Positional Cloning, Gene Expression, and Molecular Evolution of the Gene Conferring Purple Leaf Sheath in Rice

花青素可累積於水稻許多組織中，形成紫紅色或紫色外表性狀，以抵抗生物逆境和非生物逆境。亦不管是野生稻或是栽培稻皆可發現含花青素之紫色葉鞘，或不含花青素之綠色葉鞘其中稉稻台農72號之葉鞘為紫紅色，而秈稻台中秈17號之葉鞘為綠色，因此想瞭解影響水稻葉鞘顏色的基因為何。本研究以台農72號和台中秈17號之F2子代，利用定位選殖策略，找出在此分離族群調控水稻葉鞘顏色基因為OsC1，另在3個綠色葉鞘品種在OsC1調控區域R2R3 Myb序列有非同質性的SNP或序列產生缺失，序列差異造成水稻葉鞘顏色的改變。因而利用36個不同栽培品種、地方種與野生種探討OsC1基因之分子演化過程，藉由序列分析可歸類14種不同的單體型，並以中性理論Tajama’s D、 Fu and Li’s D 和 Fu and Li’s F檢測此基因，顯示OsC1可能受到天擇或選拔的影響。 為了證實OsC1基因影響著水稻葉鞘花青素之累積，挑選葉鞘顏色深淺不一的台農67號、台農72號、上季早稻和崑山五香稉等四個品種，測量花青素含量並以Real time PCR偵測OsC1和花青素相關基因OsB2、OsDfr和OsAns之表現量，結果呈現僅只有OsC1基因表現量與花青素含量有正相關，OsC1在深紫色葉鞘品種中葉鞘有較高的表現，並且隨著生育日數的增加營養生長時期基因之表現量也有顯著的提升，其他三個基因之表現與花青素含量則無相關性。由TNG72啟動子研究探討OsC1基因之表現位置，發現在三葉齡時期葉鞘無基因之表現，直到營養生長時期才有表現，因此可知OsC1確實影響著水稻葉鞘顏色之於不同生長時期變化。Anthocyanin can accumulate in several tissues in rice, leading to purple red or purple phenotypes for resistance to biotic and abiotic stresses. Purple and green leaf sheath with and without anthocyanin accumulation, respectively, can be found not only wild rice but also in cultivated rice. Japonica TNG 72 has purple leaf sheath, while indica TCS 17 has green leaf sheath. Based on the positional cloning approach, we used the F2 progenies of TNG 72 × TCS 17 to isolate the gene conferring purple leaf sheath in this segregating population, which was OsC1. Sequencing the OsC1 of 3 cultivars with green leaf sheath was found nonsynonymous SNP and deletion in the regulation region R2R3 Myb domain.. Thus, variation in OsC1 sequence changes the colors of the rice leaf sheath. In addition,, we used 36 accesions include cultivars, landraces and wild rice to study molecular evolution of OsC1, and neutrality tests, such as Tajama’s D, Fu and Li’s D, and Fu and Li’s F, revealed OsC1 alleles might be impacted by natural selection or artificial selection during rice evolution or domestication. To determine the correlation between OsC1 and the anthocyanin accumulation of leaf sheath, we used four culativars, TNG 67, TNG 72, Shang-Chi-Tsao-Tao and Kun-Shan-Wu-Siang-Keng with different levels of anthocyanin content to detect gene expression of OsC1, OsB2, OsDfr, and OsAns by real-time PCR. The results showed that gene expression of OsC1 only but not the other threewas positively correlated with anthocyanin content. Gene expression of OsC1 was high in varieties with purple leaf sheath and increased along development stages. By the GUS promoter assay, OsC1 did not express in the leaf sheath during the 3-leaf stage but did express until the vegetative stage. Thus, gene function and expression of OsC1 really affect the colors of rice leaf sheath in different development stages

