The Quantitative Biotinylproteomics Studies Reveal a WInd-Related Kinase 1 (Raf-Like Kinase 36) Functioning as an Early Signaling Component in Wind-Induced Thigmomorphogenesis and Gravitropism.

Wind is one of the most prevalent environmental forces entraining plants to develop various mechano-responses, collectively called thigmomorphogenesis. Largely unknown is how plants transduce these versatile wind force signals downstream to nuclear events and to the development of thigmomorphogenic phenotype or anemotropic response. To identify molecular components at the early steps of the wind force signaling, two mechanical signaling-related phosphoproteins, identified from our previous phosphoproteomic study of Arabidopsis touch response, mitogen-activated protein kinase kinase 1 (MKK1) and 2 (MKK2), were selected for performing in planta TurboID (ID)-based quantitative proximity-labeling (PL) proteomics. This quantitative biotinylproteomics was separately performed on MKK1-ID and MKK2-ID transgenic plants, respectively, using the genetically engineered TurboID biotin ligase expression transgenics as a universal control. This unique PTM proteomics successfully identified 11 and 71 MKK1 and MKK2 putative interactors, respectively. Biotin occupancy ratio (BOR) was found to be an alternative parameter to measure the extent of proximity and specificity between the proximal target proteins and the bait fusion protein. Bioinformatics analysis of these biotinylprotein data also found that TurboID biotin ligase favorably labels the loop region of target proteins. A WInd-Related Kinase 1 (WIRK1), previously known as rapidly accelerated fibrosarcoma (Raf)-like kinase 36 (RAF36), was found to be a putative common interactor for both MKK1 and MKK2 and preferentially interacts with MKK2. Further molecular biology studies of the Arabidopsis RAF36 kinase found that it plays a role in wind regulation of the touch-responsive TCH3 and CML38 gene expression and the phosphorylation of a touch-regulated PATL3 phosphoprotein. Measurement of leaf morphology and shoot gravitropic response of wirk1 (raf36) mutant revealed that the WIRK1 gene is involved in both wind-triggered rosette thigmomorphogenesis and gravitropism of Arabidopsis stems, suggesting that the WIRK1 (RAF36) protein probably functioning upstream of both MKK1 and MKK2 and that it may serve as the crosstalk point among multiple mechano-signal transduction pathways mediating both wind mechano-response and gravitropism.

The Integration of Genome-Wide DNA Methylation and Transcriptomics Identifies the Potential Genes That Regulate the Development of Skeletal Muscles in Ducks.

DNA methylation is a pivotal epigenetic regulatory mechanism in the development of skeletal muscles. Nonetheless, the regulators responsible for DNA methylation in the development of embryonic duck skeletal muscles remain unknown. In the present study, whole genome bisulfite sequencing (WGBS) and transcriptome sequencing were conducted on the skeletal muscles of embryonic day 21 (E21) and day 28 (E28) ducks. The DNA methylation pattern was found to fall mainly within the cytosine-guanine (CG) context, with high methylation levels in the intron, exon, and promoter regions. Overall, 7902 differentially methylated regions (DMRs) were identified, which corresponded to 3174 differentially methylated genes (DMGs). By using integrative analysis of both WGBS with transcriptomics, we identified 1072 genes that are DMGs that are negatively associated with differentially expressed genes (DEGs). The gene ontology (GO) analysis revealed significant enrichment in phosphorylation, kinase activity, phosphotransferase activity, alcohol-based receptors, and binding to cytoskeletal proteins. The Kyoto Encyclopedia of Genes and Genomes (KEGGs) analysis showed significant enrichment in MAPK signaling, Wnt signaling, apelin signaling, insulin signaling, and FoxO signaling. The screening of enriched genes showed that hyper-methylation inhibited the expression of Idh3a, Got1, Bcl2, Mylk2, Klf2, Erbin, and Klhl38, and hypo-methylation stimulated the expression of Col22a1, Dnmt3b, Fn1, E2f1, Rprm, and Wfikkn1. Further predictions showed that the CpG islands in the promoters of Klhl38, Klf2, Erbin, Mylk2, and Got1 may play a crucial role in regulating the development of skeletal muscles. This study provides new insights into the epigenetic regulation of the development of duck skeletal muscles.

