Exploring the influence of a single-nucleotide mutation in EIN4 on tomato fruit firmness diversity through fruit pericarp microstructure.

Tomato (Solanum lycopersicum) stands as one of the most valuable vegetable crops globally, and fruit firmness significantly impacts storage and transportation. To identify genes governing tomato firmness, we scrutinized the firmness of 266 accessions from core collections. Our study pinpointed an ethylene receptor gene, SlEIN4, located on chromosome 4 through a genome-wide association study (GWAS) of fruit firmness in the 266 tomato core accessions. A single-nucleotide polymorphism (SNP) (A → G) of SlEIN4 distinguished lower (AA) and higher (GG) fruit firmness genotypes. Through experiments, we observed that overexpression of SlEIN4AA significantly delayed tomato fruit ripening and dramatically reduced fruit firmness at the red ripe stage compared with the control. Conversely, gene editing of SlEIN4AA with CRISPR/Cas9 notably accelerated fruit ripening and significantly increased fruit firmness at the red ripe stage compared with the control. Further investigations revealed that fruit firmness is associated with alterations in the microstructure of the fruit pericarp. Additionally, SlEIN4AA positively regulates pectinase activity. The transient transformation assay verified that the SNP (A → G) on SlEIN4 caused different genetic effects, as overexpression of SlEIN4GG increased fruit firmness. Moreover, SlEIN4 exerts a negative regulatory role in tomato ripening by impacting ethylene evolution through the abundant expression of ethylene pathway regulatory genes. This study presents the first evidence of the role of ethylene receptor genes in regulating fruit firmness. These significant findings will facilitate the effective utilization of firmness and ripening traits in tomato improvement, offering promising opportunities for enhancing tomato storage and transportation capabilities.

Comparative transcriptome analysis of R3a and Avr3a-mediated defense responses in transgenic tomato.

Late blight caused by Phytophthora infestans is one of the most devastating diseases in potatoes and tomatoes. At present, several late blight resistance genes have been mapped and cloned. To better understand the transcriptome changes during the incompatible interaction process between R3a and Avr3a, in this study, after spraying DEX, the leaves of MM-R3a-Avr3a and MM-Avr3a transgenic plants at different time points were used for comparative transcriptome analysis. A total of 7,324 repeated DEGs were detected in MM-R3a-Avr3a plants at 2-h and 6-h, and 729 genes were differentially expressed at 6-h compared with 2-h. Only 1,319 repeated DEGs were found in MM-Avr3a at 2-h and 6-h, of which 330 genes have the same expression pattern. Based on GO, KEGG and WCGNA analysis of DEGs, the phenylpropanoid biosynthesis, plant-pathogen interaction, and plant hormone signal transduction pathways were significantly up-regulated. Parts of the down-regulated DEGs were enriched in carbon metabolism and the photosynthesis process. Among these DEGs, most of the transcription factors, such as WRKY, MYB, and NAC, related to disease resistance or endogenous hormones SA and ET pathways, as well as PR, CML, SGT1 gene were also significantly induced. Our results provide transcriptome-wide insights into R3a and Avr3a-mediated incompatibility interaction.

The HD-Zip IV transcription factor SlHDZIV8 controls multicellular trichome morphology by regulating the expression of Hairless-2.

Trichomes are specialized epidermal appendages that serve as excellent models to study cell morphogenesis. Although the molecular mechanism underlying trichome morphogenesis in Arabidopsis has been well characterized, most of the regulators essential for multicellular trichome morphology remain unknown in tomato. In this study, we determined that the recessive hairless-2 (hl-2) mutation in tomato causes severe distortion of all trichome types, along with increased stem fragility. Using map-based cloning, we found that the hl-2 phenotype was associated with a 100 bp insertion in the coding region of Nck-associated protein 1, a component of the SCAR/WAVE complex. Direct protein-protein interaction was detected between Hl-2 and Hl (SRA1, specifically Rac1-associated protein) using yeast two-hybrid and co-immunoprecipitation assays, suggesting that these proteins may work together during trichome formation. In addition, knock-down of a HD-Zip IV transcription factor, HDZIPIV8, distorted trichomes similar to the hl-2 mutant. HDZIPIV8 regulates the expression of Hl-2 by binding to the L1-box in the Hl-2 promoter region, and is involved in organizing actin filaments. The brittleness of hl-2 stems was found to result from decreased cellulose content. Taken together, these findings suggest that the Hl-2 gene plays an important role in controlling multicellular trichome morphogenesis and mechanical properties of stems in tomato plants.

Genome-wide identification, genomic organization and expression profiles of SlARR-B gene family in tomato.

