Control of inducible gene expression links cohesin to hematopoietic progenitor self-renewal and differentiation.

Cohesin is important for 3D genome organization. Nevertheless, even the complete removal of cohesin has surprisingly little impact on steady-state gene transcription and enhancer activity. Here we show that cohesin is required for the core transcriptional response of primary macrophages to microbial signals, and for inducible enhancer activity that underpins inflammatory gene expression. Consistent with a role for inflammatory signals in promoting myeloid differentiation of hematopoietic stem and progenitor cells (HPSCs), cohesin mutations in HSPCs led to reduced inflammatory gene expression and increased resistance to differentiation-inducing inflammatory stimuli. These findings uncover an unexpected dependence of inducible gene expression on cohesin, link cohesin with myeloid differentiation, and may help explain the prevalence of cohesin mutations in human acute myeloid leukemia.

CRISPR Inversion of CTCF Sites Alters Genome Topology and Enhancer/Promoter Function.

CTCF and the associated cohesin complex play a central role in insulator function and higher-order chromatin organization of mammalian genomes. Recent studies identified a correlation between the orientation of CTCF-binding sites (CBSs) and chromatin loops. To test the functional significance of this observation, we combined CRISPR/Cas9-based genomic-DNA-fragment editing with chromosome-conformation-capture experiments to show that the location and relative orientations of CBSs determine the specificity of long-range chromatin looping in mammalian genomes, using protocadherin (Pcdh) and ß-globin as model genes. Inversion of CBS elements within the Pcdh enhancer reconfigures the topology of chromatin loops between the distal enhancer and target promoters and alters gene-expression patterns. Thus, although enhancers can function in an orientation-independent manner in reporter assays, in the native chromosome context, the orientation of at least some enhancers carrying CBSs can determine both the architecture of topological chromatin domains and enhancer/promoter specificity. These findings reveal how 3D chromosome architecture can be encoded by linear genome sequences.

Chromatin jets define the properties of cohesin-driven in vivo loop extrusion.

Complex genomes show intricate organization in three-dimensional (3D) nuclear space. Current models posit that cohesin extrudes loops to form self-interacting domains delimited by the DNA binding protein CTCF. Here, we describe and quantitatively characterize cohesin-propelled, jet-like chromatin contacts as landmarks of loop extrusion in quiescent mammalian lymphocytes. Experimental observations and polymer simulations indicate that narrow origins of loop extrusion favor jet formation. Unless constrained by CTCF, jets propagate symmetrically for 1-2 Mb, providing an estimate for the range of in vivo loop extrusion. Asymmetric CTCF binding deflects the angle of jet propagation as experimental evidence that cohesin-mediated loop extrusion can switch from bi- to unidirectional and is controlled independently in both directions. These data offer new insights into the physiological behavior of in vivo cohesin-mediated loop extrusion and further our understanding of the principles that underlie genome organization.

Forces driving transposable element load variation during Arabidopsis range expansion.

Genetic load refers to the accumulated and potentially life-threatening deleterious mutations in populations. Understanding the mechanisms underlying genetic load variation of transposable element (TE) insertion, a major large-effect mutation, during range expansion is an intriguing question in biology. Here, we used 1,115 global natural accessions of Arabidopsis (Arabidopsis thaliana) to study the driving forces of TE load variation during its range expansion. TE load increased with range expansion, especially in the recently established Yangtze River basin population. Effective population size, which explains 62.0% of the variance in TE load, high transposition rate, and selective sweeps contributed to TE accumulation in the expanded populations. We genetically mapped and identified multiple candidate causal genes and TEs, and revealed the genetic architecture of TE load variation. Overall, this study reveals the variation in TE genetic load during Arabidopsis expansion and highlights the causes of TE load variation from the perspectives of both population genetics and quantitative genetics.

Arabidopsis EXECUTER1 interacts with WRKY transcription factors to mediate plastid-to-nucleus singlet oxygen signaling.