Knockdown of Tet2 Inhibits the Myogenic Differentiation of Chicken Myoblasts Induced by Ascorbic Acid.

Ascorbic acid (also called Vitamin C, VC) strengthens the function of Tets families and directly increases DNA demethylation level to affect myogenic differentiation. However, the precise regulatory mechanism of DNA methylation in chicken myogenesis remains unclear. Results of present study showed that the mRNA expression of MyoD significantly decreased and MyoG and MyHC increased in myoblasts treated with 5 µM 5-azacytidine (5-AZA) and 5 µM VC (p < 0.05). Results also indicated the formation of myotubes was induced by 5-AZA or VC, but this effect was attenuated after knockdown of Tet2. In addition, the protein expression of TET2, DESMIN and MyHC was remarkable increased by the addition of 5-AZA or VC, and the upregulation was inhibited after knockdown of Tet2 (p < 0.05). DNA dot blot and immunofluorescence staining results suggested that the level of 5hmC was significantly increased when treated with 5-AZA or VC, even by Tet2 knockdown (p < 0.05). Moreover, 5-AZA and VC reduced the level of dimethylation of lysine 9 (H3K9me2) and trimethylation of lysine 27 of histone 3 (H3K27me3), and this inhibitory effect was eliminated after Tet2 knockdown (p < 0.05). These data indicated that Tet2 knockdown antagonized the increased levels of 5hmC and H3K27me3 induced by 5-AZA and VC, and eventually reduced myotube formation by modulating the expression of genes involved in myogenic differentiation. This study provides insights that epigenetic regulators play essential roles in mediating the myogenic program of chicken myoblasts.

Testosterone Promotes the Proliferation of Chicken Embryonic Myoblasts Via Androgen Receptor Mediated PI3K/Akt Signaling Pathway.

Testosterone (T) is essential for muscle fiber formation and growth. However, the specific mechanism by which T regulates skeletal muscle development in chicken embryos remains unclear. In this study, the role of T in myoblast proliferation both in vivo and in vitro was investigated. Results showed that the T administration significantly increased the ratio of breast muscle and leg muscle. T induced a significant increase in the cross-sectional area (CSA) and density of myofiber and the ratio of PAX7-positive cells in the skeletal muscle. Exogenous T also induced the upregulation of myogenic regulatory factors (MRFs) and cyclin-dependent kinases (CDK2)/Cyclin D1 (CCND1) and protein levels of androgen receptor (AR), p-Akt and PAX7. Furthermore, T treatment significantly promoted myoblasts cultured in vitro entering a new cell cycle and increased PAX7-positive cells. The mRNA and protein expression of AR and PAX7 were upregulated when treated with T compared to that of the control. The addition of T induced proliferation accompanied by increasing AR level as well as PI3K (Phosphoinositide 3-kinase)/Akt activation. However, T-induced proliferation was attenuated by AR, PI3K, and Akt-specific inhibitors. These data indicated that the pro-proliferative effect of T was regulated though AR in response to the activation of PI3K/Akt signalling pathway.

Identification of chicken FSHR gene promoter and the correlations between polymorphisms and egg production in Chinese native hens.