The type-B authentic response regulators (ARR-Bs) function as positive regulators of cytokinin signal transduction and play important roles in abiotic stress resistance and plant development. However, little of ARR-B family is known in tomato. In this study, we performed a comprehensive analysis of ARR-B family factors in tomato. In total, 12 genes encoding ARR-B transcription factors (named as SlARR-B1-SlARR-B12) were identified from tomato. We analyzed the structures, chromosome locations, phylogeny, protein motifs, and expression profiles of these SlARR-B genes. Gene structure analysis showed that 5-12 exons and 4-11 introns existed in the SlARR-B genes. These SlARR-B genes were asymmetrically distributed on eight chromosomes in tomato. Phylogenetic tree of SlARR-B genes from tomato and other plant species revealed that SlARR-B genes were classified into 6 subfamilies. SlARR-B proteins had typical conserved domains, including Motif 1 and Motif 2. The investigation of the expression profiles of SlARR-B genes in all the examined tissues demonstrated that these genes were differentially expressed, including roots, stems, leaves, flowers, and fruits at developmental stages. Notably, the expression of SlARR-B11 and SlARR-B12 exhibited high expression levels in flowers. Each gene was induced by at least one of different phytohormones (SA, IAA, ABA, IBA, 6-BA, JA, GA, and ETH) and four abiotic stress treatments (heat, drought, salt, and cold). This study sets a good foundation for further characterization of the SlARR-B transcription factors in plant development and abiotic stress responses of tomato.

The tomato B-type cyclin gene, SlCycB2, plays key roles in reproductive organ development, trichome initiation, terpenoids biosynthesis and Prodenia litura defense.

Cyclins exist extensively in various plant species. Among them, B-type cyclins play important roles in the transition of G2-to-M. However, few B-type cyclins have been reported to participate in reproductive organ development and trichome formation. In this study, transgene analysis showed that SlCycB2 overexpression caused abnormal flower with the unclosed stamen, shortened style and aberrant pollen. In addition, nearly all non-glandular trichomes, as well as the glandular ones were disappeared. On the contrary, suppression of SlCycB2 could promote type III and type V trichomes formation. Detection of secondary metabolites indicated that the production of monoterpene and sesquiterpene were significantly decreased in SlCycB2-OE plants, which thus resulted in the reduction of the defense against Prodenia litura. Transcriptome profile demonstrated that the differentially expressed genes mainly participate in the biosynthesis of terpenes, cutin, suberine and wax. Furthermore, we identified several homologs of SlCycB2, SlCycB3, NtCycB2, AtCycB2, which have similar regulatory functions in trichome formation. These results indicate that SlCycB2 plays a critical role in reproductive organ development, multicellular trichome initiation, secondary metabolite biosynthesis and Prodenia litura defense in tomato. The similar roles of its homologs in multicellular trichome formation suggest that Solanaceous species may share common regulatory pathway.

Transcriptome profile analysis of cell proliferation molecular processes during multicellular trichome formation induced by tomato Wo (v) gene in tobacco.

Trichomes, developing from the epidermis of nearly all terrestrial plants, provide good protection from environmental stress. Regulation of trichomes in Rosids has been well characterized. However, little is known about the cell proliferation molecular processes during multicellular trichome formation in Asterids. Ectopic expression of Wo (v) in tobacco and potato induces much more trichome formation than wild type. To gain new insights into the underlying mechanisms during the processes of these trichomes formation, RNA-seq was employed for the young primary leaf tissues in Wo (v) transgenic and wild-type tobacco. We identified differentially expressed genes which are related to various biological processes and molecular functions. Here, we provide details of experimental methods, RNA-seq data (available at Gene Expression Omnibus database under GSE72310). Our data provide new insight into the molecular processes controlling multicellular formation in tobacco.

Transcriptome profile analysis of cell proliferation molecular processes during multicellular trichome formation induced by tomato Wov gene in tobacco.

BACKGROUND: Trichomes, developing from the epidermis of nearly all terrestrial plants, provide good structural resistance against insect herbivores and an excellent model for studying the molecular mechanisms underlying cell fate determination. Regulation of trichomes in Rosids has been well characterized. However, little is known about the cell proliferation molecular processes during multicellular trichome formation in Asterids. RESULTS: In this study, we identified two point mutations in a novel allele (Wov) at Wo locus. Ectopic expression of Wov in tobacco and potato induces much more trichome formation than wild type. To gain new insights into the underlying mechanisms during the processes of these trichomes formation, we compared the gene expression profiles between Wov transgenic and wild-type tobacco by RNA-seq analysis. A total of 544 co-DEGs were detected between transgenic and wild-type tobacco. Functional assignments of the co-DEGs indicated that 33 reliable pathways are altered in transgenic tobacco plants. The most noticeable pathways are fatty acid metabolism, amino acid biosynthesis and metabolism, and plant hormone signal transduction. Results suggest that these enhanced processes are critical for the cell proliferation during multicellular trichome formation in transgenic plants. In addition, the transcriptional levels of homologues of trichome regulators in Rosids were not significantly changed, whereas homologues of genes (Wo and SlCycB2) in Asterids were significantly upregulated in Wov transgenic tobacco plants. CONCLUSIONS: This study presents a global picture of the gene expression changes induced by Wov-gene in tobacco. And the results provided us new insight into the molecular processes controlling multicellular formation in tobacco. Furthermore, we inferred that trichomes in solanaceous species might share a common network.

Comprehensive analysis and expression profile of the homeodomain leucine zipper IV transcription factor family in tomato.

