Clinical efficacy of tumor organoid-guided cancer therapy for locally advanced unresectable or metastatic breast cancer.

Patient-derived organoids (PDOs) may facilitate treatment selection. This retrospective cohort study evaluated the feasibility and clinical benefit of using PDOs to guide personalized treatment in metastatic breast cancer (MBC). Patients diagnosed with MBC were recruited between January 2019 and August 2022. PDOs were established and the efficacy of customized drug panels was determined by measuring cell mortality after drug exposure. Patients receiving organoid-guided treatment (OGT) were matched 1:2 by nearest neighbor propensity scores with patients receiving treatment of physician's choice (TPC). The primary outcome was progression-free survival. Secondary outcomes included objective response rate and disease control rate. Targeted gene sequencing and pathway enrichment analysis were performed. Forty-six PDOs (46 of 51, 90.2%) were generated from 45 MBC patients. PDO drug screening showed an accuracy of 78.4% (95% CI 64.9%-91.9%) in predicting clinical responses. Thirty-six OGT patients were matched to 69 TPC patients. OGT was associated with prolonged median progression-free survival (11.0 months vs. 5.0 months; hazard ratio 0.53 [95% CI 0.33-0.85]; p = .01) and improved disease control (88.9% vs. 63.8%; odd ratio 4.26 [1.44-18.62]) compared with TPC. The objective response rate of both groups was similar. Pathway enrichment analysis in hormone receptor-positive, human epidermal growth factor receptor 2-negative patients demonstrated differentially modulated pathways implicated in DNA repair and transcriptional regulation in those with reduced response to capecitabine/gemcitabine, and pathways associated with cell cycle regulation in those with reduced response to palbociclib. Our study shows that PDO-based functional precision medicine is a feasible and effective strategy for MBC treatment optimization and customization.

Arithmetic logic unit based on the metastructure with coherent absorption: erratum.

We present a corrigendum to our Letter [Opt. Lett.48, 5699 (2023)10.1364/OL.505761]. In section 3 of the original supplementary material, the absolute value is incorrectly taken in resolving the Kubo formula for describing the conductivity of graphene. Because this is a mistake with the conductivity of graphene, the coupling of the original structure is broken when the correct result is inserted into the code of the transfer matrix. So, the structure of the arithmetic logic unit (ALU), characteristic frequency point, and phase control have been modified accordingly. However, the ultimate function and the conclusion of this work remain unchanged.

Chiral anomaly and Weyl orbit in three-dimensional Dirac semimetal Cd3As2grown on Si.

Preparing Cd3As2, which is a three-dimensional (3D) Dirac semimetal in certain crystal orientation, on Si is highly desirable as such a sample may well be fully compatible with existing Si CMOS technology. However, there is a dearth of such a study regarding Cd3As2films grown on Si showing the chiral anomaly. Here,for the first time, we report the novel preparation and fabrication technique of a Cd3As2(112) film on a Si (111) substrate with a ZnTe (111) buffer layer which explicitly shows the chiral anomaly with a nontrivial Berry's phase ofπ. Despite the Hall carrier density (n3D≈9.42×1017cm-3) of our Cd3As2film, which is almost beyond the limit for the portents of a 3D Dirac semimetal to emerge, we observe large linear magnetoresistance in a perpendicular magnetic field and negative magnetoresistance in a parallel magnetic field. These results clearly demonstrate the chiral magnetic effect and 3D Dirac semimetallic behavior in our silicon-based Cd3As2film. Our tailoring growth of Cd3As2on a conventional substrate such as Si keeps the sample quality, while also achieving a low carrier concentration.

Mettl14-Mediated m6A Modification Is Essential for Germinal Center B Cell Response.