Egg production is an important economic trait in poultry, and it is of great significance to study the key genes and functional SNPs that affect egg laying performance. Follicle-stimulating hormone (FSH) plays an important physiological role in the reproductive performance of humans and animals by binding to its receptor (FSHR). Studies have shown that there are many transcriptional regulatory elements in the 5' flanking region of the FSHR gene that interact with transcription factors to regulate FSHR transcription. In this study, DNA sequencing was used to identify SNPs in the FSHR promoter sequence in both Dongxiang and Suken chickens. To detect the activity of the chicken FSHR gene promoter, we analysed the characteristics of the sequence and constructed three deletion vectors. We confirmed that the region (-18/-544) was the core promoter. Furthermore, five polymorphisms, including a 200-bp indel at -869, C-1684T, C-1608T, G-368A and T-238A, were detected in both the Dongxiang and Suken chickens. The age at first egg (AFE) for different genotype of -869 indel in Suken chicken was significantly different (p < 0.01). For SNP C-1684T in Dongxiang chickens, the CC genotype had higher egg number at 43 weeks of age (E43) than that of the TC genotype (p < 0.05). For SNP C-1684T in Suken chickens, the TC genotype had higher AFE than that of the CC genotype (p < 0.05). For SNP C-1608T in Suken chickens, the CC genotype had higher AFE than that of the TC genotype (p < 0.05). For SNP G-368A in Suken chickens, the AG genotype had higher AFE than that of the GG genotype (p < 0.05).

Bisphenol A accelerates meiotic progression in embryonic chickens via the estrogen receptor ß signaling pathway.

Bisphenol A (BPA) as an endocrine-disrupting chemical with weak estrogenic activity affects formation of primordial follicles. This study aimed to identify the potential effects and molecular mechanisms of BPA on meiosis and primordial follicle formation in chickens. The results suggest that the cortical layer was thickened and the number of germ cells that entered into meiosis was increased in BPA-treated ovaries. The percentage of γH2AX-positive cells increased significantly. In addition, up-regulated mRNA expression of meiotic genes, including stimulated by retinoic acid gene 8 (Stra8), disrupted meiotic cDNA 1 homologue (Dmc1) and synaptonemal complex protein 3 (Scp3) were observed in BPA-treated ovaries. Therefore, progression to meiosis prophase I was accelerated by exposure to BPA. Furthermore, the results demonstrated that injection of BPA resulted in hypomethylation of Dazl (Deleted in A Zoospermia-Like gene) and Stra8 and up-regulation mRNA expression of Dazl and Stra8 during meiotic onset. Finally, the relationship between estrogen receptor (ER) expression and BPA-induced meiosis was revealed using an in vitro ovarian culture system. BPA enhanced ERß expression at the levels of mRNA and protein, while BPA exerted no significant effect on ERα and membrane-bound estrogen receptor (GPR30) expression. The inducing effects of BPA on meiosis were blocked by ER inhibitor. Collectively, these results demonstrate the dynamic ovarian response to BPA exposure, which indicate that BPA affects the formation of primordial follicles by promoting meiotic progression of oocytes via hypomethylation of Dazl and Stra8 and ERß signaling pathways.

Ebp1 regulates myogenic differentiation of myoblast cells via SMAD2/3 signaling pathway.

Regulation of skeletal muscle development requires many of the regulatory networks that are fundamental to developmental myogenesis. ErbB3 binding protein-1 (Ebp1) is involved in the control of myoblasts development in chicken. However, the expression and biological functions of Ebp1 in the progress of myogenesis are unclear. This study focused on determining the effect of Ebp1 on myogenic proliferation and differentiation using a primary myoblasts culture model. Ebp1 was found to upregulate in proliferating myoblasts and decrease at the early stage of myogenic differentiation. The level of endogenous Ebp1 increased from E9 to E20 chicken leg muscles. Knockdown of Ebp1 had no effect on myoblasts proliferation. However, myogenic differentiation into multinucleated myotubes was significantly reduced. The mRNA and protein expression of MRFs was decreased when Ebp1 was knocked down. Downregulation of Ebp1, accompanied by elevated levels of pSMAD2/3, suggests that Ebp1 is involved in regulating myogenic differentiation via SMAD2/3 inhibition. The phosphorylation of SMAD2/3 was activated and the expression of MYOD and MYOG was reduced in Ebp1 knockdown myoblasts, but addition of LY2109761 (an inhibitor specified to SMAD2/3) blocked these effects. Collectively, these results indicate that Ebp1 promotes myoblast differentiation by inhibition of SMAD2/3 signaling pathway during chicken myogenesis. These data provide new insights into the biological role of Ebp1 in embryonic chicken skeletal muscle development.

