Soybean RVE8a confers salt and drought tolerance in Arabidopsis.

Soybean is an economically important crop, which often suffers various abiotic stresses. REVEILLE (RVE) genes have been generally considered as circadian oscillators to mediate diverse developmental processes and plant response to environmental stresses. Addressing their roles is of significance for utilizing them to enhance agronomic traits in crops. However, our understanding of soybean RVEs is extremely limited. In the study, we investigated the expression patterns of soybean CCA1-like genes under salt stress using our RNA-Seq data. Subsequently, a salt stress-inducible gene, GmRVE8a, was chosen for further study. Phylogenetic analysis indicated that GmRVE8a is most closely related to Arabidopsis RVE4 and RVE8. Also, GmRVE8a showed circadian expression pattern with 24 h rhythmic period, suggesting that it might be a clock-regulated gene. Moreover, transgenic Arabidopsis lines over-expressing GmRVE8a were generated. It was observed that ectopic over-expression of GmRVE8a caused a significant delay in flowering. Further observation indicated that under salt and drought stress, transgenic seedlings were stronger than wild type. Consistently, three-week-old transgenic plants grew better than wild type under salt and drought conditions, and the MDA content in transgenic lines was significantly lower than wild type, suggesting that GmRVE8a might be a positive regulator in response to salt and drought stress. Intriguingly, Y2H assay indicated that GmRVE8a physically interacted with a drought-tolerant protein, GmNAC17. Overall, our findings provided preliminary information regarding the functional roles of GmRVE8a in response to salt and drought stress.

The miR156/SPL12 module orchestrates fruit colour change through directly regulating ethylene production pathway in blueberry.

Colour change is an important event during fruit ripening in blueberry. It is well known that miR156/SPLs act as regulatory modules mediating anthocyanin biosynthesis and ethylene plays critical roles during colour change, but the intrinsic connections between the two pathways remain poorly understood. Previously, we demonstrated that blueberry VcMIR156a/VcSPL12 affects the accumulation of anthocyanins and chlorophylls in tomato and Arabidopsis. In this study, we first showed that VcMIR156a overexpression in blueberry led to enhanced anthocyanin biosynthesis, decreased chlorophyll accumulation, and, intriguingly, concomitant elevation in the expression of ethylene biosynthesis genes and the level of the ethylene precursor ACC. Conversely, VcSPL12 enhanced chlorophyll accumulation and suppressed anthocyanin biosynthesis and ACC synthesis in fruits. Moreover, the treatment with ethylene substitutes and inhibitors attenuated the effects of VcMIR156a and VcSPL12 on pigment accumulation. Protein-DNA interaction assays indicated that VcSPL12 could specifically bind to the promoters and inhibit the activities of the ethylene biosynthetic genes VcACS1 and VcACO6. Collectively, our results show that VcMIR156a/VcSPL12 alters ethylene production through targeting VcACS1 and VcACO6, therefore governing fruit colour change. Additionally, VcSPL12 may directly interact with the promoter region of the chlorophyll biosynthetic gene VcDVR, thereby activating its expression. These findings established an intrinsic connection between the miR156/SPL regulatory module and ethylene pathway.

Curcumin inhibits the invasion and migration of pancreatic cancer cells by upregulating TFPI-2 to regulate ERK- and JNK-mediated epithelial-mesenchymal transition.