The germinal center (GC) response is essential for generating memory B and long-lived Ab-secreting plasma cells during the T cell-dependent immune response. In the GC, signals via the BCR and CD40 collaboratively promote the proliferation and positive selection of GC B cells expressing BCRs with high affinities for specific Ags. Although a complex gene transcriptional regulatory network is known to control the GC response, it remains elusive how the positive selection of GC B cells is modulated posttranscriptionally. In this study, we show that methyltransferase like 14 (Mettl14)-mediated methylation of adenosines at the position N 6 of mRNA (N 6-methyladenosine [m6A]) is essential for the GC B cell response in mice. Ablation of Mettl14 in B cells leads to compromised GC B cell proliferation and a defective Ab response. Interestingly, we unravel that Mettl14-mediated m6A regulates the expression of genes critical for positive selection and cell cycle regulation of GC B cells in a Ythdf2-dependent but Myc-independent manner. Furthermore, our study reveals that Mettl14-mediated m6A modification promotes mRNA decay of negative immune regulators, such as Lax1 and Tipe2, to upregulate genes requisite for GC B cell positive selection and proliferation. Thus, our findings suggest that Mettl14-mediated m6A modification plays an essential role in the GC B cell response.

A multiple cancer cell optical biosensing metastructure realized by CPA.

A one-dimensional optical biosensing metastructure (OBM) with graphene layers is presented in this paper. It is realized by coherent perfect absorption (CPA) and operates in the transverse electric mode. It shows a strong linear fitting relationship between the refractive index (RI) of the analysis layer and the frequency corresponding to the absorption peak, and the R-square is up to 1. Additionally, based on the principle of CPA, the OBM can realize the function of multiple cancer cell detection by adjusting the detection range by controlling the phase difference of coherent electromagnetic waves. Its detection ranges are 1.34-1.355 and 1.658-1.662. Thanks to its high-quality factor, great figure of merit, and low detection limit, whose best values are, respectively, 6.9 × 104, 1.2 × 104 RIU-1, and 3.6 × 10-6 RIU, the detection of weak changes in the RI of a cancer cell is possible. Additionally, its sensitivity can reach 26.57 THz RIU-1. This OBM based on CPA has major implications for advancing the study and investigation into the application of CPA. It also provides a simple and efficient approach to distinguish cancer cells and may be widely used in the biomedical field.

Cytosolic DNA initiates a vicious circle of aging-related endothelial inflammation and mitochondrial dysfunction via STING: the inhibitory effect of Cilostazol.

Endothelial senescence, aging-related inflammation, and mitochondrial dysfunction are prominent features of vascular aging and contribute to the development of aging-associated vascular disease. Accumulating evidence indicates that DNA damage occurs in aging vascular cells, especially in endothelial cells (ECs). However, the mechanism of EC senescence has not been completely elucidated, and so far, there is no specific drug in the clinic to treat EC senescence and vascular aging. Here we show that various aging stimuli induce nuclear DNA and mitochondrial damage in ECs, thus facilitating the release of cytoplasmic free DNA (cfDNA), which activates the DNA-sensing adapter protein STING. STING activation led to a senescence-associated secretory phenotype (SASP), thereby releasing pro-aging cytokines and cfDNA to further exacerbate mitochondrial damage and EC senescence, thus forming a vicious circle, all of which can be suppressed by STING knockdown or inhibition. Using next-generation RNA sequencing, we demonstrate that STING activation stimulates, whereas STING inhibition disrupts pathways associated with cell senescence and SASP. In vivo studies unravel that endothelial-specific Sting deficiency alleviates aging-related endothelial inflammation and mitochondrial dysfunction and prevents the development of atherosclerosis in mice. By screening FDA-approved vasoprotective drugs, we identified Cilostazol as a new STING inhibitor that attenuates aging-related endothelial inflammation both in vitro and in vivo. We demonstrated that Cilostazol significantly inhibited STING translocation from the ER to the Golgi apparatus during STING activation by targeting S162 and S243 residues of STING. These results disclose the deleterious effects of a cfDNA-STING-SASP-cfDNA vicious circle on EC senescence and atherogenesis and suggest that the STING pathway is a promising therapeutic target for vascular aging-related diseases. A proposed model illustrates the central role of STING in mediating a vicious circle of cfDNA-STING-SASP-cfDNA to aggravate age-related endothelial inflammation and mitochondrial damage.

Tunable and controllable multi-channel time-comb absorber based on continuous photonic time crystals.

