Differential Gene Regulation Research Articles

Phosphorylation-dependent activation of the bHLH transcription factor ICE1/SCRM promotes polarization of the Arabidopsis zygote.

In Arabidopsis thaliana, the asymmetric cell division (ACD) of the zygote gives rise to the embryo proper and an extraembryonic suspensor, respectively. This process is controlled by the ERECTA-YODA-MPK3/6 receptor kinase-MAP kinase-signaling pathway, which also orchestrates ACDs in the epidermis. In this context, the bHLH transcription factor ICE1/SCRM is negatively controlled by MPK3/6-directed phosphorylation. However, it is unknown whether this regulatory module is similarly involved in the zygotic ACD. We investigated the function of SCRM in zygote polarization by analyzing the effect of loss-of-function alleles and variants that cannot be phosphorylated by MPK3/6, protein accumulation, and target gene expression. Our results show that SCRM has a critical function in zygote polarization and acts in parallel with the known MPK3/6 target WRKY2 in activating WOX8. Our work further demonstrates that SCRM activity in the early embryo is positively controlled by MPK3/6-mediated phosphorylation. Therefore, the effect of MAP kinase-directed phosphorylation of the same target protein fundamentally differs between the embryo and the epidermis, shedding light on cell type-specific, differential gene regulation by common signaling pathways.

Integrative Transcriptomic and Metabolomic Analysis Reveals Quinoa Leaf Response Mechanisms to Different Phosphorus Concentrations During Filling Stage

Quinoa is an annual self-pollinating plant rich in polyphenols, flavonoids, saponins, and amino acids; its protein balance closely aligns with the ideal recommendation set by the Food and Agriculture Organization. Therefore, quinoa is considered the most suitable “all-nutrient food”. Phosphorus fertilization plays an important role in restricting the growth and development of quinoa; however, the effects of phosphorus fertilizer on quinoa growth remain unclear. Therefore, we conducted metabolome and transcriptome analyses on quinoa leaves during the filling stage, subjecting plants to different doses of phosphorus fertilizer. Overall, phosphorus treatment exerted a significant impact on the phenotypic characteristics of quinoa. Specifically, through a combined analysis of ultra-performance liquid chromatography–tandem mass spectrometry and transcriptome analysis, we identified the alteration and regulation of specific metabolites and genes within flavonoid biosynthesis pathways; this comprehensive evaluation helped elucidate the response mechanism of quinoa leaves during the grouting stage under various phosphorus conditions. Ultimately, the results of this study provide a reference for the selection of quinoa cultivars that exhibit tolerance to low- or high-phosphorus stress; additionally, we offer a theoretical basis for the rational application of phosphorus fertilizer and the enhancement of phosphorus utilization efficiency.

Deciphering temporal gene expression dynamics during epilepsy development using a rat model of focal neocortical epilepsy.

Epilepsy involves significant changes in neural cells during epileptogenesis. Although the molecular mechanism of epileptogenesis remains obscure, changes in gene regulation play a crucial role in the evolution of epilepsy. This study aimed to compare changes in a subset of specific genes during epilepsy development, focusing on the period after the first spontaneous seizure, to identify critical time windows for targeting different regulators. Using a rat model of acquired focal neocortical epilepsy induced by tetanus toxin, we characterized gene expression at acute, subacute, and chronic stages (48-72 h, 2 weeks, and 30 days after first spontaneous seizure, respectively), focusing on genes' potential contribution to epilepsy progression. We observed dynamic changes in the expression of these genes throughout the period after the first spontaneous seizure. Astrocytic reactions primarily occur early, before epilepsy is well established. Changes in Mtor (mammalian target of rapamycin) and Rest (repressor element 1 silencing transcription factor) signaling pathways are highly dynamic and correlated with the progression of epilepsy development. Ccl2 (chemokine C-C-motif ligand) is upregulated at the chronic stage, indicating activation of the neuroinflammatory pathway. Finally, Gabra5 (γ-aminobutyric acidergic signaling) is downregulated at the late stage after epilepsy is established. Surprisingly, changes in the expression of specific genes are linked to the time since the first seizure, rather than seizure frequency or duration. These results suggest that the regulation of specific genes is essentially stage-dependent during the development of epilepsy, highlighting the importance of targeting specific genes at appropriate stages of epilepsy development.