PURPOSE: Pancreatic cancer (PC) is one of the most deadly human malignancies. Curcumin is a natural polyphenolic compound with wide-ranging pharmacological effects. Growing evidence suggests that curcumin has anticancer activity against PC, but the mechanism remains incompletely elucidated. This study aimed to investigate the effects and mechanisms of curcumin on the invasion and migration of PC cells. METHODS: Effect of curcumin on tissue factor pathway inhibitor (TFPI)-2 mRNA expression in PC cells was initially identified using qRT-PCR. Cytotoxicity of curcumin was assessed with MTT assays and IC50 was calculated. Involvement of ERK and JNK pathways, as well as protein expression of TFPI-2 and epithelial-mesenchymal transition (EMT)-related markers, were detected using immunoblotting. Invasion and migration of PC cells were examined using Transwell assays. TFPI-2 expression was manipulated by transfection with siRNA and shRNA. Rescue assays were used to validate the effect of curcumin on cell invasion and migration via TFPI-2. RESULTS: Curcumin increased the expression of TFPI-2 mRNA and protein in PC cells and attenuated cell invasion and migration. Curcumin also inhibited ERK and JNK pathways and EMT in PC cells. Knockdown of TFPI-2 partially reversed the inhibition of ERK and JNK pathways and EMT by curcumin. Mechanistically, curcumin upregulated TFPI-2, thereby inhibiting the ERK and JNK pathways, leading to the inhibition of EMT in PC cells. CONCLUSION: Collectively, curcumin inhibits ERK- and JNK-mediated EMT through upregulating TFPI-2, which in turn suppresses the migration and invasion of PC cells. These findings provide new insights into the antitumor mechanism of curcumin.

Identification of R2R3-MYB family in blueberry and its potential involvement of anthocyanin biosynthesis in fruits.

BACKGROUND: Blueberries (Vaccinium corymbosum) are regarded as "superfoods" attributed to large amounts of anthocyanins, a group of flavonoid metabolites, which provide pigmentation in plant and beneficial effects for human health. MYB transcription factor is one of vital components in the regulation of plant secondary metabolism, which occupies a dominant position in the regulatory network of anthocyanin biosynthesis. However, the role of MYB family in blueberry responding to anthocyanin biosynthesis remains elusive. RESULTS: In this study, we conducted a comprehensive analysis of VcMYBs in blueberry based on the genome data, including phylogenetic relationship, conserved motifs, identification of differentially expressed MYB genes during fruit development and their expression profiling, etc. A total of 437 unique MYB sequences with two SANT domains were identified in blueberry, which were divided into 3 phylogenetic trees. Noticeably, there are many trigenic and tetragenic VcMYBs pairs with more than 95% identity to each other. Meanwhile, the transcript accumulations of VcMYBs were surveyed underlying blueberry fruit development, and they showed diverse expression patterns, suggesting various functional roles in fruit ripening. More importantly, distinct transcript profiles between skin and pulp of ripe fruit were observed for several VcMYBs, such as VcMYB437, implying the potential roles in anthocyanin biosynthesis. CONCLUSIONS: Totally, 437 VcMYBs were identified and characterized. Subsequently, their transcriptional patterns were explored during fruit development and fruit tissues (skin and pulp) closely related to anthocyanin biosynthesis. These genome-wide data and findings will contribute to demonstrating the functional roles of VcMYBs and their regulatory mechanisms for anthocyanins production and accumulation in blueberry in the future study.

Genome-wide identification of BBX gene family and their expression patterns under salt stress in soybean.

BACKGROUND: BBX genes are key players in the regulation of various developmental processes and stress responses, which have been identified and functionally characterized in many plant species. However, our understanding of BBX family was greatly limited in soybean. RESULTS: In this study, 59 BBX genes were identified and characterized in soybean, which can be phylogenetically classified into 5 groups. GmBBXs showed diverse gene structures and motif compositions among the groups and similar within each group. Noticeably, synteny analysis suggested that segmental duplication contributed to the expansion of GmBBX family. Moreover, our RNA-Seq data indicated that 59 GmBBXs showed different transcript profiling under salt stress, and qRT-PCR analysis confirmed their expression patterns. Among them, 22 GmBBXs were transcriptionally altered with more than two-fold changes by salt stress, supporting that GmBBXs play important roles in soybean tolerance to salt stress. Additionally, Computational assay suggested that GmBBXs might potentially interact with GmGI3, GmTOE1b, GmCOP1, GmCHI and GmCRY, while eight types of transcription factors showed potentials to bind the promoter regions of GmBBX genes. CONCLUSIONS: Fifty-nine BBX genes were identified and characterized in soybean, and their expression patterns under salt stress and computational assays suggested their functional roles in response to salt stress. These findings will contribute to future research in regard to functions and regulatory mechanisms of soybean BBX genes in response to salt stress.

Effects of genotype and culture conditions on microspore embryogenesis in radish (Raphanus sativus L.).

