Genome-Wide Identification and Expression Analysis of Bx Involved in Benzoxazinoids Biosynthesis Revealed the Roles of DIMBOA during Early Somatic Embryogenesis in Dimocarpus longan Lour.

Benzoxazinoids (BXs) are tryptophan-derived indole metabolites and play a role in various physiological processes, such as auxin metabolism. Auxin is essential in the process of somatic embryogenesis (SE) in plants. In this study, we used bioinformatics, transcriptome data, exogenous treatment experiments, and qPCR analysis to study the evolutionary pattern of Bx genes in green plants, the regulatory mechanism of DlBx genes during early SE, and the effect of 2,4-dihydroxy-7-methoxy-1,4-benzoxazine-3-one (DIMBOA) on the early SE in Dimocarpus longan Lour. The results showed that 27 putative DlBxs were identified in the longan genome; the Bx genes evolved independently in monocots and dicots, and the main way of gene duplication for the DlBx was tandem duplication (TD) and the DlBx were strongly constrained by purification selection during evolution. The transcriptome data indicated varying expression levels of DlBx during longan early SE, and most DlBxs responded to light, temperature, drought stress, and 2,4-dichlorophenoxyacetic acid (2,4-D) treatment; qRT-PCR results showed DlBx1, DlBx6g and DlBx6h were responsive to auxin, and treatment with 0.1mg/L DIMBOA for 9 days significantly upregulated the expression levels of DlBx1, DlBx3g, DlBx6c, DlBx6f, DlB6h, DlBx7d, DlBx8, and DlBx9b. The correlation analysis showed a significantly negative correlation between the expression level of DlBx1 and the endogenous IAA contents; DIMBOA significantly promoted the early SE and significantly changed the endogenous IAA content, and the IAA content increased significantly at the 9th day and decreased significantly at the 13th day. Therefore, the results suggested that DIMBOA indirectly promote the early SE by changing the endogenous IAA content via affecting the expression level of DlBx1 and hydrogen peroxide (H2O2) content in longan.

Genome-Wide Identification and Expression Analysis of DWARF53 Gene in Response to GA and SL Related to Plant Height in Banana.

Dwarfing is one of the common phenotypic variations in asexually reproduced progeny of banana, and dwarfed banana is not only windproof and anti-fallout but also effective in increasing acreage yield. As a key gene in the strigolactone signalling pathway, DWARF53 (D53) plays an important role in the regulation of the height of plants. In order to gain insight into the function of the banana D53 gene, this study conducted genome-wide identification of banana D53 gene based on the M. acuminata, M. balbisiana and M. itinerans genome database. Analysis of MaD53 gene expression under high temperature, low temperature and osmotic stress based on transcriptome data and RT-qPCR was used to analyse MaD53 gene expression in different tissues as well as in different concentrations of GA and SL treatments. In this study, we identified three MaD53, three MbD53 and two MiD53 genes in banana. Phylogenetic tree analysis showed that D53 Musa are equally related to D53 Asparagales and Poales. Both high and low-temperature stresses substantially reduced the expression of the MaD53 gene, but osmotic stress treatments had less effect on the expression of the MaD53 gene. GR24 treatment did not significantly promote the height of the banana, but the expression of the MaD53 gene was significantly reduced in roots and leaves. GA treatment at 100 mg/L significantly promoted the expression of the MaD53 gene in roots, but the expression of this gene was significantly reduced in leaves. In this study, we concluded that MaD53 responds to GA and SL treatments, but "Yinniaijiao" dwarf banana may not be sensitive to GA and SL.

[Identification of banana ADA1 gene family members and their expression profiles under biotic and abiotic stresses].