PATH-65. MONITORING MENINGIOMA CELL CYCLE ACTIVITY THROUGH DREAM-MMB ACTIVITY SWITCHING

Abstract Meningiomas account for over 40% of all primary intracranial brain tumors, with approximately one-fifth of these tumors recurring. Our lab has previously used molecular techniques to identify three subgroups: MenG A, B, and C, which have been independently validated. Interestingly, only MenG C tumors harbor recurring patients, making it an aggressive subtype of meningioma. One hallmark of MenG C tumors is the overexpression of a distinct set of genes that harbor in their promoter the CDE-CHR DNA-binding element, used by a transcriptional complex that in dormant cells functions as a repressor (DREAM). Before phosphorylation, this complex functions as an activator (MBB) in cycling cells. Upon a phosphorylation-dependent switch, the dual-specificity tyrosine kinase DYRK1A phosphorylates S28 on LIN52, which itself is dimerized with LIN9 of the core complex. Phosphorylation shifts the complex to DREAM, while lack thereof allows the core complex to interact with MYBL2 (now MBB) and activate the same genes repressed in quiescent cells. We seek to quantify this switch activity using an in vivo luciferase sensor using NanoBiT technology. Our biosensor is based on a split nanoluciferase, where one half is fused to the LIN52/LIN9 dimer and the other is fused to MYBL2. When MYBL2 binds to the LIN52/LIN9 dimer, a functional nanoluciferase is formed, and its activity can be measured using a luciferase-specific substrate. As proof of principle, we have increased cell proliferation by increasing FBS concentration in cultured cells as well as by blocking S28 phosphorylation using the DYRK1A-specific inhibitor Harmine. We screened all possible combinations (alternating NanoBiT fragments) for the highest luciferase activity after stimulation. In the future, this biosensor can be used to monitor cell cycle activity in vitro as well as in vivo using low-cost Luciferase reporter assays to better understand the regulation of specific genes in MenG C tumors.

Recombinant leptins affect lipid metabolism in hepatocytes of Cynoglossus semilaevis

Leptin is a peptide hormone primarily produced by adipose tissue, which plays a crucial role in regulating energy balance by controlling appetite and energy expenditure. To better understand the intricate physiological functions of leptin in hepatocytes in tongue sole (Cynoglossus semilaevis), this study presents an integrated transcriptomic and metabolomic analysis of tongue sole hepatocytes following stimulation with lepA and lepB. Comparative analysis identified 2971 and 2900 upregulated differentially expressed genes (DEGs) and 1895 and 1821 downregulated DEGs in response to lepA and lepB stimulation, respectively. Notably, 5706 genes were commonly regulated by both stimulations, suggesting overlapping functional roles of these two leptin proteins. GO and KEGG enrichment analysis indicated significant enrichment in immune response (JAK-STAT signaling pathway, NF-κB signaling pathway, etc.) and lipid metabolism pathways, such as fatty acid synthesis, with similar findings for lepB. Metabolomic analysis demonstrated clear separation between treatment and control groups, with 34 medium- to long-chain fatty acids and numerous differentially expressed metabolites (DEMs) identified. KEGG analysis of the metabolome paralleled the transcriptomic findings, with shared pathways in lipid metabolism and immune function. Finally, joint analysis highlighted co-enrichment in linoleic acid metabolism and leishmaniasis pathways. Detailed mechanistic insights revealed the activation of JAK-STAT and NF-κB signaling pathways, modulation of arachidonic acid metabolism, and the influence on the MAPK signaling pathway. Additionally, lepA uniquely co-enriched in the fatty acid synthesis pathway, with specific gene regulation affecting the synthesis of various fatty acids. The study concluded that lepA and lepB exerted significant regulatory effects on lipid metabolism and immune responses in tongue sole hepatocytes through complex molecular networks. To our current knowledge, this represents the first direct evidence of leptin's role in immune function within teleost fish and offers valuable insights into the molecular mechanisms of lipid metabolism underlying the actions of lepA and lepB.