Chloroplasts produce singlet oxygen (1O2), which causes changes in nuclear gene expression through plastid-to-nucleus retrograde signaling to increase plant fitness. However, the identity of this 1O2-triggered pathway remains unclear. Here, we identify mutations in GENOMES UNCOUPLED4 (GUN4) and GUN5 as suppressors of phytochrome-interacting factor1 (pif1) pif3 in regulating the photo-oxidative response in Arabidopsis thaliana. GUN4 and GUN5 specifically interact with EXECUTER1 (EX1) and EX2 in plastids, and this interaction is alleviated by treatment with Rose Bengal (RB) or white light. Impaired expression of GUN4, GUN5, EX1, or EX2 leads to insensitivity to excess light and overexpression of EX1 triggers photo-oxidative responses. Strikingly, upon light irradiation or RB treatment, EX1 transiently accumulates in the nucleus and the nuclear fraction of EX1 shows a similar molecular weight as the plastid-located protein. Point mutagenesis analysis indicated that nuclear localization of EX1 is required for its function. EX1 acts as a transcriptional co-activator and interacts with the transcription factors WRKY18 and WRKY40 to promote the expression of 1O2-responsive genes. This study suggests that EX1 may act in plastid-to-nucleus signaling and establishes a 1O2-triggered retrograde signaling pathway that allows plants adapt to changing light environments during chloroplast development.

Signatures of Adaptation and Purifying Selection in Highland Populations of Dasiphora fruticosa.

High mountains harbor a considerable proportion of biodiversity, but we know little about how diverse plants adapt to the harsh environment. Here we finished a high-quality genome assembly for Dasiphora fruticosa, an ecologically important plant distributed in the Qinghai-Tibetan Plateau and lowland of the Northern Hemisphere, and resequenced 592 natural individuals to address how this horticulture plant adapts to highland. Demographic analysis revealed D. fruticosa underwent a bottleneck after Naynayxungla Glaciation. Selective sweep analysis of two pairs of lowland and highland populations identified 63 shared genes related to cell wall organization or biogenesis, cellular component organization, and dwarfism, suggesting parallel adaptation to highland habitats. Most importantly, we found that stronger purging of estimated genetic load due to inbreeding in highland populations apparently contributed to their adaptation to the highest mountain. Our results revealed how plants could tolerate the extreme plateau, which could provide potential insights for species conservation and crop breeding.

Multifaceted interactions between host ESCRT-III and budded virus-related proteins involved in entry and egress of the baculovirus Autographa californica multiple nucleopolyhedrovirus.

The endosomal sorting complex required for transport (ESCRT) is a conserved protein machine mediating membrane remodeling and scission. In the context of viral infection, different components of the ESCRT-III complex, which serve as the core machinery to catalyze membrane fission, are involved in diverse viruses' entry, replication, and/or budding. However, the interplay between ESCRT-III and viral factors in the virus life cycle, especially for that of large enveloped DNA viruses, is largely unknown. Recently, the ESCRT-III components Vps2B, Vps20, Vps24, Snf7, Vps46, and Vps60 were determined for entry and/or egress of the baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV). Here, we identified the final three ESCRT-III components Chm7, Ist1, and Vps2A of Spodoptera frugiperda. Overexpression of the dominant-negative forms of these proteins or RNAi downregulation of their transcripts significantly reduced infectious budded viruses (BVs) production of AcMNPV. Quantitative PCR together with confocal and transmission electron microscopy analysis revealed that these proteins were required for internalization and trafficking of BV during entry and egress of nucleocapsids. In infected Sf9 cells, nine ESCRT-III components were distributed on the nuclear envelope and plasma membrane, and except for Chm7, the other components were also localized to the intranuclear ring zone. Y2H and BiFC analysis revealed that 42 out of 64 BV-related proteins including 35 BV structural proteins and 7 non-BV structural proteins interacted with single or multiple ESCRT-III components. By further mapping the interactome of 64 BV-related proteins, we established the interaction networks of ESCRT-III and the viral protein complexes involved in BV entry and egress.IMPORTANCEFrom archaea to eukaryotes, the endosomal sorting complex required for transport (ESCRT)-III complex is hijacked by many enveloped and nonenveloped DNA or RNA viruses for efficient replication. However, the mechanism of ESCRT-III recruitment, especially for that of large enveloped DNA viruses, remains elusive. Recently, we found the ESCRT-III components Vps2B, Vps20, Vps24, Snf7, Vps46, and Vps60 are necessary for the entry and/or egress of budded viruses (BVs) of Autographa californica multiple nucleopolyhedrovirus. Here, we demonstrated that the other three ESCRT-III components Chm7, Ist1, and Vps2A play similar roles in BV infection. By determining the subcellular localization of ESCRT-III components in infected cells and mapping the interaction of nine ESCRT-III components and 64 BV-related proteins, we built the interaction networks of ESCRT-III and the viral protein complexes involved in BV entry and egress. These studies provide a fundamental basis for understanding the mechanism of the ESCRT-mediated membrane remodeling for replication of baculoviruses.