The Spt-Ada-Gcn5-acetyltransferase (SAGA) is an ancillary transcription initiation complex which is highly conserved. The ADA1 (alteration/deficiency in activation 1, also called histone H2A functional interactor 1, HFI1) is a subunit in the core module of the SAGA protein complex. ADA1 plays an important role in plant growth and development as well as stress resistance. In this paper, we performed genome-wide identification of banana ADA1 gene family members based on banana genomic data, and analyzed the basic physicochemical properties, evolutionary relationships, selection pressure, promoter cis-acting elements, and its expression profiles under biotic and abiotic stresses. The results showed that there were 10, 6, and 7 family members in Musa acuminata, Musa balbisiana and Musa itinerans. The members were all unstable and hydrophilic proteins, and only contained the conservative SAGA-Tad1 domain. Both MaADA1 and MbADA1 have interactive relationship with Sgf11 (SAGA-associated factor 11) of core module in SAGA. Phylogenetic analysis revealed that banana ADA1 gene family members could be divided into 3 classes. The evolution of ADA1 gene family members was mostly influenced by purifying selection. There were large differences among the gene structure of banana ADA1 gene family members. ADA1 gene family members contained plenty of hormonal elements. MaADA1-1 may play a prominent role in the resistance of banana to cold stress, while MaADA1 may respond to the Panama disease of banana. In conclusion, this study suggested ADA1 gene family members are highly conserved in banana, and may respond to biotic and abiotic stress.

Transcriptome analysis provides insight into the regulatory mechanisms underlying pollen germination recovery at normal high ambient temperature in wild banana (Musa itinerans).

Introduction: Cultivated banana are polyploid, with low pollen fertility, and most cultivars are male sterile, which leads to difficulties in banana breeding research. The selection of male parent with excellent resistance and pollen fertility is therefore essential for banana breeding. Wild banana (Musa itinerans) have developed many good characteristics during natural selection and constitute an excellent gene pool for breeding. Therefore, research on wild banana breeding is very important for banana breeding. Results: In the current analysis, we examined the changes in viability of wild banana pollens at different temperatures by in vitro germination, and found that the germination ability of wild banana pollens cultured at 28°C for 2 days was higher than that of pollens cultured at 23°C (pollens that could not germinate normally under low temperature stress), 24°C (cultured at a constant temperature for 2 days) and 32°C (cultured at a constant temperature for 2 days). To elucidate the molecular mechanisms underlying the germination restoration process in wild banana pollens, we selected the wild banana pollens that had lost its germination ability under low temperature stress (23°C) as the control group (CK) and the wild banana pollens that had recovered its germination ability under constant temperature incubation of 28°C for 2 days as the treatment group (T) for transcriptome sequencing. A total of 921 differentially expressed genes (DEGs) were detected in CK vs T, of which 265 were up-regulated and 656 were down-regulated. The combined analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that the activation, metabolism of various substances (lipids, sugars, amino acids) play a major role in restoring pollen germination capacity. TCA cycle and the sesquiterpenoid and triterpenoid biosynthetic pathways were also significantly enriched in the KEGG pathway. And we found that some DEGs may be associated with pollen wall formation, DNA methylation and DNA repair. The cysteine content, free fatty acid (FFA) content, H2O2 content, fructose content, and sucrose content of pollen were increased at treatment of 28°C, while D-Golactose content was decreased. Finally, the GO pathway was enriched for a total of 24 DEGs related to pollen germination, of which 16 DEGs received targeted regulation by 14 MYBs. Discussions: Our study suggests that the balance between various metabolic processes, pollen wall remodelling, DNA methylation, DNA repairs and regulation of MYBs are essential for germination of wild banana pollens.

Trace elements in aquatic products from Shenzhen, China and their implications for human exposure.

Aquatic organisms in industrially polluted areas can accumulate large quantities of heavy metals. To assess the resulting health risks, 11 trace elements in 184 aquatic products representing 14 species of fish, crustaceans, and bivalves collected from Shenzhen, China were determined. Aluminum (Al), chromium (Cr), nickel (Ni), selenium (Se), antimony (Sb), manganese (Mn), copper (Cu), arsenic (As), cadmium (Cd), mercury (Hg), and lead (Pb) were determined by inductively coupled plasma mass spectrometry. The pollution levels of each product and the human health risk resulting from their consumption were then assessed. The concentrations of As in 57 % of samples and Cd in 11 % of samples exceeded the upper limits stipulated by the Chinese National Food Safety Standards (GB 2762-2017), which was mainly due to high concentrations of trace elements in crustaceans and bivalves. The Nemerow integrated pollution index indicated that the aquatic products accumulated high levels of As and Cd. Health risk assessments using the target hazard quotient (THQ) and hazard index (HI) suggested that As and Cd exposure due to consumption of aquatic products presents a potential health risk for residents of Shenzhen.