Radish (Raphanus sativus L.), an important annual or biennial root vegetable crop, is widely cultivated in the world for its high nutritive value. Isolated microspore culture (IMC) is one of the most effective methods for rapid development of homozygous lines. Due to imperfection of the IMC technology system, it is particularly important to establish an efficient IMC system in radish. In this study, the effects of different factors on radish microspore embryogenesis were investigated with 23 genotypes. Buds with the largest population of late-uninucleate-stage microspores were most suitable for embryogenesis, with a ratio of petal length to anther length (P/A) in buds of about 3/4 ~ 1. Cold pretreatment was found to be genotype specific, and the highest microspore-derived embryoid (MDE) yield occurred for treatment of the heat shock of 48 h. In addition, the supplement of 0.75 g/L activated charcoal (AC) could increase the yield of embryoids. It was found that genotypes, bud size, as well as temperature treatments had significant effects on microspore embryogenesis. Furthermore, somatic embryogenesis-related kinase (SERK) genes were profiled by reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis, which indicated that they are involved in the process of MDE formation and plantlet regeneration. The ploidy of microspore-derived plants was identified by chromosome counting and flow cytometry, and the microspore-derived plants were further proved as homozygous plants through expressed sequence tags-simple sequence repeats (EST-SSR) and genetic-SSR markers. The results would facilitate generating the large-scale double haploid (DH) from various genotypes, and promoting further highly efficient genetic improvement in radish. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01312-w.

A meta-analysis about the accumulation of heavy metals uptake by Sedum alfredii and Sedum plumbizincicola in contaminated soil.

Sedum alfredii and Sedum plumbizincicola typically have high heavy metal (such as Zn and Cd) accumulation capacities with fast growth rates and relatively high Pb tolerance in contaminated soils. We compared the accumulation characteristics of heavy metals in Sedum species through meta-analysis. Furthermore, we analyzed the effects of soil organic matter (SOM) and soil pH on Cd, Pb and Zn accumulation by S. alfredii and S. plumbizincicola and the correlation between various metals. Results showed that the accumulations of Cd and Zn in shoots were higher than that of roots, but Pb accumulated in roots more than shoots. Moreover, there is a significant positive correlation between the accumulation of Zn and Cd in shoots. We found that the heavy metal accumulation rate in shoots was higher with lower soil pH. Sedum species had the highest Cd adsorption capacity in 20-30 g/kg SOM and the highest Zn adsorption capacity in SOM less than 20 g/kg. The accumulation rate of Cd in shoots of S. plumbizincicola was increased with exposure time, while the accumulation rate of Zn was slightly decreased.

S. alfredii and S. plumbizincicola are two common Cd and Zn hyperaccumulators. We systematically compared the accumulation characteristics of heavy metals in Sedum plants and the effects of soil organic matter and pH on the accumulation of Cd, Pb and Zn in S. alfredii and S. plumbizincicola through meta-analysis. This provides certain theoretical knowledge for the application of sedum plants to the phytoremediation of heavy metal contaminated soils.

Analysis of the long-term effectiveness of biochar immobilization remediation on heavy metal contaminated soil and the potential environmental factors weakening the remediation effect: A review.

Currently, the research and application of biochar in the remediation of heavy metal contaminated soil has become a hotspot, especially regarding the remediation of agricultural land. Biochar has been proved to be effective in reducing the content of available heavy metals in the soil as well as the heavy metals in plants. However, the long-term effectiveness of biochar immobilization has not been widely studied. In this review, retrospective search was carried out on the published literature results concerning remediation effects of biochar on different areas of heavy metal contaminated soil in the recent years, its application in field remediation (several years), and some potential abiotic and biotic factors that may weaken the immobilization effects of biochar. This results indicate that: (1) biochar is widely used in the remediation of heavy metal contaminated soil in different areas and has excellent immobilization effect. (2) Most of the research demonstrate that the immobilization effect of biochar is effective for 2-3 years or according to few results even for 5 years. However, there have been various reports claiming that the immobilization effect of biochar decreases with time. (3) Abiotic factors such as acid rain, flooded environment, changes in soil condition (pH, redox and dissolved organic matter) and changes in biochar (Cl- and alkali leaching) can significantly weaken the immobilization effect of biochar. (4) Biotic factors such as plant roots, earthworms and soil microorganisms can also significantly reduce the immobilization effect of biochar. Therefore, field experiments having longer time span with biochar need to be further carried out, and the developmental research of modified biochar with a more stable immobilization effect also needs further attention.