Exosomal transfer of activated neutrophil-derived lncRNA CRNDE promotes proliferation and migration of airway smooth muscle cells in asthma.

Activated neutrophil-derived exosomes reportedly contribute to the proliferation of airway smooth muscle cells (ASMCs), thereby aggravating the airway wall remodeling during asthma; however, the specific mechanism remains unclear. Lipopolysaccharide (LPS)-EXO and si-CRNDE-EXO were extracted from the media of human neutrophils treated with LPS and LPS + si-CRNDE (a siRNA targets long non-coding RNA CRNDE), respectively. Human ASMCs were co-cultured with LPS-EXO or si-CRNDE-EXO, and cell viability, proliferation and migration were measured. The interplay of colorectal neoplasia differentially expressed (CRNDE), inhibitor of nuclear factor kappa B kinase subunit beta (IKKß) and nuclear receptor subfamily 2 group C member 2 (TAK1) was explored using RNA immunoprecipitation (RIP) and Co-IP assays. A mouse model of asthma was induced using ovalbumin. CRNDE was upregulated in LPS-EXO and successfully transferred from LPS-treated neutrophils to ASMCs through exosome. Mechanically, CRNDE loaded in LPS-EXO reinforced TAK1-mediated IKKß phosphorylation, thereby activating the nuclear factor kappa B (NF-κB) pathway. Functionally, silencing CRNDE in LPS-EXO, an IKKß inhibitor, and an NF-κB inhibitor all removed the upregulation of cell viability, proliferation and migration induced by LPS-EXO in ASMCs. In the end, the in vivo experiment demonstrated that CRNDE knockdown in neutrophils effectively reduced the thickness of bronchial smooth muscle in a mouse model for asthma. Activated neutrophils-derived CRNDE was transferred to ASMCs through exosomes and activated the NF-κB pathway by enhancing IKKß phosphorylation. The latter promoted the proliferation and migration of ASMCs and then contributed to airway remodeling in asthma.

From leaf to multiscale models of photosynthesis: applications and challenges for crop improvement.

To keep up with the growth of human population and to circumvent deleterious effects of global climate change, it is essential to enhance crop yield to achieve higher production. Here we review mathematical models of oxygenic photosynthesis that are extensively used, and discuss in depth a subset that accounts for diverse approaches providing solutions to our objective. These include models (1) to study different ways to enhance photosynthesis, such as fine-tuning antenna size, photoprotection and electron transport; (2) to bioengineer carbon metabolism; and (3) to evaluate the interactions between the process of photosynthesis and the seasonal crop dynamics, or those that have included statistical whole-genome prediction methods to quantify the impact of photosynthesis traits on the improvement of crop yield. We conclude by emphasizing that the results obtained in these studies clearly demonstrate that mathematical modelling is a key tool to examine different approaches to improve photosynthesis for better productivity, while effective multiscale crop models, especially those that also include remote sensing data, are indispensable to verify different strategies to obtain maximized crop yields.

Left Hepatectomy Combined with Right Hepatic Artery Resection and Reconstruction for Hilar Cholangiocarcinoma.

