise, non-pharmacological approaches to advanced pain care. MethodsThis single-blind, three-arm randomized trial recruited 95 healthy adults (18-30 years) and assigned them (1:1:1) to Conventional TENS, Acupuncture-Like TENS, or sham. Participants underwent a 30-minute TENS intervention on the left forearm (C5-C6 dermatome) with simultaneous brain-spinal fMRI before and after thermal nociceptive stimuli. The primary outcomes were changes in Numeric Rating Scale (NRS) pain scores and brain-spinal BOLD signals. Secondary outcomes included psycho-physiological interaction (PPI) and mediation analyses of periaqueductal gray (PAG) activity and brain- cord connectivity. FindingsBoth TENS modes significantly reduced pain but engaged distinct cortico-spinal pathways. Conventional TENS yielded local analgesia via dlPAG-driven spinal inhibition plus partial cortical involvement (PAG-vmPFC). By contrast, Acupuncture-Like TENS produced diffuse analgesia through vlPAG-linked top-down modulation reliant on spinal gating. Correlation and mediation analyses confirmed that Conventional TENS integrates spinal and cortical synergies, whereas Acupuncture-Like TENS is dominated by robust descending control. InterpretationDifferent TENS parameters yield distinct analgesic mechanisms. Conventional TENS couples direct spinal inhibition with partial cortical regulation, while Acupuncture-Like TENS relies heavily on top-down pathways passing through the spinal cord. Recognizing these unique descending networks can guide targeted TENS protocols for diverse pain conditions, optimizing clinical outcomes and reducing reliance on pharmacological approaches. FundingsThis work was supported by the National Key R&D Program of China (2022YFC3500603), the National Natural Science Foundation of China (32071061, 82072010, 82030121 and 32100861, 82330057), the Beijing Natural Science Foundation (IS23108, JQ22018).

neuroscience↗

bioRxiv2024

Human Amniotic Epithelial Cells Promote Chx10-/Pax6+ Muller Glia Subpopulation Reprogramming into Photoreceptor-like Cells

Reprogramming Muller glia to regenerate neurons is a promising strategy for treating retinal degeneration, but whether Muller glia contain subpopulations with different regenerative fates remains unclear. Here, using single-cell RNA-seq analysis and Muller glia lineage-tracing mice with retinal degeneration, we reveal that Muller glia were heterogeneous and identify a specific Muller glial subpopulation (Chx10-/Pax6+) in healthy retinas that is activated and migrate to the outer nuclear layer (ONL) during photoreceptor degeneration. Transplantation of human amniotic epithelial cells (hAECs) facilitates the activation and extensive migration of the Chx10-/Pax6+ Muller glial subpopulation to the ONL, where they are reprogrammed into photoreceptor-like cells. Mechanistically, hAECs degrade the inhibitory extracellular matrix through regulating matrix metalloproteinases, which probably induces remodeling of the microenvironment of Muller glia and contributes to cell reprogramming. Consequently, hAEC transplantation improves visual function in rd10 mice. Our findings uncover a distinctive Muller glial subpopulation with the potential for reprogramming into photoreceptors.

cell biology↗

bioRxiv2024

Charting the Spatial Transcriptome of the Human Cerebral Cortex at Single-Cell Resolution

In our pursuit of creating a comprehensive human cortical atlas to understand human intelligence, we examined the single-nuclei transcriptomes of 307,738 cells alongside spatial transcriptomics data from 46,948 VISIUM spots and 1,355,582 Stereo cells. Atlases reveal distinct expression patterns and spatial arrangements of cortical neural cell types. Glutamatergic neurons exhibit precise laminar patterns, often mirroring expression patterns in adjacent cortical regions. Overlaying our atlas with functional networks delineated substantial correlations between neural cell types and cortical region function. Notably, regions involved in processing sensory information (pain) display a pronounced accumulation of extratelencephalic neurons. Additionally, our atlas enabled precise localization of the thicker layer 4 of the visual cortex and an in-depth study of the stabilize the subplate structure, known as layer 6b, revealed specific marker genes and cellular compositions. Collectively, our research sheds light on the cellular foundations of the intricate and intelligent regions within the human cortex.