Riboflavin mediates m6A modification targeted by miR408, promoting early somatic embryogenesis in longan.

Plant somatic embryogenesis (SE) is an in vitro biological process wherein bipolar structures are induced to form somatic cells and regenerate into whole plants. MicroRNA (miRNA) is an essential player in plant SE. However, the mechanism of microRNA408 (miR408) in SE remains elusive. Here, we used stable transgenic technology in longan (Dimocarpus longan) embryogenic calli to verify the mechanism by which miR408 promotes cell division and differentiation of longan early SE. dlo-miR408-3p regulated riboflavin biosynthesis by targeting nudix hydrolase 23 (DlNUDT23), a previously unidentified gene mediating N6-methyladenosine (m6A) modification and influencing RNA homeostasis and cell cycle gene expression during longan early SE. We showed that DlMIR408 overexpression (DlMIR408-OE) promoted 21-nt miRNA biosynthesis. In DlMIR408-OE cell lines, dlo-miR408-3p targeted and downregulated DlNUDT23, promoted riboflavin biosynthesis, decreased flavin mononucleotide (FMN) accumulation, promoted m6A level, and influenced miRNA homeostasis. DNA replication, glycosylphosphatidylinositol (GPI)-anchor biosynthesis, the pentose phosphate pathway, and taurine and hypotaurine metabolism were also closely associated with riboflavin metabolism. In a riboflavin feeding assay, dlo-miR408-3p and pre-miR408 were upregulated and DlNUDT23 was downregulated, increasing the m6A level and cell division and differentiation in longan globular embryos. When riboflavin biosynthesis was inhibited, dlo-miR408-3p was downregulated and DlNUDT23 was upregulated, which decreased m6A modification and inhibited cell division but did not inhibit cell differentiation. FMN artificial demethylated m6A modification affected the homeostasis of precursor miRNA and miRNA. Our results revealed a mechanism underlying dlo-miR408-3p-activated riboflavin biosynthesis in which DlNUDT23 is targeted, m6A modification is dynamically mediated, and cell division is affected, promoting early SE in plants.

Genome-wide identification, evolution analysis of LysM gene family members and their expression analysis in response to biotic and abiotic stresses in banana (Musa L.).

LysM (Lysin motif), in response to pathogenic molecular stresses, is a crucial signal recognition gene. To understand the molecular characteristics of banana LysM gene family members, we used a series of bioinformatics methods. Based on the genomic databases of Musa acuminata, Musa balbisiana and Musa itinerans, a total of 53 genes and 55 proteins were identified, with 21 genes and 23 proteins in the M.acuminata, 16 genes and 16 proteins in each of M.balbisiana and M.itinerans, respectively. According to the conserved structural domains, LysM can be divided into five classes, namely LysM&MltD, LYK, LYP, LysMn, and LysMe. The LysM gene was relatively highly conserved in the evolution of the three genomes of banana, and some differences occurred. Expression analysis revealed that MaLysM4-5 was relatively highly expressed under high-temperature stress, low-temperature stress and pathogen infection; at the same time, about one-third of the members were down-regulated under low-temperature stress and high-temperature stress, while the expression of MaLysM10-1 and MaLysM4-5 were up-regulated. After the banana wilt fungus FocTR4 infected the banana roots, MaLysM1 was down-regulated and MaLysM11-1 was up-regulated. In conclusion, our study suggests that MaLysMs may be necessary in the response to high- and low-temperature stresses, as well as the banana wilt fungus infestation. Overall, this paper found that LysM genes may be involved in biotic and abiotic stresses in banana, and provided helpful information about LysM's evolution, expression and properties, which will provide theoretical references for further studies on the functions of LysM genes and resistance breeding in the future.

Genome-wide identification, evolution analysis of cytochrome P450 monooxygenase multigene family and their expression patterns during the early somatic embryogenesis in Dimocarpus longan Lour.