BACKGROUND: Hepatectomy combined with hepatic artery reconstruction in the operation for hilar cholangiocarcinoma (Klatskin tumor) is a challenging procedure. We present a video of left hepatectomy combined with right hepatic artery reconstruction for hilar cholangiocarcinoma. PATIENT AND METHODS: The patient was a 60-year-old male who presented with obstructive jaundice. The imaging examination showed that the confluence of left and right hepatic ducts and the wall of common hepatic duct were thickened, the local lumen was narrowed, the intrahepatic bile duct was dilated, and the right hepatic artery was invaded by tumors nearly 2.3 centimeters. Left hepatectomy with total caudate lobectomy, resection with reconstruction of right hepatic artery, hilar lymphadenectomy, and Roux-en-Y hepaticojejunostomy were performed. RESULTS: The operation time was 345 min, and the amount of bleeding was about 400 ml. There was no blood transfusion. The pathology showed poorly differentiated adenocarcinoma, with negative margins of common bile duct and right hepatic duct, and negative results of all lymph nodes. The patient's recovery was uneventful and he was discharged on postoperative day 14. The patient was disease free at 12-month follow-up evaluation. CONCLUSIONS: Hepatic artery resection and reconstruction procedure is safe and feasible for hilar cholangiocarcinoma in a highly tertiary hepatobiliary center.

RING induces cell cycle arrest and apoptosis in human breast cancer cells by regulating the HSF1/MT2A axis.

It was reported that lowly expressed RING1 indicates poor prognosis in breast cancer (BC) patients, while the mechanism by which RING1 is involved in BC progression is not fully understood. Here, we found that RING1 was lowly expressed in BC tissues and cells than in normal mammary tissues and epithelial cells. Overexpression of RING1 suppressed the cell proliferative and colony formation abilities, and facilitated cell cycle arrest and cell apoptosis in BC cells (T47D and MCF-7 cells). Mechanistically, as an ubiquitin ligase, RING1 bound to HSF1 and induced its proteasome-dependent degradation. HSF1 could bind to the promoter region of MT2A to promote the transcriptional level of MT2A. While RING1 overexpression hindered the transcriptional activation of MT2A induced by HSF1. Moreover, ectopic expression of MT2A reversed the inhibitory effect of RING1 on cell proliferation and clonogenesis, and antagonized the promotion effect of RING1 on cell cycle arrest and apoptosis in BC cells. Additionally, T47D cells infected with or without lentivirus-mediated RING1 overexpression vector (LV-RING1) were injected subcutaneously into the right back of nude mice to evaluate tumorigenicity. And overexpression of RING1 impeded the growth of BC xenografts in mice. In conclusion, RING1 suppressed the transcriptional activation of MT2A induced by HSF1 by facilitating the ubiquitination degradation of HSF1, resulting in cell cycle arrest and apoptosis in BC cells.

HBB contributes to individualized aconitine-induced cardiotoxicity in mice via interfering with ABHD5/AMPK/HDAC4 axis.

The root of Aconitum carmichaelii Debx. (Fuzi) is an herbal medicine used in China that exerts significant efficacy in rescuing patients from severe diseases. A key toxic compound in Fuzi, aconitine (AC), could trigger unpredictable cardiotoxicities with high-individualization, thus hinders safe application of Fuzi. In this study we investigated the individual differences of AC-induced cardiotoxicities, the biomarkers and underlying mechanisms. Diversity Outbred (DO) mice were used as a genetically heterogeneous model for mimicking individualization clinically. The mice were orally administered AC (0.3, 0.6, 0.9 mg· kg-1 ·d-1) for 7 d. We found that AC-triggered cardiotoxicities in DO mice shared similar characteristics to those observed in clinic patients. Most importantly, significant individual differences were found in DO mice (variation coefficients: 34.08%-53.17%). RNA-sequencing in AC-tolerant and AC-sensitive mice revealed that hemoglobin subunit beta (HBB), a toxic-responsive protein in blood with 89% homology to human, was specifically enriched in AC-sensitive mice. Moreover, we found that HBB overexpression could significantly exacerbate AC-induced cardiotoxicity while HBB knockdown markedly attenuated cell death of cardiomyocytes. We revealed that AC could trigger hemolysis, and specifically bind to HBB in cell-free hemoglobin (cf-Hb), which could excessively promote NO scavenge and decrease cardioprotective S-nitrosylation. Meanwhile, AC bound to HBB enhanced the binding of HBB to ABHD5 and AMPK, which correspondingly decreased HDAC-NT generation and led to cardiomyocytes death. This study not only demonstrates HBB achievement a novel target of AC in blood, but provides the first clue for HBB as a novel biomarker in determining the individual differences of Fuzi-triggered cardiotoxicity.