Genetic and Evolutionary Analysis of Purple Leaf Sheath in Rice

Background: Anthocyanin accumulates in many plant tissues or organs, in rice for example leading to red, purple red and purple phenotypes for protection from damage by biotic and abiotic stresses and for reproduction. Purple leaf, leaf sheath, stigma, pericarp, and apiculus are common in wild rice and landraces and occasionally found in modern cultivars. No gene directly conferring anthocyanin deposited in a purple leaf sheath has yet been isolated by using natural variants. An F-2 population derived from ssp. japonica cv. Tainung 72 (TNG72) with purple leaf sheath (PSH) crossed with ssp. indica cv. Taichung Sen 17 (TCS17) with green leaf sheath (GSH) was utilized to isolate a gene conferring leaf sheath color. Results: By positional cloning, 10-and 3-bp deletions in the R2R3 Myb domain of OsC1 were uncovered in GSH varieties TCS17 and Nipponbare, respectively. Allelic diversity, rather than gene expression levels of OsC1, might be responsible for anthocyanin accumulation. Parsimony-based analysis of genetic diversity in 50 accessions, including cultivars, landraces, and A-genome wild rice, suggests that independent mutation occurred in Asian, African, South American, and Australian species, while O. meridionalis had a divergent sequence. OsC1 was thought of as a domestication related gene, with up to 90 % reduction of genetic diversity in GSH; however, no values from three tests showed significant differences from neutral expectations, implying that OsC1 had not been subjected to recent selection. Haplotype network analysis revealed that species from different continents formed unique haplotypes with no gene flow. Two major groups of haplotypes corresponding to 10-bp deletion and other sequences were formed in Asian rice, including O. rufipogon, O. nivara and O. sativa. Introgressions of OsC1 between subspecies through natural and artificial hybridization were not rare. Because artificial and natural selection imposed admixture on rice germplasm in Taiwan, the genealogy of OsC1 might not be congruent with the current distribution of alleles through lineage diversification. Conclusion: OsC1 is responsible for purple leaf sheath, and much new information about OsC1 is provided e.g., new alleles, non-domestication syndrome, and incongruence of genealogy with geographic distribution

Additional file 1: Table S1. of Genetic and Evolutionary Analysis of Purple Leaf Sheath in Rice

Newly designed markers used in coarse mapping. (DOCX 17 kb

The geographic distributions and complex genetic relationships among four Sorghum taxa identified in Taiwan

The genus Sorghum consists of 25 species, including Sorghum bicolor (L.) Moench, one of the top five cereal crops cultivated globally, and S. halepense, one of the most noxious weeds. Weedy Sorghum possesses outstanding adaptability and drought tolerance thrives in diverse environments and becomes an invasive plant worldwide. Taiwan is a unique place possessing suitable habitats for four Sorghum taxa, S. bicolor ssp. bicolor, var. technicum, ssp. verticilliflorum and S. halepense, which were identified by key morphological features. The four Sorghum taxa showed distinct geographic distributions, revealing that invasiveness was influenced by their own characteristics and human activities. The sporadic distributions of cultivated S. bicolor ssp. bicolor and var. technicum may be attributed to human disturbance and agricultural activities. The rhizomatous S. halepense was widely distributed and showed the highest genetic diversity (He > 0.776) among the four taxa, with its strong adaptation to various environments threatening the agricultural practices and ecosystem in Taiwan. In contrast, the newly naturalised S. bicolor ssp. verticilliflorum was confined to and dominant in southern Taiwan, with the lowest genetic diversity (He < 0.272). Significant genetic differentiation (F-ST = 0.5207) between the two ssp. verticilliflorum subpopulations was associated with natural geographic isolation. This study concretely elucidated the geographic distributions, genetic diversity and relatedness of invasive and escaped Sorghum taxa, indicating the potential aggressiveness and hazard of weedy Sorghum in Taiwan

Anti-cancer activity of an osthole derivative, NBM-T-BMX-OS01: targeting vascular endothelial growth factor receptor signaling and angiogenesis.

Angiogenesis occurs during tissue growth, development and wound healing. It is also required for tumor progression and represents a rational target for therapeutic intervention. NBM-T-BMX-OS01 (BMX), derived from the semisynthesis of osthole, an active ingredient isolated from Chinese herb Cnidium monnieri (L.) Cuss., was recently shown to enhance learning and memory in rats. In this study, we characterized the anti-angiogenic activities of NBM-T-BMX-OS01 (BMX) in an effort to develop novel inhibitors to suppress angiogenesis and tumor growth. BMX inhibited vascular endothelial growth factor (VEGF)-induced proliferation, migration and endothelial tube formation in human umbilical endothelial cells (HUVECs). BMX also attenuated VEGF-induced microvessel sprouting from aortic rings ex vivo and reduced HCT116 colorectal cancer cells-induced angiogenesis in vivo. Moreover, BMX inhibited the phosphorylation of VEGFR2, FAK, Akt and ERK in HUVECs exposed to VEGF. BMX was also shown to inhibit HCT116 cell proliferation and to suppress the growth of subcutaneous xenografts of HCT116 cells in vivo. Taken together, this study provides evidence that BMX modulates vascular endothelial cell remodeling and leads to the inhibition of tumor angiogenesis. These results also support the role of BMX as a potential drug candidate and warrant the clinical development in the treatment of cancer

Epithelial cell adhesion molecule (EpCAM) regulates HGFR signaling to promote colon cancer progression and metastasis