To date there have been many studies on multi-channel absorbers for conventional photonic crystals (PCs). However, the number of absorption channels is small and uncontrollable, which cannot satisfy applications such as multispectral or quantitative narrowband selective filters. To address these issues, a tunable and controllable multi-channel time-comb absorber (TCA) based on continuous photonic time crystals (PTCs), is theoretically proposed. Compared with conventional PCs with fixed refractive index (RI), this system forms a stronger local electric field enhancement in the TCA by absorbing externally modulated energy, resulting in sharp multi-channel absorption peaks (APs). Tunability can be achieved by adjusting the RI, angle, and time period unit (T) of the PTCs. Diversified tunable methods allow the TCA to have more potential applications. In addition, changing T can adjust the number of multi-channels. More importantly, changing the primary term coefficient of n1(t) of PTC1 can control the number of time-comb absorption peaks (TCAPs) in multi-channels within a certain range, and the mathematical relationship between the coefficients and the number of multiple channels is summarized. This will have potential applications in the design of quantitative narrowband selective filters, thermal radiation detectors, optical detection instruments, etc.

Arithmetic logic unit based on the metastructure with coherent absorption.

An arithmetic logic unit (ALU) based on the metastructure (MS) with coherent absorption (CA) is proposed in this Letter. The ALU can perform AND and exclusive OR logical operations at the same frequency point. So it can be regarded as an optical half-adder. By controlling the chemical potential of graphene and the phase difference of coherent electromagnetic waves (EWs), two different binary numbers are input into the ALU. The dynamic absorption peaks, which are generated based on the CA, output the outcomes of the carry-digit bit and non-carry sum bit. This ALU can be used in the field of optical processing and encryption, such as processing hamming code. This given ALU based on the MS with CA has major implications for advancing the study and investigation into the application of CA. Furthermore, it also provides a new, to the best of our knowledge, idea for the study of MS with logical operation.

Advanced optical terahertz fingerprint sensor based on coherent perfect absorption.

An advanced optical terahertz (THz) fingerprint sensor based on coherent perfect absorption (CPA) is proposed. Based on a one-dimensional layered photonic structure, the sensor contains a cavity that is developed for THz fingerprint measurement. Utilizing the magneto-optical effect of magnetized InSb, CPA is excited in the structure of the sensor. Taking α-lactose as exemplar material, this numerical simulation is integrated with a Drude-Lorentz model. The transfer matrix method (TMM) is used to calculate the sensitivity (S), linear range (LR), quality (Q), the figure of merit (FOM*), and detection limit (DL) theoretically. Employing the amplitude modulation detection method, the qualitative and quantitative analysis of the α-lactose thickness of 0-0.5 µm could be realized. Because of the fragility of CPA, the S is 0.78255 µm-1, the value of average Q is up to 8019.2, the value of average FOM* is 13 234.4 (THz µm)-1, and the lower DL is 4.21 × 10-6. Moreover, the evolutions of ensemble-averaged absorption in the vicinity of the absorption peaks for different types of disorder effects are considered, which will be considered in the fabrication of sensors.

Risk factors of pancreatitis after endoscopic retrograde cholangiopancreatography in patients with biliary tract diseases.