neuroscience↗

bioRxiv2024

Loss of ZNRF3/RNF43 Unleashes EGFR in Cancer

ZNRF3 and RNF43 are closely related transmembrane E3 ubiquitin ligases with significant roles in development and cancer. Conventionally, their biological functions have been associated with regulating WNT signaling receptor ubiquitination and degradation. However, our proteogenomic studies have revealed EGFR as the protein most negatively correlated with ZNRF3/RNF43 mRNA levels in multiple human cancers. Through biochemical investigations, we demonstrate that ZNRF3/RNF43 interact with EGFR via their extracellular domains, leading to EGFR ubiquitination and subsequent degradation facilitated by the E3 ligase RING domain. Overexpression of ZNRF3 reduces EGFR levels and suppresses cancer cell growth in vitro and in vivo, whereas knockout of ZNRF3/RNF43 stimulates cell growth and tumorigenesis through upregulated EGFR signaling. Together, these data highlight ZNRF3 and RNF43 as novel E3 ubiquitin ligases of EGFR and establish the inactivation of ZNRF3/RNF43 as a driver of increased EGFR signaling, ultimately promoting cancer progression. This discovery establishes a connection between two fundamental signaling pathways, EGFR and WNT, at the level of cytoplasmic membrane receptors, uncovering a novel mechanism underlying the frequent co-activation of EGFR and WNT signaling in development and cancer.

cancer biology↗

bioRxiv2024

Unraveling wheat endosperm development: epigenetic regulation and novel regulators for enhanced yield and quality

Starch content and seed storage protein (SSP) composition are critical factors influencing wheat grain yield and quality. To uncover the molecular mechanisms governing their biosynthesis, we conducted transcriptome and epigenome profiling across key endosperm developmental stages, revealing that chromatin accessibility, H3K27ac, and H3K27me3 collectively regulate SSP and starch genes with varying impact. Population transcriptome and phenotype analyses highlighted the crucial role of accessible promoter regions as a genetic variation resource, influencing grain yield and quality in a core collection of wheat accessions. By integrating time-serial RNA-seq and ATAC-seq data, we constructed a hierarchical transcriptional regulatory network (TRN) governing starch and SSP biosynthesis, identifying 42 high-confidence novel candidates. These candidates exhibited overlap with genetic regions associated with grain size and quality traits, and their functional significance was validated through expression-phenotype association analysis among wheat accessions and TILLING mutants. In-depth functional analysis of wheat abscisic acid insensitive 3-A1 (TaABI3-A1) with genome editing knock-out lines demonstrated its role in promoting SSP accumulation while repressing starch biosynthesis through transcriptional regulation. An elite haplotype of TaABI3-A1 with higher grain weight was identified during the breeding process in China, and its superior trait was associated with altered TaABI3-A1 expression levels. Additionally, we identified the potential upstream regulator, wheat GAGA-binding transcription factor 1 (TaGBP1), influencing TaABI3-A1 expression. Our study provides novel and high-confidence regulators, presenting an effective strategy for understanding the regulation of SSP and starch biosynthesis and contributing to breeding enhancement.

plant biology↗

bioRxiv2023

TaNF-Y-PRC2 orchestrates temporal control of starch and protein synthesis in wheat

The endosperm in cereal grains is instrumental in determining grain yield and seed quality, as it controls the production of starch and protein. In this study, we identified a specific TaNF-Y trimeric complex, consisting of TaNF-YA3-D, TaNF-YB7-B, and TaNF-YC6-B, exhibiting robust expression within endosperm during grain filling stage in wheat. Knock-down of either TaNF-YA3 or TaNF-YC6 led to less starch but more gluten proteins. Detailed analyses have unveiled that the TaNF-Y indirectly boosts starch biosynthesis genes by reducing TaNAC019, a repressor of TaAGPS1a, TaSuS2, thereby regulating starch biosynthesis. Conversely, the TaNF-Y directly inhibits the expression of gliadin and low molecular weight (LMW)-GS coding genes, including TaGli-{gamma}-700 and TaLMW-400. Furthermore, the TaNF-Y components interact with TaSWN, the histone methyltransferase subunit of Polycomb repressive complex 2 (PRC2), to repress the expression of TaNAC019, TaGli-{gamma}-700, and TaLMW-400 through H3K27me3 modification. Notably, weak mutation of TaFIE, core subunit of PRC2, has reduced starch but elevated gliadin and LMW-GS levels. Intriguingly, DNA variations of TaNF-Y components are widely associated with seed developmental traits. In particular, variation within the coding region of TaNF-YB7-B is linked to differences in starch and protein content. Distinct haplotypes of TaNF-YB7-B affect its interaction with TaSWN, influencing the repression of targets like TaNAC019 and TaGli-{gamma}-700. Our findings illuminate the intricate molecular mechanisms governing epigenetic regulation by the TaNF-Y-PRC2 for wheat endosperm development. Manipulating the TaNF-Y complex holds potential for optimizing grain yield and enhancing quality.

plant biology↗

bioRxiv2023

Distinct roles of H3K27me3 and H3K36me3 in vernalization response, maintenance and resetting in winter wheat