Transcriptomics combined with physiological analysis provided new insights into the enhancing Dendrobium nobile resilience to Cu stress through Ce modulation

In the face of escalating environmental challenges, understanding the mechanisms through which plants can resist heavy metal stress is paramount. This study delves into the efficacy of cerium (Ce), a rare earth element, in mitigating copper (Cu) toxicity in Dendrobium nobile Lindl., an orchid plant with significant medicinal and ornamental value. Through a comprehensive experimental approach that encompasses physiological, biochemical, and molecular analyses, we demonstrate that Ce application notably enhances D. nobile's tolerance to Cu stress. The investigation revealed that Ce supplementation significantly ameliorated the adverse effects of Cu on plant growth, evidenced by a notable decrease in Cu accumulation within the plant tissues. At the molecular level, transcriptome sequencing uncovered the upregulation of genes involved in antioxidant defense mechanisms, heavy metal chelation, and stress response signaling pathways. Key findings include the enhanced activities of antioxidant enzymes such as superoxide dismutase, peroxidase, and catalase, alongside an increase in the levels of antioxidants and phytochelatins, which collectively contribute to the detoxification of Cu. Furthermore, Ce was found to influence the subcellular distribution of Cu, promoting its sequestration in less harmful cellular compartments. Our study provides novel insights into the role of Ce in modulating plant responses to heavy metal stress, highlighting its potential as a mitigative agent against Cu toxicity. The elucidation of specific gene regulations and pathway activations offers a foundation for future research aimed at enhancing plant resilience to environmental stresses through biotechnological interventions.

Transcriptional regulation of hormone signalling genes in black pepper in response to Phytophthora capsici

IntroductionBlack pepper (Piper nigrum L.) is a non-model spice crop of significant agricultural and biological importance. The ‘quick wilt’ disease caused by the oomycete Phytophthora capsici is a major threat, leading to substantial crop loss. The molecular mechanisms governing the plant immune responses to this pathogen remain unclear. This study employs RNA sequencing and transcriptome analysis to explore the defense mechanisms of P. nigrum against P. capsici.ResultsTwo-month-old P. nigrum plantlets were subjected to infection with P. capsici, and leaf samples were collected at 6- and 12-hours post-inoculation. RNA was extracted, sequenced, and the resulting data were processed and assembled. Differential gene expression analysis was conducted to identify genes responding to the infection. Additionally, the study investigated the involvement of Salicylic acid (SA), Jasmonic acid (JA), and Ethylene (ET) signalling pathways. Our transcriptome assembly comprised 64,667 transcripts with 96.7% completeness, providing valuable insights into the P. nigrum transcriptome. Annotation of these transcripts identified functional categories and domains, provided details on molecular processes. Gene expression analysis identified 4,714 transcripts at 6 h post-infection (hpi) and 9,416 at 12 hpi as differentially expressed, revealing dynamic regulation of immune-related genes. Furthermore, the study investigated key genes involved in biosynthesis pathways of Salicylic acid, Jasmonic acid, and Ethylene signalling. Notably, we found differential regulation of critical genes associated with these pathways while comparing data before and after infection, thereby shedding light on their roles in defense mechanism in P. nigrum defense.ConclusionsThis comprehensive transcriptome analysis of P. nigrum response to P. capsici attack provides valuable insights into the plant defense mechanisms. The dynamic regulation of innate immunity and the involvement of key signalling pathways highlight the complexity of the plant-pathogen interaction. This study contributes to our understanding of plant immunity and offers potential strategies for enhancing P. nigrum resistance to this harmful pathogen.