BACKGROUND: To investigate the risk factors of pancreatitis after endoscopic retrograde cholangiopancreatography (ERCP) in patients with biliary tract diseases. METHODS: We retrospectively analyzed the clinical data of 480 patients who underwent ERCP for biliary tract diseases at the Affiliated Zhongshan Hospital of Dalian University from October 2011 to October 2016. The patients were divided into a study group (n = 75, with PEP) and a control group (n = 405, without PEP) based on whether they developed post-ERCP pancreatitis (PEP), and their clinical baseline data and intraoperative conditions were retrieved and compared. Then, factors associated with PEP were analyzed using logistic regression model, based on which a nomogram prediction model was constructed. The receiver operating characteristic (ROC) curve and calibration curve were used to evaluate the performance of the prediction model. RESULTS: Significant differences in age, sex, history of pancreatitis, history of choledocholithiasis, pancreatic duct imaging, pancreatic sphincterotomy, difficult cannulation, multiple cannulation attempts and juxtapapillary duodenal diverticula were observed between the two groups. Multivariate logistic regression analysis showed that age less than 60 years (OR, 0.477; 95% CI, 0.26-0.855), female sex (OR, 2.162; 95% CI, 1.220-3.831), history of pancreatitis (OR, 2.567; 95% CI, 1.218-5.410), history of choledocholithiasis (OR, 2.062; 95% CI, 1.162-3.658), pancreatic sphincterotomy (OR, 2.387; 95% CI, 1.298-4.390), pancreatic duct imaging (OR, 4.429; 95% CI, 1.481-13.242), multiple cannulation attempts (OR, 2.327; 95% CI, 1.205-4.493), difficult cannulation (OR, 2.421; 95% CI, 1.143-5.128), and JPD (OR, 2.002; 95% CI, 1.125-3.564) were independent risk factors for PEP. The nomogram for predicting the occurrence of PEP demonstrated an area under the ROC curve (AUC) of 0.787, and the calibration curves of the model showed good consistency between the predicted and actual probability of PEP. CONCLUSION: Our results showed that age less than 60 years, female sex, history of pancreatitis, history of choledocholithiasis, pancreatic sphincterotomy, pancreatic duct imaging, multiple cannulation attempts, difficult cannulation and juxtapapillary duodenal diverticula were independent risk factors for PEP. In addition, the established nomogram demonstrated promising clinical efficacy in predicting PEP risk in patients who underwent ERCP for biliary tract diseases.

Asymmetric Allylic Substitution-Isomerization for the Modular Synthesis of Axially Chiral N-Vinylquinazolinones.

Axially chiral N-substituted quinazolinones are important bioactive molecules, which are presented in many synthetic drugs. However, most strategies toward their atroposelective synthesis are mainly limited to the axially chiral arylquinazolinone frameworks. The development of modular synthetic methods to access diverse quinazolinone-based atropisomers remains scarce and challenging. Herein, we report the regio- and atroposelective synthesis of axially chiral N-vinylquinazolinones via the strategy of asymmetric allylic substitution-isomerization. The catalysis system utilized both asymmetric transition-metal catalysis and organocatalysis to efficiently afford trisubstituted and tetrasubstituted N-vinylquinazolinone atropisomers, respectively. With the meticulous design of ß-substituted allylic substrates, both Z- and E-tetrasubstituted axially chiral N-vinylquinazolinones were obtained in good yields and high enantioselectivities.

Visualizing an Electrochemically Induced Radical Cation of Bipyridine at Au(111)/Ionic Liquid Interfaces toward a Single-Molecule Switch.

Room-temperature ionic liquids (RTILs) emerged as ideal solvents, and bipyridine as one of the most used ligands have been widely employed in surface science, catalysis, and molecular electronics. Herein, in situ shell-isolated nanoparticle-enhanced Raman spectroscopy (SHINERS) and STM break junction (STM-BJ) technique has been employed to probe the electrochemical process of bipyridine at Au(111)/IL interfaces. It is interestingly found that these molecules undertake a redox process with a pair of well-defined reversible peaks in cyclic voltammograms (CVs). The spectroscopic evidence shows a radical cation generated with rising new Raman peaks related to parallel CC stretching of a positively charged pyridyl ring. Furthermore, these electrochemically charged bipyridine is also confirmed by electrochemical STM-BJ at the single-molecule level, which displays a binary conductance switch ratio of about 400% at the redox potentials. This present work offers a molecular-level insight into the pyridine-mediated reaction process and electron transport in RTILs.

The Role of Klotho Protein Against Sevoflurane-Induced Neuronal Injury.