Winter plants rely on vernalization, a vital process for adapting to cold and ensuring successful reproduction. However, understanding the role of histone modifications in guiding the vernalization process in winter wheat is limited. In this study, we investigate the transcriptome and chromatin dynamics in the shoot apex throughout the life cycle of winter wheat in the field. Two core histone modifications, H3K27me3 and H3K36me3, exhibit opposite pattern on the key vernalization gene VERNALIZATION1 (VRN1), correlated with its induction during cold exposure. Additionally, H3K36me3 remains high at VRN1 after cold exposure, maintaining its active state. Mutations in FERTILIZATION-INDEPENDENT ENDOSPERM (TaFIE) and SET DOMAIN GROUP 8 (TaSDG8), writer complex components of H3K27me3 and H3K36me3, respectively, affect flowering time. Interestingly, VRN1 loses its high expression after cold exposure memory in the absence of H3K36me3. During embryo development, VRN1 is silenced with the removal of H3K36me3 in both winter and spring alleles. H3K27me3 is selectively added to the winter allele, influencing the cold exposure requirement for the next generation. Integrating gene expression with H3K27me3 and H3K36me3 patterns identified potential regulators of flowering. This study reveals distinct roles of H3K27me3 and H3K36me3 in controlling vernalization response, maintenance, and resetting in winter wheat. Significance StatementVernalization, initially observed in cereals, lacks a comprehensive understanding of its underlying mechanism, particularly regarding chromatin-mediated transcriptional regulation in winter wheat. By delving into the transcriptome and chromatin dynamics in the shoot apex throughout winter wheats life cycle, we pinpointed two crucial histone modifications, H3K27me3 and H3K36me3, each playing distinct roles at different vernalization stages. H3K27me3 is implicated in establishing and resetting the extended cold exposure requirement for winter wheat, gradually diminishing during vernalization. On the other hand, H3K36me3 is crucial for maintaining VRN1s active state post-cold exposure, contributing to the memory of the vernalization treatment. Additionally, the integration of transcriptome and histone modification profiles unveiled potential novel regulators of flowering in winter wheat.

plant biology↗

bioRxiv2023

PUPpy: a primer design pipeline for substrain-level microbial detection and absolute quantification.

Characterizing microbial communities at high-resolution and with absolute quantification is crucial to unravel the complexity and diversity of microbial ecosystems. This can be achieved with PCR assays, which enable highly selective detection and absolute quantification of microbial DNA. However, a major challenge that has hindered PCR applications in microbiome research is the design of highly specific primer sets that exclusively amplify intended targets. Here, we introduce Phylogenetically Unique Primers in python (PUPpy), a fully automated pipeline to design microbe- and group-specific primers within a given microbial community. PUPpy can be executed from a user-friendly GUI, or two simple terminal commands, and it only requires coding sequence files of the community members as input. PUPpy-designed primers enable the detection of individual microbes and quantification of absolute microbial abundance in defined communities below the strain level. We experimentally evaluated the performance of PUPpy-designed primers using two bacterial communities as benchmarks. Each community was comprised of 10 members, exhibiting a range of genetic similarities that spanned from different phyla to substrains. PUPpy-designed primers also enable the detection of groups of bacteria in an undefined community, such as the detection of a gut bacterial family in a complex stool microbiota sample. Taxon-specific primers designed with PUPpy showed 100% specificity to their intended targets, without unintended amplification, in each community tested. Lastly, we show absolute quantification of microbial abundance using PUPpy-designed primers in ddPCR, benchmarked against 16S rRNA and shotgun sequencing. Our data shows that PUPpy-designed microbe-specific primers can be used to quantify substrain-level absolute counts, providing more resolved and accurate quantification in defined communities than short-read 16S rRNA and shotgun sequencing. ImportanceProfiling microbial communities at high resolution and with absolute quantification is essential to uncover hidden ecological interactions within microbial ecosystems. Nevertheless, achieving resolved and quantitative investigations has been elusive due to methodological limitations in distinguishing and quantifying highly related microbes. Here, we describe PUPpy, an automated computational pipeline to design taxon-specific primers within defined microbial communities. Taxon-specific primers can be used to selectively detect and quantify individual microbes and larger taxa within a microbial community. PUPpy achieves substrain-level specificity without the need for computationally intensive databases and prioritises user-friendliness by enabling both terminal and graphical user interface (GUI) applications. Altogether, PUPpy enables fast, inexpensive, and highly accurate perspectives into microbial ecosystems, supporting the characterization of bacterial communities in both in vitro and complex microbiota settings.

microbiology↗