Combined analysis of transcriptome and WGCNA reveals the mediating role of JA in the low male fertility of loquat H30-6

The number of seeds is an important factor limiting the palatability and processing efficiency of loquat. Loquat H30-6 has been found to bear fruits with few seeds, which is attributed to its low male fertility. To investigate the molecular mechanism underlying the low male fertility of H30-6, in this study, RNA-seq was performed using pre-meiotic, meiotic and mature anthers from H30-6 and the other two loquat varieties with normal male fertility. The results indicated that a total of 331 differentially expressed genes (DEGs) were identified between H30-6 and the other two loquats and these DEGs were mainly enriched in pathways related to jasmonic acid (JA). WGCNA revealed that module ‘green’ were highly positively correlated with seed quantity (r = 0.46), suggesting that module ‘green’ is the candidate module. Enrichment analysis showed that genes in module ‘green’ were also primarily enriched in JA biosynthetic process, regulation of JA mediated signaling pathway and response to JA, implying that JA may play a key role in the molecular mechanism underlying the low male fertility of H30-6. The detection of UPLC-MS/MS indicated that the contents of JA and its derivatives in the flower bud of H30-6 were significantly lower than those in ‘Huabai 1’ (with normal male fertility). Besides, exogenous application of MeJA improved the pollen quantity per anther and seed numbers of H30-6. Our work indicates that the suppression of JA-related gene expression and the low JA content in immature flower buds may be key factors for H30-6 male sterility. These findings provide a theoretical basis for seedless breeding of loquats and lay the foundation for the development of stamens in fruit trees

Multi-scale modeling and mechanical performance analysis of finger seals with plain woven C/C composite

Abstract The application of carbon fibers reinforced carbon matrix (C/C) composite can solve the local wear of metallic finger seals effectively. However, the performance of C/C composite finger seal is complex and variable, which further decreases the sealing performance and life. Therefore, a method of multi-scale modeling and mechanical performance analysis for plain woven C/C composite finger seals was conducted. The circumferential finger beams of C/C composite were modeled by multi-scale structural analysis and weaving simulation. The radial static and dynamic stiffness characteristics of finger beams were investigated. The results showed that the radial static stiffness of the finger beam with three layers was about 3 times that with single layer. The radial stiffness of circumferential finger beams presented a periodic distribution pattern with a period of 90°. The radial dynamic stiffness of C/C composite finger beams increased with the excitation displacement amplitude and rotor speed. But the magnitude and fluctuation degree of dynamic stiffness were greater than those of static stiffness. A large difference in radial stiffness will lead to local wear and hysteretic leakage. This study lays a foundation for the analysis and optimization of the hysteresis and wear characteristics of C/C composite finger seals

The Putative Role of the NAC Transcription Factor EjNACL47 in Cell Enlargement of Loquat (Eriobotrya japonica Lindl.)

NAC transcription factors (TFs) are plant-specific TFs that play essential roles in plant development; however, the function of NAC TFs in loquat development remains unknown. The natural triploid loquat (Eriobotrya japonica Lindl.), Longquan No.1. B355, has larger organs than its corresponding diploid loquat (B2). Here, we cloned an NAC-like TF (EjNACL47 (NAC-like 47)) from the cDNA of triploid loquat B355 flowers. EjNACL47 has a conserved domain of NAC TFs and is homologous to AtNAC47. Transient expression in tobacco leaves revealed that EjNACL47 localized to the nucleus, and yeast-two-hybrid screening confirmed that the C-terminus displayed transcriptional activity. Interestingly, real-time qRT-PCR indicated that the expression levels of EjNACL47 in leaves and flower organs in triploid loquat (B355) were higher than those in diploid loquat (B2), implying that EjNACL47 might be associated with the larger organ size in B355. Moreover, Arabidopsis lines ectopically expressing EjNACL47 presented obviously larger leaves, flowers, and siliques than the wild-type variant, suggesting that EjNACL47 plays a positive role in Arabidopsis organ enlargement. These results offer insight into the molecular mechanism of NAC TFs involved in regulating organ size in loquat.</jats:p

The Putative Role of the NAC Transcription Factor EjNACL47 in Cell Enlargement of Loquat (Eriobotrya japonica Lindl.)