Fabrication of levofloxacin-loaded porcine acellular dermal matrix hydrogel and functional assessment in urinary tract infection.

Bacterial cystitis, a commonly occurring urinary tract infection (UTI), is renowned for its extensive prevalence and tendency to recur. Despite the extensive utilization of levofloxacin as a conventional therapeutic approach for bacterial cystitis, its effectiveness is impeded by adverse toxic effects, drug resistance concerns, and its influence on the gut microbiota. This study introduces Lev@PADM, a hydrogel with antibacterial properties that demonstrates efficacy in the treatment of bacterial cystitis. Lev@PADM is produced by combining levofloxacin with decellularized porcine acellular dermal matrix hydrogel and exhibits remarkable biocompatibility. Lev@PADM demonstrates excellent stability as a hydrogel at body temperature, enabling direct administration to the site of infection through intravesical injection. This localized delivery route circumvents the systemic circulation of levofloxacin, resulting in a swift and substantial elevation of the antimicrobial agent's concentration specifically at the site of infection. The in vivo experimental findings provide evidence that Lev@PADM effectively prolongs the duration of levofloxacin's action, impedes the retention and invasion of E.coli in the urinary tract, diminishes the infiltration of innate immune cells into infected tissues, and simultaneously preserves the composition of the intestinal microbiota. These results indicate that, in comparison to the exclusive administration of levofloxacin, Lev@PADM offers notable benefits in terms of preserving the integrity of the bladder epithelial barrier and suppressing the recurrence of urinary tract infections.

Deficiency of ribosomal proteins reshapes the transcriptional and translational landscape in human cells.

Human ribosomes have long been thought to be uniform factories with little regulatory function. Accumulating evidence emphasizes the heterogeneity of ribosomal protein (RP) expression in specific cellular functions and development. However, a systematic understanding of functional relevance of RPs is lacking. Here, we surveyed translational and transcriptional changes after individual knockdown of 75 RPs, 44 from the large subunit (60S) and 31 from the small subunit (40S), by Ribo-seq and RNA-seq analyses. Deficiency of individual RPs altered specific subsets of genes transcriptionally and translationally. RP genes were under cotranslational regulation upon ribosomal stress, and deficiency of the 60S RPs and the 40S RPs had opposite effects. RP deficiency altered the expression of genes related to eight major functional classes, including the cell cycle, cellular metabolism, signal transduction and development. 60S RP deficiency led to greater inhibitory effects on cell growth than did 40S RP deficiency, through P53 signaling. Particularly, we showed that eS8/RPS8 deficiency stimulated apoptosis while eL13/RPL13 or eL18/RPL18 deficiency promoted senescence. We also validated the phenotypic impacts of uL5/RPL11 and eL15/RPL15 deficiency on retina development and angiogenesis, respectively. Overall, our study provides a valuable resource for and novel insights into ribosome regulation in cellular activities, development and diseases.

Ribosomes are the main effector of the translational machinery to synthesize proteins. In this study, the authors characterized genome-wide transcriptional and translational changes after knocking-down 75 individual human ribosomal proteins (RPs). They revealed that deficiency of individual RPs perturbed expression of specific subsets of genes, enriched in eight major functional classes, such as cell cycle and development. RPs were subjected to co-translational regulation under ribosomal stress where deficiency of the 60S RPs and the 40S RPs had opposite effects on the two subunits. They also showed that RPS8 deficiency stimulated cellular apoptosis while RPL13 and RPL18 deficiency promoted cellular senescence. They further showed functional and regulatory roles of RPL11 and RPL15 in retina development and angiogenesis, respectively.