Toxicity and Behavioral Effects of Amending Soils with Biochar on Red Imported Fire Ants, Solenopsis invicta.

Solenopsis invicta, often known as the red imported fire ants (RIFAs), is a well-known global invasive ant species that can be found in agricultural, urban, and natural environments worldwide. Simultaneously, it also inhabits the soil. Biochar is generated by the pyrolysis of organic matter under high-temperature anoxic environments and widely used in agricultural ecosystems and soil amendment. However, to date, it remains unknown as to whether soil application of biochar has a negative effect on RIFAs. In our study, we investigated the toxicity and irritability effects of different amounts of biochar (0%, 1%, 2%, 5%, 10%, and 20%) introduced into the soil on red fire ants; upon comparison with the control soil (0% biochar), the application of 1%, 2%, and 5% biochar did not result in significantly different results. But the utilization of biochar at a concentration over 10% effectively repelled the RIFAs, resulting in their departure from the treated soils. High doses of biochar were able to cause death of red fire ants; the mortality rate of red fire ants reached 55.56% after 11 days of 20% biochar treatment. We also evaluated the effects of biochar on four behaviors of red fire ants, namely aggregation, walking, grasping, and attacking; 20% of the biochar treatment group reduced aggregation by 64.22% and this value was 55.22%, 68.44%, and 62.36% for walking, grasping, and attacking. Finally, we measured the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) enzyme activity and malondialdehyde (MDA) content in red fire ants; the results showed that the activities of the three enzymes increased with the increase in biochar addition, which indicated that a high dose of biochar induced oxidative stress in red fire ants. Our results indicate that biochar has the potential to cause toxicity and repel red imported fire ants (RIFAs) in a manner that is dependent on the concentration. We propose that biochar could be utilized in the control and manufacturing of baits for red fire ant management. This work establishes a foundation for the prevention and management of red fire ants and the logical utilization of biochar.

Ten-eleven translocation 1 mediating DNA demethylation regulates the proliferation of chicken primordial germ cells through the activation of Wnt4/ß-catenin signaling pathway.

OBJECTIVE: The objective of this study was to investigate the regulation relationship of Teneleven translocation 1 (Tet1) in DNA demethylation and the proliferation of primordial germ cells (PGCs) in chickens. METHODS: siRNA targeting Tet1 was used to transiently knockdown the expression of Tet1 in chicken PGCs, and the genomic DNA methylation status was measured. The proliferation of chicken PGCs was detected by flow cytometry analysis and cell counting kit-8 assay when activation or inhibition of Wnt4/ß-catenin signaling pathway. And the level of DNA methylation and hisotne methylation was also tested. RESULTS: Results revealed that knockdown of Tet1 inhibited the proliferation of chicken PGCs and downregulated the mRNA expression of Cyclin D1 and cyclin-dependent kinase 6 (CDK6), as well as pluripotency-associated genes (Nanog, PouV, and Sox2). Flow cytometry analysis confirmed that the population of PGCs in Tet1 knockdown group displayed a significant decrease in the proportion of S and G2 phase cells, which meant that there were less PGCs entered the mitosis process than that of control. Furthermore, Tet1 knockdown delayed the entrance to G1/S phase and this inhibition was rescued by treated with BIO. Consistent with these findings, Wnt/ß-catenin signaling was inactivated in Tet1 knockdown PGCs, leading to aberrant proliferation. Further analysis showed that the methylation of the whole genome increased significantly after Tet1 downregulation, while hydroxymethylation obviously declined. Meanwhile, the level of H3K27me3 was upregulated and H3K9me2 was downregulated in Tet1 knockdown PGCs, which was achieved by regulating Wnt/ß-catenin signaling pathway. CONCLUSION: These results suggested that the self-renewal of chicken PGCs and the maintenance of their characteristics were regulated by Tet1 mediating DNA demethylation through the activation of Wnt4/ß-catenin signaling pathway.

Comparative genome-wide association study on body weight in Chinese native ducks using four models.