Abstract Background Epithelial cell adhesion molecule (EpCAM) is known to highly expression and promotes cancer progression in many cancer types, including colorectal cancer. While metastasis is one of the main causes of cancer treatment failure, the involvement of EpCAM signaling in metastatic processes is unclear. We propose the potential crosstalk of EpCAM signaling with the HGFR signaling in order to govern metastatic activity in colorectal cancer. Methods Immunoprecipitation (IP), enzyme-linked immunosorbent assay (ELISA), and fluorescence resonance energy transfer (FRET) was conducted to explore the extracellular domain of EpCAM (EpEX) and HGFR interaction. Western blotting was taken to determine the expression of proteins in colorectal cancer (CRC) cell lines. The functions of EpEX in CRC were investigated by proliferation, migration, and invasion analysis. The combined therapy was validated via a tail vein injection method for the metastasis and orthotopic colon cancer models. Results This study demonstrates that the EpEX binds to HGFR and induces downstream signaling in colon cancer cells. Moreover, EpEX and HGF cooperatively mediate HGFR signaling. Furthermore, EpEX enhances the epithelial-to-mesenchymal transition and metastatic potential of colon cancer cells by activating ERK and FAK-AKT signaling pathways, and it further stabilizes active β-catenin and Snail proteins by decreasing GSK3β activity. Finally, we show that the combined treatment of an anti-EpCAM neutralizing antibody (EpAb2-6) and an HGFR inhibitor (crizotinib) significantly inhibits tumor progression and prolongs survival in metastatic and orthotopic animal models of colon cancer. Conclusion Our findings illuminate the molecular mechanisms underlying EpCAM signaling promotion of colon cancer metastasis, further suggesting that the combination of EpAb2-6 and crizotinib may be an effective strategy for treating cancer patients with high EpCAM expression

Linking CO to Urea Production from CO₂ and NO₃^–/NO₂^– Co-Electrolysis on Transition Metals

Electrochemical reduction of CO2 (CO2ER) has the potential to advance carbon neutrality and renewable energy storage. Advanced CO2ER catalysts can selectively produce a wide array of products. Their importance is amplified when coreducing CO2 with nitrate/nitrite ions (NO3–/NO2–) to generate organic compounds containing C–N bonds, enhancing product diversity and value. Some transition metals effectively catalyze the coreduction of CO2 and NO3–/NO2– to yield urea. However, a disparity exists between the experimental observations that underscore the significance of CO production in urea synthesis and the theoretical perspectives that dismiss the role of CO in C–N bond creation. To reconcile this disparity, we utilized density functional theory combined with a constant electrode potential model to investigate four facile CO2 + *N1 (the intermediates from NO3–/NO2– reduction to NH3) couplingsrepresenting the primary C–N formation pathways on a range of transition metal surfaces. Our comprehensive study elucidates the relationships among C–N coupling barriers, *N1, and CO adsorption energies. Notably, we found that while CO is not involved in C–N formation, a catalyst’s proficiency in generating CO from CO2ER is indicative of its reduced adsorption strength. This result indicates a heightened reactivity in forming C–N bonds via the CO2 + *N1 couplings. Our theoretical exploration adeptly bridges the discrepancies observed between earlier experimental and theoretical studies

Linking CO to Urea Production from CO₂ and NO₃^–/NO₂^– Co-Electrolysis on Transition Metals

Electrochemical reduction of CO2 (CO2ER) has the potential to advance carbon neutrality and renewable energy storage. Advanced CO2ER catalysts can selectively produce a wide array of products. Their importance is amplified when coreducing CO2 with nitrate/nitrite ions (NO3–/NO2–) to generate organic compounds containing C–N bonds, enhancing product diversity and value. Some transition metals effectively catalyze the coreduction of CO2 and NO3–/NO2– to yield urea. However, a disparity exists between the experimental observations that underscore the significance of CO production in urea synthesis and the theoretical perspectives that dismiss the role of CO in C–N bond creation. To reconcile this disparity, we utilized density functional theory combined with a constant electrode potential model to investigate four facile CO2 + *N1 (the intermediates from NO3–/NO2– reduction to NH3) couplingsrepresenting the primary C–N formation pathways on a range of transition metal surfaces. Our comprehensive study elucidates the relationships among C–N coupling barriers, *N1, and CO adsorption energies. Notably, we found that while CO is not involved in C–N formation, a catalyst’s proficiency in generating CO from CO2ER is indicative of its reduced adsorption strength. This result indicates a heightened reactivity in forming C–N bonds via the CO2 + *N1 couplings. Our theoretical exploration adeptly bridges the discrepancies observed between earlier experimental and theoretical studies