Polygenic adaptation of rosette growth in Arabidopsis thaliana.

The rate at which plants grow is a major functional trait in plant ecology. However, little is known about its evolution in natural populations. Here, we investigate evolutionary and environmental factors shaping variation in the growth rate of Arabidopsis thaliana. We used plant diameter as a proxy to monitor plant growth over time in environments that mimicked latitudinal differences in the intensity of natural light radiation, across a set of 278 genotypes sampled within four broad regions, including an outgroup set of genotypes from China. A field experiment conducted under natural conditions confirmed the ecological relevance of the observed variation. All genotypes markedly expanded their rosette diameter when the light supply was decreased, demonstrating that environmental plasticity is a predominant source of variation to adapt plant size to prevailing light conditions. Yet, we detected significant levels of genetic variation both in growth rate and growth plasticity. Genome-wide association studies revealed that only 2 single nucleotide polymorphisms associate with genetic variation for growth above Bonferroni confidence levels. However, marginally associated variants were significantly enriched among genes with an annotated role in growth and stress reactions. Polygenic scores computed from marginally associated variants confirmed the polygenic basis of growth variation. For both light regimes, phenotypic divergence between the most distantly related population (China) and the various regions in Europe is smaller than the variation observed within Europe, indicating that the evolution of growth rate is likely to be constrained by stabilizing selection. We observed that Spanish genotypes, however, reach a significantly larger size than Northern European genotypes. Tests of adaptive divergence and analysis of the individual burden of deleterious mutations reveal that adaptive processes have played a more important role in shaping regional differences in rosette growth than maladaptive evolution.

Synthesis of 2,3-Fused Quinazolinones via the Radical Cascade Pathway and Reaction Mechanistic Studies by DFT Calculations.

An efficient radical cascade cyclization of unactivated alkenes toward the synthesis of a series of ring-fused quinazolinones has been developed in moderate to excellent yields using commercially available ethers, alkanes, and alcohols, respectively, under a base-free condition in a short time without a transition metal as catalyst. Notably, the transformations can be carried out with the advantages of a broad substrate scope and high atomic economy. Density functional theory calculations and wavefunction analyses were performed to elucidate the radical reaction mechanism.

Crypt-like patterned electrospun nanofibrous membrane and probiotics promote intestinal epithelium models close to tissues.

In vitro intestinal epithelium models have drawn great attention to investigating intestinal biology in recent years. However, the difficulty to maintain the normal physiological status of primary intestinal epithelium in vitro limits the applications. Here, we designed patterned electrospun polylactic acid (PLA) nanofibrous membranes with crypt-like topography and mimic ECM fibrous network to support crypt culture and construct in vitro intestinal epithelium models. The patterned electrospun PLA nanofibrous membranes modified with Matrigels at 0 °C showed high biocompatibility and promoted cell growth and proliferation. The constructed duodenum epithelium models and colon epithelium models on the patterned electrospun PLA nanofibrous membranes expressed the typical differentiation markers of intestinal epithelia and the gene expression levels were close to the original tissues, especially with the help of probiotics. The constructed intestinal epithelium models could be used to assess probiotic adhesion and colonization, which were verified to show significant differences with the Caco-2 cell models due to the different cell types. These findings provide new insights and a better understanding of the roles of biophysical, biochemical, and biological signals in the construction of in vitro intestinal epithelium models as well as the potential applications of these models in the study of host-gut microbes interactions. KEY POINTS: • Patterned electrospun scaffold has crypt-like topography and ECM nanofibrous network. • Matrigels at 0°C modify scaffolds more effectively than at 37°C. • Synergy of biomimic scaffold and probiotics makes in vitro model close to tissue.

Nacre-mimetic cerium-doped nano-hydroxyapatite/chitosan layered composite scaffolds regulate bone regeneration via OPG/RANKL signaling pathway.