Phylogenomic instructed target analysis reveals ELAV complex binding to multiple optimally spaced U-rich motifs.

ELAV/Hu RNA-binding proteins are gene-specific regulators of alternative pre-mRNA processing. ELAV/Hu family proteins bind to short AU-rich motifs which are abundant in pre-mRNA, making it unclear how they achieve gene specificity. ELAV/Hu proteins multimerize, but how multimerization contributes to decode degenerate sequence environments remains uncertain. Here, we show that ELAV forms a saturable complex on extended RNA. Through phylogenomic instructed target analysis we identify the core binding motif U5N2U3, which is repeated in an extended binding site. Optimally spaced short U5N2U3 binding motifs are key for high-affinity binding in this minimal binding element. Binding strength correlates with ELAV-regulated alternative poly(A) site choice, which is physiologically relevant through regulation of the major ELAV target ewg in determining synapse numbers. We further identify a stem-loop secondary structure in the ewg binding site unwound upon ELAV binding at three distal U motifs. Base-pairing of U motifs prevents ELAV binding, but N6-methyladenosine (m6A) has little effect. Further, stem-loops are enriched in ELAV-regulated poly(A) sites. Additionally, ELAV can nucleate preferentially from 3' to 5'. Hence, we identify a decisive mechanism for ELAV complex formation, addressing a fundamental gap in understanding how ELAV/Hu family proteins decode degenerate sequence spaces for gene-specific mRNA processing.

The gut microbiota and its metabolite butyrate shape metabolism and antiviral immunity along the gut-lung axis in the chicken

The gut microbiota exerts profound influence on poultry immunity and metabolism through mechanisms that yet need to be elucidated. Here we used conventional and germ-free chickens to explore the influence of the gut microbiota on transcriptomic and metabolic signatures along the gut-lung axis in poultry. Our results demonstrated a differential regulation of certain metabolites and genes associated with innate immunity and metabolism in peripheral tissues of germ-free birds. Furthermore, we evidenced the gut microbiota’s capacity to regulate mucosal immunity in the chicken lung during avian influenza virus infection. Finally, by fine-analysing the antiviral pathways triggered by the short-chain fatty acid (SCFA) butyrate in chicken respiratory epithelial cells, we found that it regulates interferon-stimulated genes (ISGs), notably OASL, via the transcription factor Sp1. These findings emphasize the pivotal role of the gut microbiota and its metabolites in shaping homeostasis and immunity in poultry, offering crucial insights into the mechanisms governing the communication between the gut and lungs in birds.

Study on molecular mechanism of intervertebral disc degeneration by single cell hdWGCNA combined with transcriptome sequencing

BackgroundIntervertebral disc degeneration (IVDD) is one of the important causes of lower back pain, seriously affecting people's health and quality of life. This research employs single-cell analysis to identify the specific cellular subtypes and key regulatory genes associated with IVDD. MethodsWe analyzed the single-cell data and screened cells that closely associated with the development of IVDD. The differential expression of feature genes between IVDD and control groups was analyzed. Additionally, drugs and regulatory transcription factors that interact with feature genes were predicted and clinically validated by reverse transcription quantitative real-time PCR (RT-qPCR), immunohistochemistry (IHC), and enzyme-linked immunosorbent assay (ELISA). ResultsOur study identified the Chond2 cell subtype associated with IVDD and selected four feature genes influencing the development of IVDD, namely IGFBP3, ACAN, VAPA and TMEM45A, through the high-dimensional weighted gene co-expression network analysis (hdWGCNA) analysis, least absolute shrinkage and selection operator (LASSO), and random forest (RF). Besides, compared to the MDD group, IGFBP3 and TMEM45A were significantly upregulated in the SDD group, while ACAN and VAPA showed no significant difference between the two groups. ELISA testing revealed a positive correlation between IGFBP3 concentration and the grading of IVDD. Furthermore, Celecoxib may be used to treat IVDD by inhibiting IGFBP3. ConclusionOur study identified the Chond2 cell subtype associated with IVDD and selected four feature genes influencing the development of IVDD, namely IGFBP3, ACAN, VAPA and TMEM45A. Our findings establish a robust theoretical foundation for the clinical diagnosis and treatment of IVDD patients.