The Jinling White duck represents a newly developed breed characterized by a rapid growth rate and a superior meat quality, offering significant economic value and research potential; however, the genetic basis underlying their body weight traits remains less understood. Here, we performed whole-genome resequencing for 201 diverse Jinling White male ducks and conducted population genomic analyses, suggesting a rich genetic diversity within the Jinling White duck population. Equipped with our genomic resources, we applied genome-wide association analysis for body weight on birth (BWB), body weight on 1 wk (BW1), body weight on 3 wk (BW3), body weight on 5 wk (BW5) and body weight on 7 wk (BW7) using 4 statistical models. Comparative studies indicated that factored spectrally transformed linear mixed models (FaST-LMM) demonstrated the most superior efficiency, yielding more results with the minimal false positives. We discovered that PUS7, FBXO11, FOXN2, MSH6, and SLC4A4 were associated with BWB. RAG2, and TMEFF2 were candidate genes for BW1, and STARD13, Klotho, ZAR1L are likely candidates for BW3 and BW5. PLXNC1, ATP1A1, CD58, FRYL, OCIAD1, and OCIAD2 were linked to BW7. These findings provide a genetic reference for the selection and breeding of Jinling White ducks, while also deepened our understanding of Growth and development phenotypic in ducks.

Analysis of carcass traits, meat quality, amino acid and fatty acid profiles between different duck lines.

To investigate the effect of genetic selection on meat quality in ducks, twenty of each fast growth ducks (LCA) and slow growth ducks (LCC) selected from F6 generation of Cherry Valley ducks (♂) x Liancheng white ducks (♀) were analyzed for carcass characteristics, meat quality (physicochemical and textural characteristics), amino acid and fatty acid profiles at 7 wk. Results showed that live body weight, slaughter weight, eviscerated yield and abdominal fat percentage of LCA were significantly higher than those in LCC ducks (P < 0.01). Moreover, the average area and diameter of myofiber were larger in LCA than LCC ducks (P < 0.01). The breast and thigh muscles of LCA exhibited significantly lower water holding capacity and thermal loss compared with LCC ducks (P < 0.01). In addition, the content of nonessential amino acids (Glu, Asp, and Arg) in breast muscles and Asp, Ser, Thr, and Met in thigh muscles was higher in LCC than LCA ducks (P < 0.05). The proportion of polyunsaturated fatty acids (PUFA) in breast muscles of LCC was higher than LCA ducks (P < 0.05). However, the content of saturated fatty acids (SFA) in breast and thigh muscles of LCA was higher compared with LCC ducks (P < 0.05). The proportion of monounsaturated fatty acids (MUFA) in thigh muscles was significantly higher in LCC compared with LCA ducks (P < 0.01). Finally, multiple traits were evaluated by applying principal component analysis (PCA) and the results indicated that PUFA and SFA in breast muscles of LCA played important roles in meat quality, followed by Warner-Bratzler shear force (WBSF) and MUFA. However, water holding capacity (WHC) had a dominant effect in meat quality of thigh muscles in both LCA and LCC ducks.

Identification of relevant differential genes to the divergent development of pectoral muscle in ducks by transcriptomic analysis.

OBJECTIVE: The objective of this study was to identify candidate genes that play important roles in skeletal muscle development in ducks. METHODS: In this study, we investigated the transcriptional sequencing of embryonic pectoral muscles from two specialized lines: Liancheng white ducks (female) and Cherry valley ducks (male) hybrid Line A (LCA) and Line C (LCC) ducks. In addition, prediction of target genes for the differentially expressed mRNAs was conducted and the enriched gene ontology (GO) terms and Kyoto encyclopedia of genes and genomes signaling pathways were further analyzed. Finally, a protein-to-protein interaction network was analyzed by using the target genes to gain insights into their potential functional association. RESULTS: A total of 1,428 differentially expressed genes (DEGs) with 762 being up-regulated genes and 666 being down-regulated genes in pectoral muscle of LCA and LCC ducks identified by RNA-seq (p<0.05). Meanwhile, 23 GO terms in the down-regulated genes and 75 GO terms in up-regulated genes were significantly enriched (p<0.05). Furthermore, the top 5 most enriched pathways were ECM-receptor interaction, fatty acid degradation, pyruvate degradation, PPAR signaling pathway, and glycolysis/gluconeogenesis. Finally, the candidate genes including integrin b3 (Itgb3), pyruvate kinase M1/2 (Pkm), insulinlike growth factor 1 (Igf1), glucose-6-phosphate isomerase (Gpi), GABA type A receptorassociated protein-like 1 (Gabarapl1), and thyroid hormone receptor beta (Thrb) showed the most expression difference, and then were selected to verification by quantitative realtime polymerase chain reaction (qRT-PCR). The result of qRT-PCR was consistent with that of transcriptome sequencing. CONCLUSION: This study provided information of molecular mechanisms underlying the developmental differences in skeletal muscles between specialized duck lines.