A blueberry MIR156a-SPL12 module coordinates the accumulation of chlorophylls and anthocyanins during fruit ripening.

Color change is an important event during fruit maturation in blueberry, usually depending on chlorophyll degradation and anthocyanin accumulation. MicroRNA156 (miR156)-SPL modules are an important group of regulatory hubs involved in the regulation of anthocyanin biosynthesis. However, little is known regarding their roles in blueberry or in chlorophyll metabolism during color change. In this study, a MIR156 gene (VcMIR156a) was experimentally identified in blueberry (Vaccinium corymbosum). Overexpression of VcMIR156a in tomato (Solanum lycopersicum) enhanced anthocyanin biosynthesis and chlorophyll degradation in the stem by altering pigment-associated gene expression. Further investigation indicated that the VcSPL12 transcript could be targeted by miR156, and showed the reverse accumulation patterns during blueberry fruit development and maturation. Noticeably, VcSPL12 was highly expressed at green fruit stages, while VcMIR156a transcripts mainly accumulated at the white fruit stage when expression of VcSPL12 was dramatically decreased, implying that VcMIR156a-VcSPL12 is a key regulatory hub during fruit coloration. Moreover, VcSPL12 decreased the expression of several anthocyanin biosynthetic and regulatory genes, and a yeast two-hybrid assay indicated that VcSPL12 interacted with VcMYBPA1. Intriguingly, expression of VcSPL12 significantly enhanced chlorophyll accumulation and altered the expression of several chlorophyll-associated genes. Additionally, the chloroplast ultrastructure was altered by the expression of VcMIR156a and VcSPL12. These findings provide a novel insight into the functional roles of miR156-SPLs in plants, especially in blueberry fruit coloration.

Soybean CCA1-like MYB transcription factor GmMYB133 modulates isoflavonoid biosynthesis.

MYB transcription factors play important roles in the regulation of phenylpropanoid biosynthesis. However, the knowledge regarding the roles of CCA1-like MYBs in phenylpropanoid pathway is limited in plants. Previously, we identified 54 CCA1-like proteins in soybean. In the study, a CCA1-like MYB (GmMYB133) was functionally characterized as a positive regulator in isoflavonoid synthesis. GmMYB133 encodes a 330 aa protein featured with one CCA1 conserved motif. Further analysis indicated that the expression pattern of GmMYB133 was near-perfectly correlated with isoflavonoid accumulation as soybean embryos develop. GmMYB133 over-expression promoted the expression of two key isoflavonoid biosynthetic genes (GmCHS8 and GmIFS2) and increased total isoflavonoid content in hairy roots. Protein-protein interaction assays indicated that GmMYB133 might form hetero- and homodimers with an isoflavonoid regulator GmMYB176 and itself, respectively, while the subcellular localization of GmMYB133 can be altered by its interaction with 14-3-3 protein. These findings provided new insights into the functional roles of CCA1-like MYB proteins in the regulation of phenylpropanoid pathway, and will contribute to the future genetic engineering in the improvement of soybean seed quality.

[Mediating effect of psychological symptoms, coping styles and impulsiveness on the relationship between childhood abuses and non-suicidal self-injuries among middle school students].