Differential regulation of viral entry-associated genes modulated by inflammatory cytokines in the nasal epithelium.

This study aimed to investigate the impact of different types of nasal inflammation on the regulation of entry-associated genes of respiratory viruses, including severe acute respiratory syndrome coronavirus 2 (SARS CoV-2), Middle East respiratory syndrome coronavirus (MERS-CoV), human coronavirus 229E (HCoV-229E), and influenza virus, in the nasal epithelium. Subjects were classified into three groups: control, eosinophilic chronic rhinosinusitis (ECRS), and noneosinophilic CRS (NECRS) groups. Angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine subtype 2 (TMPRSS2), alanyl aminopeptidase (ANPEP), dipeptidyl peptidase 4 (DPP4), and beta-galactoside alpha-2,6-sialyltransferase 1 (ST6GAL1), and beta-galactoside alpha-2,3-sialyltransferase 4 (ST3GAL4) were selected as key entry-associated genes for SARS-CoV-2, HCoV-229E, MERS-CoV, and influenza, respectively, and were evaluated. Brushing samples obtained from each group and human nasal epithelial cells cultured using an air-liquid interface system were treated for 7 days with typical inflammatory cytokines and analyzed using real-time polymerase chain reaction. Western blot analysis and confocal microscopy were performed. The entry-associated genes showed distinct regulation patterns in response to each interleukin-4 (IL-4), interleukin-13 (IL-13), tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ). Specifically, ACE2 significantly decreased in type 2 cytokines (IL-4 and IL-13), while TMPRSS2 significantly decreased in type 1 cytokines (TNF-α and IFN-γ). ANPEP significantly decreased in both types of cytokines. Remarkably, DPP4 significantly increased in type 2 cytokines and decreased in type 1 cytokines. Moreover, ST6GAL1 and ST3GAL4 significantly increased in type 2 cytokines and decreased in type 1 cytokines, particularly IFN-γ. These findings were supported by western blot analysis and confocal imaging results, especially for ACE2 and DPP4. The findings regarding differential regulation suggest that patients with ECRS, primarily mediated by type 2 inflammation, may have lower susceptibility to SARS-CoV-2 and HCoV-229E infections but higher susceptibility to MERS-CoV and influenza infections.

Green Synthesis of Silver Nanoparticles and their Potential Applications in Mitigating Cancer.

In recent years, the field of nanotechnology has brought about significant advancements that have transformed the landscape of disease diagnosis, prevention, and treatment, particularly in the realm of medical science. Among the various approaches to nanoparticle synthesis, the green synthesis method has garnered increasing attention. Silver nanoparticles (AgNPs) have emerged as particularly noteworthy nanomaterials within the spectrum of metallic nanoparticles employed for biomedical applications. AgNPs possess several key attributes that make them highly valuable in the biomedical field. They are biocompatible, cost-effective, and environmentally friendly, rendering them suitable for various bioengineering and biomedical applications. Notably, AgNPs have found a prominent role in the domain of cancer diagnosis. Research investigations have provided evidence of AgNPs' anticancer activity, which involves mechanisms such as DNA damage, cell cycle arrest, induction of apoptosis, and the regulation of specific cytokine genes. The synthesis of AgNPs primarily involves the reduction of silver ions by reducing agents. Interestingly, natural products and living organisms have proven to be effective sources for the generation of precursor materials used in AgNP synthesis. This comprehensive review aims to summarize the key aspects of AgNPs, including their characterization, properties, and recent advancements in the field of biogenic AgNP synthesis. Furthermore, the review highlights the potential applications of these nanoparticles in combating cancer.

IL23R mutations associated with decreased risk of psoriasis lead to the differential expression of genes implicated in the disease.