The effects of general anesthetics on the developing brain have aroused much attention in recent years. Sevoflurane, a commonly used inhalation anesthetic especially in pediatric anesthesia, can induce developmental neurotoxicity. In this study, the differentially expressed mRNAs in the hippocampus of newborn rats exposed to 3% sevoflurane for 6 h were detected by RNA-Sequencing. Those data indicated that the mRNA of Klotho was increased after exposure to sevoflurane. Moreover, the protein expression of Klotho was assayed by Western Blot. Besides over-expression and under-expression of Klotho protein, we also detected changes of cell proliferation, ROS, JC-1, and Bcl-2/Bax ratio in PC12 cells exposed to sevoflurane. After exposure to 3% sevoflurane, the expression of Klotho protein increased in the hippocampus of neonatal rats. In PC12 cells, exposure to sevoflurane could increase cellular ROS level, reduce mitochondrial membrane potential and Bcl-2/Bax ratio. While overexpression of Klotho alleviated the above changes, knockdown of Klotho aggravated the injury of sevoflurane. Klotho protein could reduce oxidative stress and mitochondrial injury induced by sevoflurane in the neuron.

Fingerprint combining with quantitative analysis of multi-components by single marker for quality control of Chenxiang Huaqi tablets.

BACKGROUND: Chenxiang Huaqi tablets (CXHQTs) are a traditional Chinese medicine (TCM) commonly used to treat stomach-related diseases. Currently, the ministerial standards do not provide detailed guidance and regulations on the content determination of CXHQTs, and the reported studies only use individual active components as indicators for determining effective ingredients. OBJECTIVE: The present study aimed to propose a methodology for quality control of CXHQTs based on high-performance liquid chromatography (HPLC) fingerprinting combined with the quantitative analysis of multi-components by single marker (QAMS) method. METHODS: HPLC method was used to determine seven active ingredients and performed fingerprint analysis of CXHQTs. To further process chemometric assessment, technical analysis-model including similarity analysis (SA), hierarchical clustering analysis (HCA), principal components analysis (PCA) and orthogonal partial least squares discrimination analysis (OPLS-DA) was set up to differentiate and classify the 20 batches of samples. RESULTS: After comparing the results of QAMS method with the external standard method (ESM), we found there is no significant difference. Besides, the fingerprint of CXHQT was also established. CONCLUSION: HPLC fingerprint combined with the QAMS could be an efficient and selective analysis technique to achieve a qualitative and quantitative evaluation of executing quality processes.

Lysine Deprivation Regulates Npy Expression via GCN2 Signaling Pathway in Mandarin Fish (Siniperca chuatsi).

Regulation of food intake is associated with nutrient-sensing systems and the expression of appetite neuropeptides. Nutrient-sensing systems generate the capacity to sense nutrient availability to maintain energy and metabolism homeostasis. Appetite neuropeptides are prominent factors that are essential for regulating the appetite to adapt energy status. However, the link between the expression of appetite neuropeptides and nutrient-sensing systems remains debatable in carnivorous fish. Here, with intracerebroventricular (ICV) administration of six essential amino acids (lysine, methionine, tryptophan, arginine, phenylalanine, or threonine) performed in mandarin fish (Siniperca chuatsi), we found that lysine and methionine are the feeding-stimulating amino acids other than the reported valine, and found a key appetite neuropeptide, neuropeptide Y (NPY), mainly contributes to the regulatory role of the essential amino acids on food intake. With the brain cells of mandarin fish cultured in essential amino acid deleted medium (lysine, methionine, histidine, valine, or leucine), we showed that only lysine deprivation activated the general control nonderepressible 2 (GCN2) signaling pathway, elevated α subunit of eukaryotic translation initiation factor 2 (eIF2α) phosphorylation, increased activating transcription factor 4 (ATF4) protein expression, and finally induced transcription of npy. Furthermore, pharmacological inhibition of GCN2 and eIF2α phosphorylation signaling by GCN2iB or ISRIB, effectively blocked the transcriptional induction of npy in lysine deprivation. Overall, these findings could provide a better understanding of the GCN2 signaling pathway involved in food intake control by amino acids.

Subcutaneous phaeohyphomycosis caused by plant pathogenic Corynespora cassiicola: A case report.