OBJECTIVE: To estimate the mediationl effects of psychological symptoms, coping styles and impulsiveness on relationship between childhood abuse and non-suicidal self-injuries( NSSI) among middle school students. METHODS: A total of 9704 participants( boys( 4600), girls( 5104); urban( 4049), rural( 5655); junior( 5168), senior( 4536)) were retained from all epidemiological study, involving students from junior and senior middle schools in Zhengzhou, Guiyang by random cluster sampling. The Childhood Trauma Questionnaire( CTQ-SF), Non-Suicidal Self-Injury Questionnaire, Multidimensional Sub-health Questionnaire of Adolescents( MSQA), Trait Coping Style Questionnaire( TCSQ) and Barratt Impulsiveness Scale( BIS-11) were used to measure childhood abuse assessment, NSSI frequency, psychological symptoms, coping styles and impulsiveness scores. Chi-square test was used to compare the frequency of childhood abuse and NSSI among different groups. Pearson's correlation analysis was used in analyzing the relationship, Bootstrap method and PROCESS software were applied to examine the mediationl effects. RESULTS: A total of 44. 6% of middle school students had childhood abuse experiences, girls were higher than boys( 45. 9% vs. 43. 1%), junior secondary students were higher than senior secondary students( 47. 3% vs. 41. 5%), the differences were statistically significant( χ~2= 7. 558 and 32. 433; P < 0. 01). However, there was no statistical significance in the residence area( P > 0. 05). 38. 5% of high school students had NSSI, of which girls were lower than boys( 37. 1% vs. 40. 1%), junior secondary students were higher than senior secondary students( 41. 4% vs. 35. 3%), the differences were statistically significant( χ~2= 9. 151 and 37. 955; P <0. 01). The middle of perceived family SES who the detection rate of childhood abuse and NSSI were lower than poor and high family SES, the differences were statistically significant( χ~2= 9. 916 and 25. 735; P < 0. 01). The psychological symptoms, coping style and impulsiveness scores were related with childhood abuse scores and NSSI frequency respectively( P < 0. 01). These mediating effects persisted after controlling for sex, grade, residence area and perceived family SES, the total mediationl effects was 52. 24%, specific mediationl effect of psychological symptoms, negative and positive coping styles, impulsiveness was 33. 41%, 14. 87%, 0. 49% and 3. 48%( P < 0. 01). CONCLUSION: It suggests that psychological symptoms, coping styles, impulsiveness may play a partial mediating effect on the relationship between childhood abuse and NSSI among middle school students. The alleviating of psychological symptoms, negative coping style may contribute to the prevention and control of NSSI among middle school students who have experienced childhood abuse.

Conservation and diversification of the miR166 family in soybean and potential roles of newly identified miR166s.

BACKGROUND: microRNA166 (miR166) is a highly conserved family of miRNAs implicated in a wide range of cellular and physiological processes in plants. miR166 family generally comprises multiple miR166 members in plants, which might exhibit functional redundancy and specificity. The soybean miR166 family consists of 21 members according to the miRBase database. However, the evolutionary conservation and functional diversification of miR166 family members in soybean remain poorly understood. RESULTS: We identified five novel miR166s in soybean by data mining approach, thus enlarging the size of miR166 family from 21 to 26 members. Phylogenetic analyses of the 26 miR166s and their precursors indicated that soybean miR166 family exhibited both evolutionary conservation and diversification, and ten pairs of miR166 precursors with high sequence identity were individually grouped into a discrete clade in the phylogenetic tree. The analysis of genomic organization and evolution of MIR166 gene family revealed that eight segmental duplications and four tandem duplications might occur during evolution of the miR166 family in soybean. The cis-elements in promoters of MIR166 family genes and their putative targets pointed to their possible contributions to the functional conservation and diversification. The targets of soybean miR166s were predicted, and the cleavage of ATHB14-LIKE transcript was experimentally validated by RACE PCR. Further, the expression patterns of the five newly identified MIR166s and 12 target genes were examined during seed development and in response to abiotic stresses, which provided important clues for dissecting their functions and isoform specificity. CONCLUSION: This study enlarged the size of soybean miR166 family from 21 to 26 members, and the 26 soybean miR166s exhibited evolutionary conservation and diversification. These findings have laid a foundation for elucidating functional conservation and diversification of miR166 family members, especially during seed development or under abiotic stresses.

Overexpressed HSPA2 correlates with tumor angiogenesis and unfavorable prognosis in pancreatic carcinoma.