Psoriasis is an incurable immune-mediated skin disease, affecting around 1%-3% of the population. Various lines of evidence implicate IL23 as being pivotal in disease. Genetic variants within the IL23 receptor (IL23R) increase the risk of developing psoriasis, and biologic therapies specifically targeting IL23 demonstrated high efficacy in treating disease. IL23 acts via the IL23R, signalling through the STAT3 pathway, mediating the cascade of events that ultimately results in the clinical presentation of psoriasis. Given the essential role of IL23R in disease, it is important to understand the impact of genetic variants on receptor function with respect to downstream gene regulation. Here we developed model systems in CD4+ (Jurkat) and CD8+ (MyLa) T cells to express either the wild type risk or mutant (R381Q) protective form of IL23R. After confirmation that the model system expressed the genes/proteins and had a differential effect on the phosphorylation of STAT3, we used RNAseq to explore differential gene regulation, in particular for genes implicated with risk to psoriasis, at a single time point for both cell types, and in a time course experiment for Jurkat CD4+ T cells. These experiments discovered differentially regulated genes in the cells expressing wild type and mutant IL23R, including HLA-B, SOCS1, RUNX3, CCR5, CXCR3, CCR9, KLF3, CD28, IRF, SOCS6, TNFAIP and ICAM5, that have been implicated in both the IL23 pathway and psoriasis. These genes have the potential to define a IL23/psoriasis pathway in disease, advancing our understanding of the biology behind the disease.

Tannic acid elicits differential gene regulation in prostate cancer apoptosis.

Prostate cancer is a significant global health concern that requires innovative therapeutic investigations. Here, the potential anticancer properties of tannic acid were evaluated by examining its effects on apoptosis in prostate cancer cell lines. PC-3 and LnCaP prostate adeno carcinoma cells, along with PNT1A prostate control cells, were cultured and divided into untreated and tannic acid-treated groups. Cell proliferation, cytotoxicity, and effects of tannic acid on the cell death mechanism were evaluated. mRNA expression levels of 84 genes were explored in cells following tannic acid treatment. Notably, tannic acid-induced down-regulation of several pro-survival genes, including ATM, BCL2, BCL2A1, BIK, BIRC2, BIRC3, BRE, CASP3, CASP6, CASP8, CHEK2, CRADD, PPIA, RPA3, TNFSF18, TRAF1, TRAF2, TRAF4, and TRAF5 in both cell lines. Moreover, tannic acid treatment led to the up-regulation of various pro-apoptotic genes, such as BCL10, BIRC3, BNIP3, CASP1, CASP5, CD40, CIDEB, DAPK2, FASLG, GADD45A, MYD88, RPA 3, TNFRSF10D, TNFRSF17, TNFRSF8, TNFSF13B, TNFSF4, TNFSF7, TNFSF8, TNFSF9, TP53, TRAF1, and TRAF2 in both PC-3 and LnCap cells. These findings highlight tannic acid's ability to induce apoptosis in prostate cancer cells through pro-apoptotic pathways. This study concludes that tannic acid selectively inhibits prostate cancer cell growth.

Molecular mechanisms of natural antifreeze phenomena and their application in cryopreservation.

Cryopreservation presents a critical challenge due to cryo-damage, such as crystallization and osmotic imbalances that compromise the integrity of biological tissues and cells. In contrast, various organisms in nature exhibit remarkable freezing tolerance, leveraging complex molecular mechanisms to survive extreme cold. This review explores the adaptive strategies of freeze-tolerant species, including the regulation of specific genes, proteins, and metabolic pathways, to enhance survival in low-temperature environments. We then discuss recent advancements in cryopreservation technologies that aim to mimic these natural phenomena to preserve cellular and tissue integrity. Special focus is given to the roles of glucose metabolism, microRNA expression, and cryoprotective protein modulation in improving cryopreservation outcomes. The insights gained from studying natural antifreeze mechanisms offer promising directions for advancing cryopreservation techniques, with potential applications in medical, agricultural, and conservation fields. Future research should aim to further elucidate these molecular mechanisms to develop more effective and reliable cryopreservation methods.