Transcriptome Profiling Identifies Differentially Expressed Genes in Skeletal Muscle Development in Native Chinese Ducks.

China boasts a rich diversity of indigenous duck species, some of which exhibit desirable economic traits. Here, we generated transcriptome sequencing datasets of breast muscle tissue samples from 1D of four groups: Pekin duck pure breeding group (P), Jinling White duck breeding group (J), P ♂ × J ♀ orthogonal group (PJ) and J ♂ × P ♀ reciprocal-cross group (JP) (n = 3), chosen based on the distinctive characteristics of duck muscle development during the embryonic period. We identified 5053 differentially expressed genes (DEGs) among the four groups. Network prediction analysis showed that ribosome and oxidative phosphorylation-related genes were the most enriched, and muscular protein-related genes were found in the 14-day-old embryonic group. We found that previously characterized functional genes, such as FN1, AGRN, ADNAMST3, APOB and FGF9, were potentially involved in muscle development in 14-day-old embryos. Functional enrichment analysis suggested that genes that participated in molecular function and cell component and key signaling pathways (e.g., hippo, ribosome, oxidative phosphorylation) were significantly enriched in the development of skeletal muscle at 14 days of embryonic age. These results indicate a possible role of muscle metabolism and myoglobin synthesis in skeletal muscle development in both duck parents and hybrids.

Metastasis-associated gene, mag-1 improves tumour microenvironmental adaptation and potentiates tumour metastasis.

Metastasis is a major cause of death from malignant diseases, and the underlying mechanisms are still largely not known. A detailed probe into the factors which may regulate tumour invasion and metastasis contributes to novel anti-metastatic therapies. We previously identified a novel metastasis-associated gene 1 (mag-1) by means of metastatic phenotype cloning. Then we characterized the gene expression profile of mag-1 and showed that it promoted cell migration, adhesion and invasion in vitro. Importantly, the disruption of mag-1 via RNA interference not only inhibited cellular metastatic behaviours but also significantly reduced tumour weight and restrained mouse breast cancer cells to metastasize to lungs in spontaneous metastatic assay in vivo. Furthermore, we proved that mag-1 integrates dual regulating mechanisms through the stabilization of HIF-1α and the activation of mTOR signalling pathway. We also found that mag-1-induced metastatic promotion could be abrogated by mTOR specific inhibitor, rapamycin. Taken together, the findings identified a direct role that mag-1 played in metastasis and implicated its function in cellular adaptation to tumour microenvironment.

Evaluating cell migration in vitro by the method based on cell patterning within microfluidic channels.

Cell migration is an early-stage and critical step for cancer metastasis. The most common approach to monitor this process is wound-healing assay. However, this traditional method has some unavoidable limitations. We observed that simply scratching the monolayer of cultured cells might cause local cell damage around the injury line. The cells along the scratched border seemed to be irritated and exhibited abnormal distribution of cytoskeleton reassembly with protruding "cell islands" and "pseudopodia" during wound healing, which might potentially affect the assessment of cell migration behavior. Herein, we applied a microfluidic device that mechanically constrained cells seeded in a designed pattern inside microchannels, and monitored cell movement in a way of mimicking the natural microenvironment of cancerous tissues. We illustrated the capacity of this simple method to probe cellular migration behaviors and to screen some biological active agents that reflected in their influence on cellular motility.