Cytochrome P450 (CYP), a multi-gene superfamily, is involved in a broad range of physiological processes, including hormone responses and secondary metabolism throughout the plant life cycle. Longan (Dimocarpus longan), a subtropical and tropical evergreen fruit tree, its embryonic development is closely related to the yield and quality of fruits. And a large number of secondary metabolites, such as flavonoids and carotenoids, are also produced during the longan somatic embryogenesis (SE). It is important, therefore, to study potential functions of CYPs in longan. However, the knowledge of longan CYPs is still very limited. Here, a total of 327 DlCYPs were identified using the genome-search method, which could be classified into nine clans. The expansion of the DlCYP family was mainly caused by tandem duplication (TD) events. Promoter cis-acting elements analysis elucidated that DlCYPs played important roles in hormonal responses. A total of 246 DlCYPs exhibited six different expression patterns during the early SE based on longan transcriptomic data. Eight DlCYPs underwent alternative splicing (AS) events, and they might produce one to six isoforms. And the AS transcript of DlCYP97C1 might act as an alternative to the full-length transcript in ICpEC and GE stages. Finally, protein-protein interaction (PPI) networks and miRNA target prediction elucidated that DlCYPs might be involved in the phenylpropanoid metabolic pathway and primarily regulated and targeted by miR413. In summary, our results provided valuable inventory for understanding the classification and biological functions of DlCYPs and provided insight into further functional verification of DlCYPs during the longan early SE.

Genome-wide identification, expression and functional analysis of the core cell cycle gene family during the early somatic embryogenesis of Dimocarpus longan Lour.

Core cell cycle genes (CCCs) are essential regulators of cell cycle operation. In this study, a total of 69 CCCs family members, including 37 CYCs, 20 CDKs, five E2F/DPs, three KRPs, two RBs, one CKS and one Wee1, were identified from the longan genome. Phylogenetic and motifs analysis showed the evolutionary conservation of CCCs. Transcriptome dataset showed that CCCs had various expression patterns during longan early somatic embryogenesis (SE). Either CKS or CYCD3;2 silencing increased the expression of RB-E2F pathway genes, and the silencing of CYCD3;2 might induce the process of apoptosis in longan embryogenic callus (EC) cells. In addition, The qRT-PCR results showed that the expression levels of CDKG2, CYCD3;2, CYCT1;2, CKS and KRP1 were elevated by ABA, 2,4-D and PEG4000 treatments, while CDKG2 and CYCT1;2 were inhibited by NaCl treatment. In conclusion, our study provided valuable information for understanding the characterization and biological functions of longan CCCs.

PAs Regulate Early Somatic Embryo Development by Changing the Gene Expression Level and the Hormonal Balance in Dimocarpus longan Lour.

Polyamines (PAs) play an important regulatory role in many basic cellular processes and physiological and biochemical processes. However, there are few studies on the identification of PA biosynthesis and metabolism family members and the role of PAs in the transition of plant embryogenic calli (EC) into globular embryos (GE), especially in perennial woody plants. We identified 20 genes involved in PA biosynthesis and metabolism from the third-generation genome of longan (Dimocarpus longan Lour.). There were no significant differences between longan and other species regarding the number of members, and they had high similarity with Citrus sinensis. Light, plant hormones and a variety of stress cis-acting elements were found in these family members. The biosynthesis and metabolism of PAs in longan were mainly completed by DlADC2, DlSAMDC2, DlSAMDC3, DlSPDS1A, DlSPMS, DlCuAOB, DlCuAO3A, DlPAO2 and DlPAO4B. In addition, 0.01 mmol∙L-1 1-aminocyclopropane-1-carboxylic acid (ACC), putrescine (Put) and spermine (Spm), could promote the transformation of EC into GE, and Spm treatment had the best effect, while 0.01 mmol∙L-1 D-arginine (D-arg) treatment inhibited the process. The period between the 9th and 11th days was key for the transformation of EC into GE in longan. There were higher levels of gibberellin (GA), salicylic acid (SA) and abscisic acid (ABA) and lower levels of indole-3-acetic acid (IAA), ethylene and hydrogen peroxide (H2O2) in this key period. The expression levels in this period of DlADC2, DlODC, DlSPDS1A, DlCuAOB and DlPAO4B were upregulated, while those of DlSAMDC2 and DlSPMS were downregulated. These results showed that the exogenous ACC, D-arg and PAs could regulate the transformation of EC into GE in longan by changing the content of endogenous hormones and the expression levels of PA biosynthesis and metabolism genes. This study provided a foundation for further determining the physicochemical properties and molecular evolution characteristics of the PA biosynthesis and metabolism gene families, and explored the mechanism of PAs and ethylene for regulating the transformation of plant EC into GE.