Corynespora cassiicola is a common plant pathogen responsible for leaf-spotting diseases in the tropical and subtropical areas. C. cassiicola seldom causes human infections. Here we describe a case of subcutaneous phaeohyphomycosis caused by C. cassiicola in a 76-year-old Chinese man, who presented to our hospital with a purulent discharge and painful sensation on his right leg. Skin biopsy revealed an abscess, and culture confirmed C. cassiicola to be the causative agent. The result was further identified by sequence analysis of the internal transcribed spacer region. The patient was successfully treated with systemic voriconazole and wound debridement: the lesion disappeared after 20 days.

[Pharmacokinetic study of Polydopamine Guttate Pills loaded with active components of Sarcandrae Herba in rats].

An ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS) method was established for the determination of active components of Sarcandrae Herba, and applied to the pharmacokinetics study of multiple dosage forms. After SD rats were administered by gavage with three dosage forms [Sarcandrae Herba extract, commercial Sarcandrae Herba Guttate Pills, and polydopamine guttate pills loaded with active components of Sarcandrae Herba(PDA-Sg Guttate Pills)], blood samples were collected from the inner canthus at different time points. After protein precipitation, plasma samples were separated on ACQUITY UPLC C_(18) column(2.1 mm×100 mm, 1.7 µm). The mobile phase consisted of water containing 0.2% formic acid and acetonitrile in gradient elution. The negative ions were measured simultaneously in the multi-reaction monitoring(MRM) mode. The pharmacokinetic parameters were calculated and fitted by DAS 2.0. All four components could be detected in the plasma of rats in each group at each time point except the neochlorogenic acid and cryptochlorogenic acid in the Sarcandrae Herba extract group. The guttate pills group showed a significant increase in drug content at each time point. The exposure of the main components of Sarcandrae Herba in blood was effectively increased by PDA-drug loading effect in PDA-Sg Guttate Pills(The AUC_(0-24 h) of neochlorogenic acid, cryptochlorogenic acid, isaziridin and rosmarinic acid reached 2.45, 32.90, 1.54, 4.81 times that of the commercial guttate pills). This study proves the measurability of the above-mentioned multi-component in vitro-in vivo delivery process. The pharmacokinetic study has shown that PDA-Sg Guttate Pills can effectively delay the elimination time and improve the bioavailability of the four components, which can provide theoretical data for the production of the drug.

IκB kinase promotes Nrf2 ubiquitination and degradation by phosphorylating cylindromatosis, aggravating oxidative stress injury in obesity-related nephropathy.

BACKGROUND: Obesity-related nephropathy (ORN) has become one of the leading causes of end-stage renal disease and has tripled over the past decade. Previous studies have demonstrated that decreased reactive oxygen species production may contribute to improving ORN by ameliorating oxidative stress injury. Here, IκB kinase (IKK) was hypothesized to inactivate the deubiquitination activity of cylindromatosis (CYLD) by activating the phosphorylation of CYLD, thus promoting the ubiquitination of NF-E2-related factor 2 (Nrf2) and further aggravating oxidative stress injury of the kidney in ORN. This study was aimed to confirm this hypothesis. METHODS: Haematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Oil Red O staining were performed to assess histopathology. Dihydroethidium (DHE) staining and MDA, SOD, CAT, and GSH-PX assessments were performed to measure reactive oxygen species (ROS) production. Immunohistochemical (IHC) staining, qRT-PCR and/or western blotting were performed to assess the expression of related genes. JC-1 assays were used to measure the mitochondrial membrane potential (ΔΨm) of treated HK-2 cells. Co-immunoprecipitation experiments (Co-IP) were used to analyse the interaction between CYLD and Nrf2 in ORN. RESULTS: ORN in vivo and in vitro models were successfully constructed, and oxidative stress injury was detected in the model tissues and cells. Compared with the control groups, the phosphorylation level of CYLD increased while Nrf2 levels decreased in ORN model cells. An IKK inhibitor reduced lipid deposition, ROS production, CYLD phosphorylation levels and ΔΨm in vitro, which were reversed by knockdown of CYLD. Nrf2 directly bound to CYLD and was ubiquitinated in ORN cells. The proteasome inhibitor MG132 activated the Nrf2/ARE signalling pathway, thereby reversing the promoting effect of CYLD knockdown on oxidative stress. CONCLUSION: IKK inactivates the deubiquitination activity of CYLD by activating the phosphorylation of CYLD, thus promoting the ubiquitination of Nrf2 and further aggravating oxidative stress injury of the kidney in ORN. This observation provided a feasible basis for the treatment of kidney damage caused by ORN.