Homeodomain leucine zipper IV (HD-ZIP IV) proteins are plant-specific transcription factors that play important roles in development of epidermal cell layers and cuticle formation. The functions of two HD-ZIP IV family genes, CD2 and Wo, have been well characterized in tomato (Solanum lycopersicum). CD2 and Wo are involved in cuticle biosynthesis and trichome formation, respectively. In this study, we identified 13 novel tomato HD-ZIP IV (SlHDZIV) genes. We analyzed the structures, chromosome locations, phylogeny, protein motifs, and expression profiles of these SlHDZIV genes. Gene structure analysis revealed that a module of 11 exons and 10 introns existed in the SlHDZIV genes. These genes were asymmetrically distributed on chromosomes, except on chromosome 4 and 5. Segmental duplication possibly contributed to the expansion of tomato HD-ZIP IV genes. The expression profiles of these genes revealed their broad expression pattern and high expression in young leaves and flowers. Each gene responded to more than one of different phytohormones [abscisic acid, ethephon, 4-(indolyl)-butyric acid, jasmonic acid, salicylic acid, gibberellic acid, and 6-benzylaminopurine] and four abiotic stress treatments (cold, heat, salt, and drought). This study provided significant insights into the diverse roles of SlHDZIV genes in tomato growth and development.

Polygalasaponin F inhibits secretion of inflammatory cytokines via NF-κB pathway regulation.

To study the anti-neuroinflammatory mechanisms of polygalasaponin F (PS-F), ELISA method was used to detect the secretion of inflammatory cytokines. Western blot was used to detect the protein expression and phosphorylation levels. Immunofluorescence assay was used to observe the NF-κB nuclear translocation. PS-F could inhibit the release of inflammatory cytokines TNF-α and NO induced by lipopolysaccharides (LPS) and reduce the expression of inducible nitric oxide synthases (iNOS). As for MAPK-signaling pathway, PS-F could only inhibit the phosphorylation levels of p38 MAPK, but did not significantly affect the phosphorylation levels of JNK and ERK1/2 protein kinases. PS-F could inhibit NF-κB nuclear translocation in a dose-dependent manner. The results of Western blot assay were consistent with immunofluorescence assays. Meanwhile, p38-specific inhibitor SB203580 (20 µM) and p65-specific inhibitor PDTC (100 µM) were, respectively, administered as a positive control. In addition, PS-F could significantly inhibit the cytotoxicity of conditioned medium prepared by LPS-stimulated BV-2 microglia (LPS conditioned media) to neuronal PC12 cells and improve cell viability. PS-F inhibits the secretions of neuroinflammatory cytokines by the regulation of NF-κB-signaling pathway.

Overexpression of human E46K mutant α-synuclein impairs macroautophagy via inactivation of JNK1-Bcl-2 pathway.

Parkinson's disease (PD) is pathologically characterized by selective loss of dopaminergic neurons in the midbrain and the existence of intracellular protein inclusions termed Lewy bodies, largely composed of α-synuclein. Genetic studies have revealed that rare point mutations in the gene encoding α-synuclein including A30P, A53T, and E46K are associated with familial forms of PD, indicating a pathological role for mutant α-synuclein in PD etiology. However, the mechanisms underlying the neuronal toxicity of mutant α-synuclein are still to be elucidated. Growing evidence has suggested a deleterious effect of mutant α-synuclein on the autophagy-lysosome pathway. In this study, we discovered that overexpression of human E46K mutant α-synuclein impaired macroautophagy in mammalian cells. Our data showed that overexpression of E46K mutant α-synuclein impaired autophagy at an early stage of autophagosome formation via the c-Jun N-terminal kinase 1 (JNK1)-Bcl-2 but not the mammalian target of rapamycin (mTOR) pathway. Overexpressed E46K mutant α-synuclein inhibited JNK1 activation, leading to a reduced Bcl-2 phosphorylation and increased association between Bcl-2 and Beclin1, further disrupting the formation of Beclin1/hVps34 complex, which is essential for autophagy initiation. Furthermore, overexpression of E46K mutant α-synuclein increased the vulnerability of differentiated PC12 cells to rotenone treatment, which would be partly due to its inhibitory effects on autophagy. Our findings may shed light on the potential roles of mutant α-synuclein in the pathogenesis of PD.

Perceptions of nursing profession and learning experiences of male students in baccalaureate nursing program in Changsha, China.

Using Colaizzi's framework, this qualitative study explored perceptions of the nursing profession and learning experiences of male students in a baccalaureate nursing program in Changsha, China. Data were collected through in-depth interviews on 14 subjects and analyzed with software Nvivo 8.0. Six theme clusters emerged as the following: (1) entering nursing field, (2) perceptions of nursing profession, (3) difficulties in studying nursing, (4) inner feelings, (5) impact of being a nursing student, and (6) career plans. The experiences and perceptions of nursing and studying nursing were mainly negative, revealing issues stemming from the method of student recruitment for the baccalaureate nursing program, gender bias in nursing teaching, and social views on nursing work. In addition, psychological pressure on male nursing students is significant factor and should not be ignored. The implications for nurse educators are outlined, with suggestions to facilitate the recruitment and retention of more male nursing students.