NAC transcription factors (TFs) are plant-specific TFs that play essential roles in plant development; however, the function of NAC TFs in loquat development remains unknown. The natural triploid loquat (Eriobotrya japonica Lindl.), Longquan No.1. B355, has larger organs than its corresponding diploid loquat (B2). Here, we cloned an NAC-like TF (EjNACL47 (NAC-like 47)) from the cDNA of triploid loquat B355 flowers. EjNACL47 has a conserved domain of NAC TFs and is homologous to AtNAC47. Transient expression in tobacco leaves revealed that EjNACL47 localized to the nucleus, and yeast-two-hybrid screening confirmed that the C-terminus displayed transcriptional activity. Interestingly, real-time qRT-PCR indicated that the expression levels of EjNACL47 in leaves and flower organs in triploid loquat (B355) were higher than those in diploid loquat (B2), implying that EjNACL47 might be associated with the larger organ size in B355. Moreover, Arabidopsis lines ectopically expressing EjNACL47 presented obviously larger leaves, flowers, and siliques than the wild-type variant, suggesting that EjNACL47 plays a positive role in Arabidopsis organ enlargement. These results offer insight into the molecular mechanism of NAC TFs involved in regulating organ size in loquat

An Integrative Analysis of Transcriptome, Proteome and Hormones Reveals Key Differentially Expressed Genes and Metabolic Pathways Involved in Flower Development in Loquat

Flower development is a vital developmental process in the life cycle of woody perennials, especially fruit trees. Herein, we used transcriptomic, proteomic, and hormone analyses to investigate the key candidate genes/proteins in loquat (Eriobotrya japonica) at the stages of flower bud differentiation (FBD), floral bud elongation (FBE), and floral anthesis (FA). Comparative transcriptome analysis showed that differentially expressed genes (DEGs) were mainly enriched in metabolic pathways of hormone signal transduction and starch and sucrose metabolism. Importantly, the DEGs of hormone signal transduction were significantly involved in the signaling pathways of auxin, gibberellins (GAs), cytokinin, ethylene, abscisic acid (ABA), jasmonic acid, and salicylic acid. Meanwhile, key floral integrator genes FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (SOC1) and floral meristem identity genes SQUAMOSA PROMOTER BINDING LIKE (SPL), LEAFY (LFY), APETALA1 (AP1), and AP2 were significantly upregulated at the FBD stage. However, key floral organ identity genes AGAMOUS (AG), AP3, and PISTILLATA (PI) were significantly upregulated at the stages of FBE and FA. Furthermore, transcription factors (TFs) such as bHLH (basic helix-loop-helix), NAC (no apical meristem (NAM), Arabidopsis transcription activation factor (ATAF1/2) and cup-shaped cotyledon (CUC2)), MYB_related (myeloblastosis_related), ERF (ethylene response factor), and C2H2 (cysteine-2/histidine-2) were also significantly differentially expressed. Accordingly, comparative proteomic analysis of differentially accumulated proteins (DAPs) and combined enrichment of DEGs and DAPs showed that starch and sucrose metabolism was also significantly enriched. Concentrations of GA3 and zeatin were high before the FA stage, but ABA concentration remained high at the FA stage. Our results provide abundant sequence resources for clarifying the underlying mechanisms of the flower development in loquat.</jats:p

An Efficient System for Mesophyll Protoplast Isolation, Purification, and Transformation in Loquat: Studies on Fluorescent Marker Analysis and Subcellular Localization

Loquat (Eriobotrya japonica Lindl.) is one of the most important subtropical evergreen fruit trees. However, due to the lack of widely applicable genetic transformation platforms, the research about gene functional characterization and molecular mechanisms is largely confined. In this study, the efficient protocol of protoplast isolation (the enzyme solution composed of 2.4% macerozyme R-10, 4.8% cellulase RS, dissolved in a 0.6 M mannitol solution) and the method of protoplast purification (CPW solution containing 5% sucrose and 11% mannitol) have been achieved with protoplast yields of 12.6 &times; 106/g&middot;FW, reaching a viability rate of up to 91%. A protoplast transient gene expression system has been established with an efficiency of approximately 40% using GFP reporter gene. Using this reliable and efficient system, the protein localization characteristics of transcription factor EjDELLA, EjbHLH79, and marker gene OsPHT4 were also utilized for further analysis. To our knowledge, this is the first report on establishing an efficient system for protoplast isolation, purification, and transformation of loquat mesophyll. The system reported here will definitely promote rapid progress in breeding, genetic transformation, and molecular research