Ectopic expression γ-glutamylcysteine synthetase of Vicia sativa increased cadmium tolerance in Arabidopsis.

Glutathione (GSH) is a multifunctional essential biothiol, and its metabolism is important for plant against toxic metals and metalloids. γ-Glutamylcysteine (γ-EC), which is catalyzed by γ-Glutamylcysteine synthetase (γ-ECS), is a rate-limiting intermediate in GSH synthesis. Here, a γ-ECS gene (Vsγ-ECS) from Vicia sativa was cloned, and its function in modulating Cd tolerance was studied. Vsγ-ECS is a chloroplast localization protein, and the expression of Vsγ-ECS was upregulated by Cd stress in root of V. sativa. Heterologous expression of Vsγ-ECS (35S::Vsγ-ECS) in Arabidopsis enhanced the Cd tolerance of plants through improved primary root length, fresh weight, chlorophyll content and low degree of oxidation associated with reduced H2O2 and lipid peroxidation. However, the Cd accumulation of Arabidopsis had no effect on Vsγ-ECS overexpression. Further analysis showed that the increased Cd tolerance in 35S::Vsγ-ECS was mainly due to the capacity of increasing GSH synthesis that improved Cd chelation by GSH and phytochelatins (PCs) and alleviated the oxidative stress caused by Cd stress. In summary, a γ-ECS was characterized from V. sativa, and it demonstrated a property for increasing GSH and PC synthesis to protect plants from Cd poisoning.

Ascorbic acid and all-trans retinoic acid promote proliferation of chicken blastoderm cells (cBCs) by mediating DNA demethylation.

Chicken blastoderm cells (cBCs) obtained from stage X (EG&K) embryos are easily available materials for the study of cell development. However, cBCs are not widely used because they are hard to maintain in long-term culture in vitro. To solve this problem, ascorbic acid (AA; also known as vitamin C (VC)) and all-trans retinoic acid (ATRA) were added into basic culture medium to promote cell growth. Results suggested that cultured cBCs possessed strongly proliferative activity and maintained their pluripotency on the support of chicken embryonic fibroblast (CEF) feeder. Moreover, when VC or/and ATRA was added, the number and area of cBC colonies increased significantly compared with the control group. The expression of pluripotency genes (Sox2 and Nanog) and cell cycle-regulated genes (CCND1 and CDK6) was upregulated obviously. Furthermore, results showed that 5hmC levels in VC and RA groups increased significantly by DNA dot blot and immunofluorescence staining. These results provide strong evidence that VC and ATRA induced DNA demethylation and enhanced 5hmC level. The level of H3K27me3 was raised, while the level of H3K9me2 was reduced by addition of VC and ATRA. Finally, the expression of Tet1 and Dnmt3b was upregulated remarkably. Therefore, these results indicated that VC and ATRA enhanced DNA demethylation and then promoted cBC survival and proliferation in vitro.

Novel gene rearrangement in the mitochondrial genome of Anastatus fulloi (Hymenoptera Chalcidoidea) and phylogenetic implications for Chalcidoidea.

The genus Anastatus comprises a large group of parasitoids, including several biological control agents in agricultural and forest systems. The taxonomy and phylogeny of these species remain controversial. In this study, the mitogenome of A. fulloi Sheng and Wang was sequenced and characterized. The nearly full-length mitogenome of A. fulloi was 15,692 bp, compromising 13 protein-coding genes (PCGs), 2 rRNA genes, 22 tRNA genes and a control region (CR). The total A + T contents were 83.83%, 82.18%, 87.58%, 87.27%, and 82.13% in the whole mitogenome, 13 PCGs, 22 tRNA genes, 2 rRNA genes, and CR, respectively. The mitogenome presented negative AT skews and positive GC skews, except for the CR. Most PCGs were encoded on the heavy strand, started with ATN codons, and ended with TAA codons. Among the 3736 amino acid-encoding codons, TTA (Leu1), CGA (Arg), TCA (Ser2), and TCT (Ser2) were predominant. Most tRNAs had cloverleaf secondary structures, except trnS1, with the absence of a dihydrouridine (DHU) arm. Compared with mitogenomes of the ancestral insect and another parasitoid within Eupelmidae, large-scale rearrangements were found in the mitogenome of A. fulloi, especially inversions and inverse transpositions of tRNA genes. The gene arrangements of parasitoid mitogenomes within Chalcidoidea were variable. A novel gene arrangement was presented in the mitogenome of A. fulloi. Phylogenetic analyses based on the 13 protein-coding genes of 20 parasitoids indicated that the phylogenetic relationship of 6 superfamilies could be presented as Mymaridae + (Eupelmidae + (Encyrtidae + (Trichogrammatidae + (Pteromalidae + Eulophidae)))). This study presents the first mitogenome of the Anastatus genus and offers insights into the identification, taxonomy, and phylogeny of these parasitoids.