Natural hybridization among three Rhododendron species (Ericaceae) revealed by morphological and genomic evidence.

BACKGROUND: Natural hybridization can influence the adaptive response to selection and accelerate species diversification. Understanding the composition and structure of hybrid zones may elucidate patterns of hybridization processes that are important to the formation and maintenance of species, especially for taxa that have experienced rapidly adaptive radiation. Here, we used morphological traits, ddRAD-seq and plastid DNA sequence data to investigate the structure of a Rhododendron hybrid zone and uncover the hybridization patterns among three sympatric and closely related species. RESULTS: Our results show that the hybrid zone is complex, where bi-directional hybridization takes place among the three sympatric parental species: R. spinuliferum, R. scabrifolium, and R. spiciferum. Hybrids between R. spinuliferum and R. spiciferum (R. ×duclouxii) comprise multiple hybrid classes and a high proportion of F1 generation hybrids, while a novel hybrid taxon between R. spinuliferum and R. scabrifolium dominated the F2 generation, but no backcross individuals were detected. The hybrid zone showed basically coincident patterns of population structure between genomic and morphological data. CONCLUSIONS: Natural hybridization exists among the three Rhododendron species in the hybrid zone, although patterns of hybrid formation vary between hybrid taxa, which may result in different evolutionary outcomes. This study represents a unique opportunity to dissect the ecological and evolutionary mechanisms associated with adaptive radiation of Rhododendron species in a biodiversity hotspot.

Differential expressions of anthocyanin synthesis genes underlie flower color divergence in a sympatric Rhododendron sanguineum complex.

BACKGROUND: The Rhododendron sanguineum complex is endemic to alpine mountains of northwest Yunnan and southeast Tibet of China. Varieties in this complex exhibit distinct flower colors even at the bud stage. However, the underlying molecular regulations for the flower color variation have not been well characterized. Here, we investigated this via measuring flower reflectance profiles and comparative transcriptome analyses on three coexisting varieties of the R. sanguineum complex, with yellow flush pink, bright crimson, and deep blackish crimson flowers respectively. We compared the expression levels of differentially-expressed-genes (DEGs) of the anthocyanin / flavonoid biosynthesis pathway using RNA-seq and qRT-PCR data. We performed clustering analysis based on transcriptome-derived Single Nucleotide Polymorphisms (SNPs) data, and finally analyzed the promoter architecture of DEGs. RESULTS: Reflectance spectra of the three color morphs varied distinctively in the range between 400 and 700 nm, with distinct differences in saturation, brightness, hue, and saturation/hue ratio, an indirect measurement of anthocyanin content. We identified 15,164 orthogroups that were shared among the three varieties. The SNP clustering analysis indicated that the varieties were not monophyletic. A total of 40 paralogous genes encoding 12 enzymes contributed to the flower color polymorphism. These anthocyanin biosynthesis-related genes were associated with synthesis, modification and transportation properties (RsCHS, RsCHI, RsF3H, RsF3'H, RsFLS, RsANS, RsAT, RsOMT, RsGST), as well as genes involved in catabolism and degradation (RsBGLU, RsPER, RsCAD). Variations in sequence and cis-acting elements of these genes might correlate with the anthocyanin accumulation, thus may contribute to the divergence of flower color in the R. sanguineum complex. CONCLUSIONS: Our results suggested that the varieties are very closely related and flower color variations in the R. sanguineum complex correlate tightly with the differential expression levels of genes involved in the anabolic and catabolic synthesis network of anthocyanin. Our study provides a scenario involving intricate relationships between genetic mechanisms for floral coloration accompanied by gene flow among the varieties that may represent an early case of pollinator-mediated incipient sympatric speciation.