Assessment of macular microvasculature features before and after vitrectomy in the idiopathic macular epiretinal membrane using a grading system: An optical coherence tomography angiography study.

PURPOSE: To evaluate pre- and postoperative microvasculature features in eyes with different idiopathic macular epiretinal membrane (iERM) classifications using optical coherence tomography angiography (OCTA). METHODS: In this retrospective study, 100 eyes with iERM were enrolled; 62 eyes underwent pars plana vitrectomy (PPV). All iERM eyes were evaluated and graded using optical coherence tomography (OCT). According to the thickness of the fovea relative to the surrounding macula from OCT radial line scans, we classified iERM into three grades. Optical coherence tomography angiography (OCTA) was used to measure the foveal avascular zone (FAZ)-related parameters and the superficial and deep capillary plexus layers using 3 × 3 mm scans. Measurements were taken at baseline and 3 months postoperatively. Best corrected visual acuity (BCVA), vessel density (VD), FAZ area, FAZ perimeter (PERIM), acircularity index (AI), and foveal vessel density (FD) were evaluated. RESULTS: Idiopathic macular epiretinal membrane (iERM) eyes with a higher grade had a lower FAZ area and perimeter (p < 0.0001), higher foveal vessel density (FVD) both in the superficial capillary plexus (SCP) (p < 0.0001) and in the deep capillary plexus (DCP) (p < 0.05), and a lower parafoveal vessel density (PRVD) in the DCP (p < 0.0001). The macular vessel density ratio (MVR = FVD/PRVD) increased with an increase in grade both in the SCP and in the DCP (p < 0.0001). For grade 1 iERM eyes, only PRVD in the DCP significantly changed before versus after the operation (p < 0.05). For grade 2 iERM eyes, the FAZ area and perimeter became larger after the operation (p < 0.05). The MVR of grade 2 iERM eyes decreased postoperatively both in the SCP (p < 0.05) and in the DCP (p < 0.001). For grade 1 and grade 2 iERM eyes, preoperative LogMAR BCVA was negatively correlated with the FAZ area (p < 0.01) and perimeter (p < 0.01), and was positively correlated with the MVR in the SCP (p < 0.05). Postoperative LogMAR BCVA was positively correlated with the FVD in the DCP (p < 0.05). CONCLUSIONS: Idiopathic macular epiretinal membrane (iERM) eyes of different grades have significant differences in microvasculature features. According to OCTA, eyes with higher grades have more serious microvascular changes. Grading plays a part in predicting postoperative microvascular characteristics, and grade 1 iERM eyes have a better visual outcome compared with grade 2 iERM eyes.

Quality suitability regionalization analysis of Angelica sinensis in Gansu, China.