Impact of pesticide/fertilizer mixtures on the rhizosphere microbial community of field-grown sugarcane.

The rhizosphere microbial community is important for plant health and is shaped by numerous environmental factors. This study aimed to unravel the effects of a pesticide/fertilizer mixture on the soil rhizosphere microbiome of field-grown sugarcane. A field trial on sugarcane was conducted in Zhanjian City, Guangdong Province, China, and soil samples from the rhizosphere were collected after clothianidin pesticide and/or organic fertilizer treatments. The effects of pesticide and/or organic fertilizer treatments on the composition, diversity, and predictive function of the rhizosphere microbial communities were examined using 16S rRNA gene and ITS1 amplicon sequencing. Compared with the controls (no pesticide or fertilizer used), the microbial community that resulted from treatment with the pesticide/fertilizer mixture (SPF) had a higher relative bacterial diversity and fungal richness, and contributed more beneficial functions to sugarcane, including xenobiotics biodegradation and metabolism of amino acids. The bacterial and fungal compositions at various taxonomic levels were not significantly different in SPF and SP (pesticide only) treatments compared to treatments without the pesticide, suggesting that the clothianidin addition did not cause a detrimental impact on the soil microbiome. Moreover, five bacterial genera, including Dyella, Sphingomonas, Catenulispora, Mucilaginibacter, and Tumebacillus, were significantly more abundant in the SPF and SP treatments, which could be associated with the pesticide addition. With the addition of organic fertilizers in SPF, the abundances of some soil-beneficial bacteria Bacillus, Paenibacillus, and Brevibacillus were highly increased. Our study provides insights into the interactions between the rhizosphere soil microbiome and pesticide-fertilizer integration, which may help improve the application of pesticide-fertilizer to sugarcane fields. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02770-3.

Transcriptome analysis of yellow passion fruit in response to cucumber mosaic virus infection.

The cultivation and production of passion fruit (Passiflora edulis) are severely affected by viral disease. Yet there have been few studies of the molecular response of passion fruit to virus attack. In the present study, RNA-based transcriptional profiling (RNA-seq) was used to identify the gene expression profiles in yellow passion fruit (Passiflora edulis f. flavicarpa) leaves following inoculation with cucumber mosaic virus (CMV). Six RNA-seq libraries were constructed comprising a total of 42.23 Gb clean data. 1,545 differentially expressed genes (DEGs) were obtained (701 upregulated and 884 downregulated). Gene annotation analyses revealed that genes associated with plant hormone signal transduction, transcription factors, protein ubiquitination, detoxification, phenylpropanoid biosynthesis, photosynthesis and chlorophyll metabolism were significantly affected by CMV infection. The represented genes activated by CMV infection corresponded to transcription factors WRKY family, NAC family, protein ubiquitination and peroxidase. Several DEGs encoding protein TIFY, pathogenesis-related proteins, and RNA-dependent RNA polymerases also were upregualted by CMV infection. Overall, the information obtained in this study enriched the resources available for research into the molecular-genetic mechanisms of the passion fruit/CMV interaction, and might provide a theoretical basis for the prevention and management of passion fruit viral disease in the field.