Autogenous bone grafting has long been considered the gold standard for treating critical bone defects. However, its use is plagued by numerous drawbacks, such as limited supply, donor site morbidity, and restricted use for giant-sized defects. For this reason, there is an increasing need for effective bone substitutes to treat these defects. Mollusk nacre is a natural structure with outstanding mechanical property due to its notable "brick-and-mortar" architecture. Inspired by the nacre architecture, our team designed and fabricated a nacre-mimetic cerium-doped layered nano-hydroxyapatite/chitosan layered composite scaffold (CeHA/CS). Hydroxyapatite can provide a certain strength to the material like a brick. And as a polymer material, chitosan can slow down the force when the material is impacted, like an adhesive. As seen in natural nacre, the combination of these inorganic and organic components results in remarkable tensile strength and fracture toughness. Cerium ions have been demonstrated exceptional anti-osteoclastogenesis capabilities. Our scaffold featured a distinct layered HA/CS composite structure with intervals ranging from 50 to 200 µm, which provided a conducive environment for human bone marrow mesenchymal stem cell (hBMSC) adhesion and proliferation, allowing for in situ growth of newly formed bone tissue. In vitro, Western-blot and qPCR analyses showed that the CeHA/CS layered composite scaffolds significantly promoted the osteogenic process by upregulating the expressions of osteogenic-related genes such as RUNX2, OCN, and COL1, while inhibiting osteoclast differentiation, as indicated by reduced TRAP-positive osteoclasts and decreased bone resorption. In vivo, calvarial defects in rats demonstrated that the layered CeHA/CS scaffolds significantly accelerated bone regeneration at the defect site, and immunofluorescence indicated a lowered RANKL/OPG ratio. Overall, our results demonstrate that CeHA/CS scaffolds offer a promising platform for bone regeneration in critical defect management, as they promote osteogenesis and inhibit osteoclast activation.

Transcriptional repression specifies the central cell for double fertilization.

Double fertilization is a key innovation for the evolutionary success of angiosperms by which the two fertilized female gametes, the egg cell and central cell, generate the embryo and endosperm, respectively. The female gametophyte (embryo sac) enclosed in the sporophyte is derived from a one-celled haploid cell lineage. It undergoes successive events of mitotic divisions, cellularization, and cell specification to give rise to the mature embryo sac, which contains the two female gametes accompanied by two types of accessory cells, namely synergids and antipodals. How the cell fate of the central cell is specified has long been equivocal and is further complicated by the structural diversity of female gametophyte across plant taxa. Here, MADS-box protein AGL80 was verified as a transcriptional repressor that directly suppresses the expression of accessory cell-specific genes to specify the central cell. Further genetic rescue and phylogenetic assay of the AGL80 orthologs revealed a possible conserved mechanism in the Brassicaceae family. Results from this study provide insight into the molecular determination of the second female gamete cell in Brassicaceae.

Cytotoxic Compounds from Belamcanda chinensis (L.) DC Induced Apoptosis in Triple-Negative Breast Cancer Cells.

Four compounds (1, 5, 7, and 8) were first isolated from the genus Belamcanda Adans. nom. conserv., and six known compounds (2-4, 6, 9, and 10) were isolated from the rhizome of Belamcanda chinensis (L.) DC. Their structures were confirmed by spectroscopic data. Herein, compounds 1-10 were rhapontigenin, trans-resveratrol, 5,7,4'-trihydroxy-6,3',5'-trimethoxy-isoflavone, irisflorentin, 6-hydroxybiochannin A, iridin S, pinoresinol, 31-norsysloartanol, isoiridogermanal, and iristectorene B, respectively. All compounds were evaluated for their antiproliferative effects against five tumor cell lines (BT549, 4T1, MCF7, MDA-MB-231, and MDA-MB-468). Among them, compound 9 (an iridal-type triterpenoid) showed the highest activity against 4T1 and MDA-MB-468 cells. Further studies displayed that compound 9 inhibited cell metastasis, induced cells cycle arrest in the G1 phase, exhibited significant mitochondrial damage in 4T1 and MDA-MB-468 cells including excess reactive oxygen species, decreased mitochondrial membrane potential, and induced 4T1 and MDA-MB-468 cell apoptosis for the first time. In summary, these findings demonstrate that compound 9 exerts promising potential for triple-negative breast cancer treatment and deserves further evaluation.