A Spatiotemporally Controlled Gene-Regulation Strategy for Combined Tumor Therapy Based on Upconversion Hybrid Nanosystem.

The lack of precise spatiotemporal gene modulation and therapy impedes progress in medical applications. Herein, a 980nm near-infrared (NIR) light-controlled nanoplatform, namely URMT, is developed, which can allow spatiotemporally controlled photodynamic therapy and trigger the enzyme-activated gene expression regulation in tumors. URMT is constructed by engineering an enzyme-activatable antisense oligonucleotide, which combined with an upconversion nanoparticle (UCNP)-based photodynamic nanosystem, followed by the surface functionalization of triphenylphosphine (TPP), a mitochondria-targeting ligand. URMT allows for the 980nm NIR light-activated generation of reactive oxygen species, which can induce the translocation of a DNA repair enzyme (namely apurinic/apyrimidinic endonuclease 1, APE1) from the nucleus to mitochondria. APE1 can recognize the basic apurinic/apyrimidinic (AP) sites in DNA double-strands and perform cleavage, thereby releasing the functional single-strands for gene regulation. Overall, an augmented antitumor effect is observed due to NIR light-controlled mitochondrial damage and enzyme-activated gene regulation. Altogether, the approach reported in this study offers high spatiotemporal precision and shows the potential to achieve precise and specific gene regulation for targeted tumor treatment.

Actin Polymerization Status Regulates Tendon Homeostasis through Myocardin-Related Transcription Factor-A.

The actin cytoskeleton is a potent regulator of tenocyte homeostasis. However, the mechanisms by which actin regulates tendon homeostasis are not entirely known. This study examined the regulation of tenocyte molecule expression by actin polymerization via the globular (G-) actin-binding transcription factor, myocardin-related transcription factor-a (MRTF). We determined that decreasing the proportion of G-actin in tenocytes by treatment with TGFβ1 increases nuclear MRTF. These alterations in actin polymerization and MRTF localization coincided with favorable alterations to tenocyte gene expression. In contrast, latrunculin A increases the proportion of G-actin in tenocytes and reduces nuclear MRTF, causing cells to acquire a tendinosis-like phenotype. To parse out the effects of F-actin depolymerization from regulation by MRTF, we treated tenocytes with cytochalasin D. Similar to latrunculin A treatment, exposure of cells to cytochalasin D increases the proportion of G-actin in tenocytes. However, unlike latrunculin A treatment, cytochalasin D increases nuclear MRTF. Compared to latrunculin A treatment, cytochalasin D led to opposing effects on the expression of a subset of genes. The differential regulation of genes by latrunculin A and cytochalasin D suggests that actin signals through MRTF to regulate a specific subset of genes. By targeting the deactivation of MRTF through the inhibitor CCG1423, we verify that MRTF regulates Type I Collagen, Tenascin C, Scleraxis, and α-smooth muscle actin in tenocytes. Actin polymerization status is a potent regulator of tenocyte homeostasis through the modulation of several downstream pathways, including MRTF. Understanding the regulation of tenocyte homeostasis by actin may lead to new therapeutic interventions against tendinopathies, such as tendinosis.

Ontogeny and organ-specific steroidal glycoside diversity is associated with differential expression of steroidal glycoside pathway genes in two Solanum dulcamara leaf chemotypes.