The genus Angelica encompasses 80 species worldwide. Among them, only Angelica sinensis is widely used in China and Japan. To explore the quality and geographical distribution of A. sinensis, we collected 1,530 plants from Gansu Province and analyzed them for their contents of chlorogenic acid (CA), ferulic acid (FA), senkyunolide I(SI), senkyunolide A(SA), senkyunolide H (SH), coniferyl ferulate (CF), ligustilide (LI), and butenyl phthalide (BP) using UPLC. We also assessed the relationship between the ecological environment and quality of A. sinensis through maximum entropy modeling and a geographical information system. The habitat suitability distribution demonstrated that the most influential ecological factors for the growth of A. sinensis were altitude, precipitation during March, May, and December, precipitation during the wettest month, and the soil pH. The most suitable areas for cultivation are concentrated to the south of Gansu Province, including Linxia Hui Autonomous Prefecture, Dingxi City, Tianshui City, south of Wuwei City, east of Gannan Tibetan Autonomous Prefecture, north of Longnan City, and northwest of Pingliang City. The quality suitability regionalization analysis divulged that the most influential ecological factors for the index components of A. sinensis were the altitude, sunshine, rainfall, temperature, and soil pH. The highest quality A. sinensis grow in Dingxi City, Tangchang, Lixian, and Wen counties in Longnan City, Wushan County in Tianshui City, Lintan, Zhouqu, and Zhuoni counties in Gannan Tibetan Autonomous Prefecture, Kangle and Linxia counties in Linxia Hui Autonomous Prefecture. The experiment yielded highly accurate results (accuracy of 0.955), suggesting that the results were consistent with the actual distribution of A. sinensis in Gansu. The inferences of this research will naturally draw the attention of the authorities in the fields of natural resources and agriculture departments and provide a scientific basis for the rational selection of A. sinensis cultivation areas.

Measurement and Analysis of Central Corneal Thickness at Different Postnatal Stages in Chinese Premature Infants.

PURPOSE: The objective of this study was to evaluate central corneal thickness (CCT) in Chinese premature infants at different postnatal stages to study the peak point and analyze influential factors on CCT development. METHODS: This was a cross-sectional study of premature infants. Initial CCT measurement was taken at 34 weeks of gestational age (GA) and at intervals until 88 weeks of postmenstrual age (PMA) was reached. The comparison and correlation analysis were carried out to access the association of CCT with gender, birth weight (BW), GA, and retinopathy of prematurity (ROP) for each PMA. The premature infants were divided into the thick CCT group and the thin CCT group according to the average CCT at 40 w. And the difference in CCT between the two groups at subsequent 52 w and 64 w was compared. RESULTS: A total of 1726 premature infants (3463 measurements) with an average of 2.21 ± 1.57 measurements were included in this study. The CCT decreased from 34 w GA to 52 w PMA (R = 92.36, P < 0.0001) and then reached a plateau (R = 2.541, P=0.3567). Male (P < 0.05), low BW (P < 0.05), and low GA (P < 0.05) were associated with thicker CCT at the early stage of PMA. The premature infants who had thick CCT at 40 w would have thick CCT at 52 w and 64 w accordingly. CONCLUSIONS: The CCT values of premature infants decreased over time and plateaued at 52 w PMA. Gender, BW, and GA were considered as the influential factors of CCT at the early stage of PMA. Moreover, CCT at 40 w could forecast its development trend at 52 w or 64 w after birth.

Protective effect of extract of Astragalus on learning and memory impairments and neurons' apoptosis induced by glucocorticoids in 12-month-old male mice.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder marked by a progressive loss of memory and cognitive function. Stress-level glucocorticoids are correlated with dementia progression in patients with AD. In this study, 12-month male mice were chronically treated with stress-level dexamethasone (DEX, 5 mg/kg) and extract of Astragalus (EA, 10, 20, and 40 mg/kg) or Ginsenoside Rg1 (Rg1, 6.5 mg/kg) for 21 days. We investigated the protective effect of EA against DEX injury in mice and its action mechanism. Our results indicate that DEX can induce learning and memory impairments and neuronal cell apoptosis. The mRNA levels of caspase-3 are selectively increased after DEX administration. The results of immunohistochemistry demonstrate that caspase-3 and cytochrome c in hippocampus (CA1, CA3) and neocortex are significantly increased. Furthermore, DEX treatment increased the activity of caspase-9 and caspase-3. Treatment groups with EA (20 and 40 mg/kg) or Rg1 (6.5 mg/kg) significantly improve learning and memory, downregulate the mRNA level of caspase-3, decrease expression of caspase-3 and cytochrome c in hippocampus (CA1, CA3) and neocortex, and inhibit activity of caspase-9 and caspase-3. The present findings highlight a possible mechanism by which stress level of DEX accelerates learning and memory impairments and increases neuronal apoptosis and the potential neuronal protection of EA.