Antagonistic MADS-box transcription factors SEEDSTICK and SEPALLATA3 form a transcriptional regulatory network that regulates seed oil accumulation.

Transcriptional regulation is essential for balancing multiple metabolic pathways that influence oil accumulation in seeds. Thus far, the transcriptional regulatory mechanisms that govern seed oil accumulation remain largely unknown. Here, we identified the transcriptional regulatory network composed of MADS-box transcription factors SEEDSTICK (STK) and SEPALLATA3 (SEP3), which bridges several key genes to regulate oil accumulation in seeds. We found that STK, highly expressed in the developing embryo, positively regulates seed oil accumulation in Arabidopsis (Arabidopsis thaliana). Furthermore, we discovered that SEP3 physically interacts with STK in vivo and in vitro. Seed oil content is increased by the SEP3 mutation, while it is decreased by SEP3 overexpression. The chromatin immunoprecipitation, electrophoretic mobility shift assay, and transient dual-luciferase reporter assays showed that STK positively regulates seed oil accumulation by directly repressing the expression of MYB5, SEP3, and SEED FATTY ACID REDUCER 4 (SFAR4). Moreover, genetic and molecular analyses demonstrated that STK and SEP3 antagonistically regulate seed oil production and that SEP3 weakens the binding ability of STK to MYB5, SEP3, and SFAR4. Additionally, we demonstrated that TRANSPARENT TESTA 8 (TT8) and ACYL-ACYL CARRIER PROTEIN DESATURASE 3 (AAD3) are direct targets of MYB5 during seed oil accumulation in Arabidopsis. Together, our findings provide the transcriptional regulatory network antagonistically orchestrated by STK and SEP3, which fine tunes oil accumulation in seeds.

Role of SiPHR1 in the Response to Low Phosphate in Foxtail Millet via Comparative Transcriptomic and Co-Expression Network Analyses.

Enhancing the absorption and utilization of phosphorus by crops is an important aim for ensuring food security worldwide. However, the gene regulatory network underlying phosphorus use in foxtail millet remains unclear. In this study, the molecular mechanism underlying low-phosphorus (LP) responsiveness in foxtail millet was evaluated using a comparative transcriptome analysis. LP reduced the chlorophyll content in shoots, increased the anthocyanin content in roots, and up-regulated purple acid phosphatase and phytase activities as well as antioxidant systems (CAT, POD, and SOD). Finally, 13 differentially expressed genes related to LP response were identified and verified using transcriptomic data and qRT-PCR. Two gene co-expression network modules related to phosphorus responsiveness were positively correlated with POD, CAT, and PAPs. Of these, SiPHR1, functionally annotated as PHOSPHATE STARVATION RESPONSE 1, was identified as an MYB transcription factor related to phosphate responsiveness. SiPHR1 overexpression in Arabidopsis significantly modified the root architecture. LP stress caused cellular, physiological, and phenotypic changes in seedlings. SiPHR1 functioned as a positive regulator by activating downstream genes related to LP tolerance. These results improve our understanding of the molecular mechanism underlying responsiveness to LP stress, thereby laying a theoretical foundation for the genetic modification and breeding of new LP-tolerant foxtail millet varieties.

Genome-wide investigation of histone acetyltransferase gene family and its responses to biotic and abiotic stress in foxtail millet (Setaria italica [L.] P. Beauv).

BACKGROUND: Modification of histone acetylation is a ubiquitous and reversible process in eukaryotes and prokaryotes and plays crucial roles in the regulation of gene expression during plant development and stress responses. Histone acetylation is co-regulated by histone acetyltransferase (HAT) and histone deacetylase (HDAC). HAT plays an essential regulatory role in various growth and development processes by modifying the chromatin structure through interactions with other histone modifications and transcription factors in eukaryotic cells, affecting the transcription of genes. Comprehensive analyses of HAT genes have been performed in Arabidopsis thaliana and Oryza sativa. However, little information is available on the HAT genes in foxtail millet (Setaria italica [L.] P. Beauv). RESULTS: In this study, 24 HAT genes (SiHATs) were identified and divided into four groups with conserved gene structures via motif composition analysis. Phylogenetic analysis of the genes was performed to predict functional similarities between Arabidopsis thaliana, Oryza sativa, and foxtail millet; 19 and 2 orthologous gene pairs were individually identified. Moreover, all identified HAT gene pairs likely underwent purified selection based on their non-synonymous/synonymous nucleotide substitutions. Using published transcriptome data, we found that SiHAT genes were preferentially expressed in some tissues and organs. Stress responses were also examined, and data showed that SiHAT gene transcription was influenced by drought, salt, low nitrogen, and low phosphorus stress, and that the expression of four SiHATs was altered as a result of infection by Sclerospora graminicola. CONCLUSIONS: Results indicated that histone acetylation may play an important role in plant growth and development and stress adaptations. These findings suggest that SiHATs play specific roles in the response to abiotic stress and viral infection. This study lays a foundation for further analysis of the biological functions of SiHATs in foxtail millet.