Solanaceous plants, such as Solanum dulcamara, produce steroidal glycosides (SGs). Leaf SG profiles vary among S. dulcamara individuals, leading to distinct phytochemical phenotypes ('chemotypes') and intraspecific phytochemical diversity ('chemodiversity'). However, if and how SG chemodiversity varies among organs and across ontogeny, and how this relates to SG metabolism gene expression is unknown. Among organs and across ontogeny, S. dulcamara plants with saturated (S) and unsaturated (U) SG leaf chemotypes were selected and clonally propagated. Roots, stems and leaves were harvested from vegetative and flowering plants. Extracts were analysed using untargeted LC-MS. Expression of candidate genes in SG metabolism (SdGAME9, SdGAME4, SdGAME25, SdS5αR2 and SdDPS) was analysed using RT-qPCRs. Our analyses showed that SG chemodiversity varies among organs and across ontogeny in S. dulcamara; SG richness (Dmg) was higher in flowering than vegetative plants. In vegetative plants, Dmg was higher for leaves than for roots. Lack of SdGAME25 expression in U-chemotype leaves, while readily expressed in roots and stems, suggests a pivotal role for SdGAME25 in differentiation of leaf chemotypes in vegetative and flowering plants. By acting as an ontogeny-dependent chemotypic switch, differential regulation of SdGAME25 enables adaptive allocation of SGs, thereby increasing SG chemodiversity in leaves. This indicates that differential expression and/or regulation of glycoalkaloid metabolism genes, rather than their presence or absence, explains observed chemotypic variation in SG chemodiversity among organs and across ontogeny.

LIKE HETEROCHROMATIN PROTEIN 1 (LHP1) partially inhibits the transcriptional activation of FT by MYB73 and regulates flowering in Arabidopsis.

Polycomb group (PcG) proteins are essential gene repressors in higher eukaryotes. However, how PcG proteins mediate transcriptional regulation of specific genes remains unknown. LIKE HETEROCHROMATIN PROTEIN 1 (LHP1), as a component of Polycomb Repression Complexes (PRC), epigenetically mediates several plant developmental processes together with PcG proteins. We observed physical interaction between MYB73 and LHP1 in vitro and in vivo. Genetic analysis indicated that myb73 mutants showed slightly late flowering, and the lhp1-3 myb73-2 double mutant exhibited delayed flowering and downregulated FT expression compared to lhp1-3. Chromatin immunoprecipitation and yeast one-hybrid assays revealed that MYB73 preferentially binds to the FT promoter. Additionally, our protoplast transient assays demonstrated that MYB73 activates to the FT promoter. Interestingly, the LHP1-MYB73 interaction is necessary to repress the FT promoter, suggesting that the LHP1-MYB73 interaction prevents FT activation by MYB73 in Arabidopsis. Our results show an example in which a chromatin regulator affects transcriptional regulation by negatively regulating a transcription factor through direct interaction.

Cross-species transcriptomics reveals differential regulation of essential photosynthesis genes in Hirschfeldia incana.

Photosynthesis is the only yield-related trait not yet substantially improved by plant breeding. Previously, we have established H. incana as the model plant for high photosynthetic light-use efficiency (LUE). Now we aim to unravel the genetic basis of this trait in H. incana, potentially contributing to the improvement of photosynthetic LUE in other species. Here, we compare its transcriptomic response to high light with that of Arabidopsis thaliana, Brassica rapa, and Brassica nigra, 3 fellow Brassicaceae members with lower photosynthetic LUE. We built a high-light, high-uniformity growing environment, in which the plants developed normally without signs of stress. We compared gene expression in contrasting light conditions across species, utilizing a panproteome to identify orthologous proteins. In-depth analysis of 3 key photosynthetic pathways showed a general trend of lower gene expression under high-light conditions for all 4 species. However, several photosynthesis-related genes in H. incana break this trend. We observed cases of constitutive higher expression (like antenna protein LHCB8), treatment-dependent differential expression (as for PSBE), and cumulative higher expression through simultaneous expression of multiple gene copies (like LHCA6). Thus, H. incana shows differential regulation of essential photosynthesis genes, with the light-harvesting complex as the first point of deviation. The effect of these expression differences on protein abundance and turnover, and ultimately the high photosynthetic LUE phenotype is relevant for further investigation. Furthermore, this transcriptomic resource of plants fully grown under, rather than briefly exposed to, a very high irradiance, will support the development of highly efficient photosynthesis in crops.