Heat shock-related 70-kDa protein 2 (HSPA2) is known to correlate with tumor development and progression. This work aimed to determine the expression and prognostic roles of HSPA2 in pancreatic carcinoma. Tumor and their corresponding non-tumor tissues were obtained from 80 patients with pancreatic carcinoma. HSPA2 expression in tumor and non-tumor tissues was evaluated by immunohistochemistry. Expression of vascular endothelial growth factor (VEGF) and CD31 in tumor tissues were also evaluated by immunostaining. The relationships of HSPA2 with clinicopathological data, tumor angiogenesis and prognosis were analyzed. The results showed that HSPA2 expression was significantly elevated in tumor tissues compared with adjacent non-tumor tissues (P < 0.05). High HSPA2 expression was significantly associated with aggressive clinicopathological characteristics. HSPA2 staining was positively correlated with VEGF (r = 0.466, P < 0.001) and microvessel density (MVD) (r = 0.366, P = 0.001) in tumor tissues. Patients with high HSPA2 expression showed worse relapse-free survival (RFS) (P < 0.001) and overall survival (OS) (P < 0.001) than those with low HSPA2 expression. Multivariate analysis indicated that high HSPA2 expression was an independent predictor for poor RFS (P < 0.001) and OS (P = 0.001). Taken together, overexpressed HSPA2 is correlated with tumor angiogenesis and poor prognosis in pancreatic carcinoma. HSPA2 may play an important role in tumor progression, and serve as a potential biomarker for the prediction of adverse prognosis in pancreatic carcinoma.

Genome-Wide Analysis of CCA1-Like Proteins in Soybean and Functional Characterization of GmMYB138a.

Plant CIRCADIAN CLOCK ASSOCIATED1 (CCA1)-like proteins are a class of single-repeat MYELOBLASTOSIS ONCOGENE (MYB) transcription factors generally featured by a highly conserved motif SHAQK(Y/F)F, which play important roles in multiple biological processes. Soybean is an important grain legume for seed protein and edible vegetable oil. However, essential understandings regarding CCA1-like proteins are very limited in soybean. In this study, 54 CCA1-like proteins were identified by data mining of soybean genome. Phylogenetic analysis indicated that soybean CCA1-like subfamily showed evolutionary conservation and diversification. These CCA1-like genes displayed tissue-specific expression patterns, and analysis of genomic organization and evolution revealed 23 duplicated gene pairs. Among them, GmMYB138a was chosen for further investigation. Our protein-protein interaction studies revealed that GmMYB138a, but not its alternatively spliced isoform, interacts with a 14-3-3 protein (GmSGF14l). Although GmMYB138a was predominately localized in nucleus, the resulting complex of GmMYB138a and GmSGF14l was almost evenly distributed in nucleus and cytoplasm, supporting that 14-3-3s interact with their clients to alter their subcellular localization. Additionally, qPCR analysis suggested that GmMYB138a and GmSGF14l synergistically or antagonistically respond to drought, cold and salt stresses. Our findings will contribute to future research in regard to functions of soybean CCA1-like subfamily, especially regulatory mechanisms of GmMYB138a in response to abiotic stresses.

Comparative Analysis of Fruit Ripening-Related miRNAs and Their Targets in Blueberry Using Small RNA and Degradome Sequencing.

MicroRNAs (miRNAs) play vital roles in the regulation of fruit development and ripening. Blueberry is an important small berry fruit crop with economical and nutritional value. However, nothing is known about the miRNAs and their targets involved in blueberry fruit ripening. In this study, using high-throughput sequencing of small RNAs, 84 known miRNAs belonging to 28 families and 16 novel miRNAs were identified in white fruit (WF) and blue fruit (BF) libraries, which represent fruit ripening onset and in progress, respectively. Among them, 41 miRNAs were shown to be differentially expressed during fruit maturation, and 16 miRNAs representing 16 families were further chosen to validate the sRNA sequencing data by stem-loop qRT-PCR. Meanwhile, 178 targets were identified for 41 known and 7 novel miRNAs in WF and BF libraries using degradome sequencing, and targets of miR160 were validated using RLM-RACE (RNA Ligase-Mediated (RLM)-Rapid Amplification of cDNA Ends) approach. Moreover, the expression patterns of 6 miRNAs and their targets were examined during fruit development and ripening. Finally, integrative analysis of miRNAs and their targets revealed a complex miRNA-mRNA regulatory network involving a wide variety of biological processes. The findings will facilitate future investigations of the miRNA-mediated mechanisms that regulate fruit development and ripening in blueberry.