Vasoactive agonists exert dynamic and coordinated effects on vascular smooth muscle cell elasticity, cytoskeletal remodelling and adhesion.

In this study, we examined the ability of vasoactive agonists to induce dynamic changes in vascular smooth muscle cell (VSMC) elasticity and adhesion, and tested the hypothesis that these events are coordinated with rapid remodelling of the cortical cytoskeleton. Real-time measurement of cell elasticity was performed with atomic force microscopy (AFM) and adhesion was assessed with AFM probes coated with fibronectin (FN). Temporal data were analysed using an Eigen-decomposition method. Elasticity in VSMCs displayed temporal oscillations with three components at approximately 0.001, 0.004 and 0.07 Hz, respectively. Similarly, adhesion displayed a similar oscillatory pattern. Angiotensin II (ANG II, 10(-6) M) increased (+100%) the amplitude of the oscillations, whereas the vasodilator adenosine (ADO, 10(-4) M) reduced oscillation amplitude (-30%). To test whether the oscillatory changes were related to the architectural alterations in cortical cytoskeleton, the topography of the submembranous actin cytoskeleton (100-300 nm depth) was acquired with AFM. These data were analysed to compare cortical actin fibre distribution and orientation before and after treatment with vasoactive agonists. The results showed that ANG II increased the density of stress fibres by 23%, while ADO decreased the density of the stress fibres by 45%. AFM data were supported by Western blot and confocal microscopy. Collectively, these observations indicate that VSMC cytoskeletal structure and adhesion to the extracellular matrix are dynamically altered in response to agonist stimulation. Thus, vasoactive agonists probably invoke unique mechanisms that dynamically alter the behaviour and structure of both the VSMC cytoskeleton and focal adhesions to efficiently support the normal contractile behaviour of VSMCs.

Prolonged vasoconstriction of resistance arteries involves vascular smooth muscle actin polymerization leading to inward remodelling.

AIMS: Inward remodelling of the resistance vasculature is predictive of hypertension and life-threatening cardiovascular events. We hypothesize that the contractile mechanisms responsible for maintaining a reduced diameter over time in response to prolonged stimulation with vasoconstrictor agonists are in part responsible for the initial stages of the remodelling process. Here we investigated the role of vascular smooth muscle (VSM) actin polymerization on agonist-induced vasoconstriction and development of inward remodelling. METHODS AND RESULTS: Experiments were conducted in Sprague-Dawley rat resistance vessels isolated from the cremaster and mesentery. Within blood vessels, actin dynamics of VSM were monitored by confocal microscopy after introduction of fluorescent actin monomers through electroporation and by differential centrifugation to probe globular (G) and filamentous (F) actin content. Results indicated that 4 h of agonist-dependent vasoconstriction induced inward remodelling and caused significant actin polymerization, elevating the F-/total-actin ratio. Inhibition of actin polymerization prevented vessels from maintaining prolonged vasoconstriction and developing inward remodelling. Activation of the small GTPases Rho/Rac/Cdc42 also increased the F-/total-actin ratio and induced inward remodelling, while inhibition of Rho kinase or Rac-1 prevented inward remodelling. Disruption of the actin cytoskeleton reversed the inward remodelling caused by prolonged vasoconstriction, but did not affect the passive diameter of freshly isolated vessels. CONCLUSION: These results indicate that vasoconstriction-induced inward remodelling is in part caused by the polymerization of actin within VSM cells through activation of small GTPases.

Mono-dispersed bioactive glass nanospheres: preparation and effects on biomechanics of mammalian cells.

Mono-dispersed SiO(2)-CaO bioactive glass nanospheres (BGNS) were prepared by a two step sol-gel method in the absence of surfactant. The size of BGNS ranged from 200 to 350 nm in diameter and exhibited a rough surface texture. In vitro biomineralization tests showed that BGNS could rapidly induce the deposition of an apatite layer in simulated body fluid (SBF). The effect of bioactive glass on the biomechanical properties of various mammalian cells was first reported in this paper. Atomic force microscopy (AFM) was used for measuring the biomechanical properties of mammalian cells. The result showed that BGNS-medium could significantly decrease the plasma membrane stiffness of bone marrow stem cells (BMSCs) by ∼50% and stimulate BMSCs spreading. The effect of BGNS on biomechanical properties of bovine aortic endothelial cells (BAECs) was opposite to that on BMSCs. BGNS increased the BAECs' stiffness and stimulated the elongation of endothelial cells and the formation of endothelial networks, which might potentially facilitate the vascularization of implanted BGNS-based biomaterials in tissue engineering as a scaffold or as an injectable system.