The present study investigated the effect of silibinin, the principal potential anti-inflammatory flavonoid contained in silymarin, a mixture of flavonolignans extracted from Silybum marianum seeds, on palmitate-induced insulin resistance in C2C12 myotubes and its potential molecular mechanisms. Silibinin prevented the decrease of insulin-stimulated 2-NBDG (2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-D-glucose) uptake and the downregulation of glutamate transporter type 4 (GLUT4) translocation in C2C12 myotubes induced by palmitate. Meanwhile, silibinin suppressed the palmitate-induced decrease of insulin-stimulated Akt Ser473 phosphorylation, which was reversed by wortmannin, a specific inhibitor of phosphatidylinositol-3-kinase (PI3K). We also found that palmitate downregulated insulin-stimulated Tyr632 phosphorylation of insulin receptor substrate 1 (IRS-1) and up-regulated IRS-1 Ser307 phosphorylation. These effects were rebalanced by silibinin. Considering several serine/threonine kinases reported to phosphorylate IRS-1 at Ser307, treatment with silibinin downregulated the phosphorylation of both c-Jun N-terminal kinase (JNK) and nuclear factor-κB kinase β (IKKβ), which was increased by palmitate in C2C12 myotubes mediating inflammatory status, whereas the phosphorylation of PKC-θ was not significantly modulated by silibinin. Collectively, the results indicated that silibinin prevented inhibition of the IRS-1/PI3K/Akt pathway, thus ameliorating palmitate-induced insulin resistance in C2C12 myotubes.

Background The basic leucine zipper (bZIP) transcription factor (TF) is one of the largest families of transcription factors (TFs). It is widely distributed and highly conserved in animals, plants, and microorganisms. Previous studies have shown that the bZIP TF family is involved in plant growth, development, and stress responses. The bZIP family has been studied in many plants; however, there is little research on the bZIP gene family in tobacco. Results In this study, 77 bZIPs were identified in tobacco and named NtbZIP01 through to NtbZIP77. These 77 genes were then divided into eleven subfamilies according to their homology with Arabidopsis thaliana. NtbZIPs were unevenly distributed across twenty-two tobacco chromosomes, and we found sixteen pairs of segmental duplication. We further studied the collinearity between these genes and related genes of six other species. Quantitative real-time polymerase chain reaction analysis identified that expression patterns of bZIPs differed, including in different organs and under various abiotic stresses. NtbZIP49 might be important in the development of flowers and fruits; NtbZIP18 might be an important regulator in abiotic stress. Conclusions In this study, the structures and functions of the bZIP family in tobacco were systematically explored. Many bZIPs may play vital roles in the regulation of organ development, growth, and responses to abiotic stresses. This research has great significance for the functional characterisation of the tobacco bZIP family and our understanding of the bZIP family in higher plants.

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Background The filamentous temperature-sensitive H protease (ftsH) gene family plays an important role in plant growth and development. FtsH proteins belong to the AAA protease family. Studies have shown that it is a key gene for plant chloroplast development and photosynthesis regulation. In addition, the ftsH gene is also involved in plant response to stress. At present, the research and analysis of the ftsH gene family are conducted in microorganisms such as Escherichia coli and Oenococcus and various plants such as Arabidopsis, pear, rice, and corn. However, analysis reports on ftsH genes from tobacco (Nicotiana tabacum L.), an important model plant, are still lacking. Since ftsH genes regulate plant growth and development, it has become necessary to systematically study this gene in an economically important plant like tobacco. Results This is the first study to analyze the ftsH gene from Nicotiana tabacum L. K326 (NtftsH). We identified 20 ftsH genes from the whole genome sequence, renamed them according to their chromosomal locations, and divided them into eight subfamilies. These 20 NtftsH genes were unevenly distributed across the 24 chromosomes. We found four pairs of fragment duplications. We further investigated the collinearity between these genes and related genes in five other species. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis identified differential expression patterns of NtftsH in different tissues and under various abiotic stress conditions. Conclusions This study provides a comprehensive analysis of the NtftsH gene family. The exon–intron structure and motif composition are highly similar in NtftsH genes that belong to the same evolutionary tree branch. Homology analysis and phylogenetic comparison of ftsH genes from several different plants provide valuable clues for studying the evolutionary characteristics of NtftsH genes. The NtftsH genes play important roles in plant growth and development, revealed by their expression levels in different tissues as well as under different stress conditions. Gene expression and phylogenetic analyses will provide the basis for the functional analysis of NtftsH genes. These results provide a valuable resource for a better understanding of the biological role of the ftsH genes in the tobacco plant.

Heterosis is a common biological phenomenon that can be used to optimize yield and quality of crops. Using heterosis breeding, hybrids with suitable nicotine content have been applied to tobacco leaf production. However, the molecular mechanism of the formation of nicotine heterosis has never been explained from the perspective of protein. The DIA proteomics technique was used to compare the differential proteomics of the hybrid Va116 × Basma, showing strong heterosis in nicotine content from its parent lines Va116 and Basma. Proteomics analysis indicated that 65.2% of DEPs showed over-dominant expression patterns, and these DEPs included QS, BBL, GS, ARAF and RFC1 which related to nicotine synthesis. In addition, some DEPs (including GST, ABCE2 and ABCF1 and SLY1) that may be associated with nicotinic transport exhibited significant heterosis over the parental lines. These findings demonstrated that the efficiency of the synthesis and transport of nicotine in hybrids was significantly higher than that in the parent lines, and the accumulation of over-dominant expression proteins may be the cause of heterosis of nicotinic content in hybrids.

Potassium (K+) is essential for crop growth. Increasing the K+ content can often directly promote the improvement of crop yield and quality. Heterosis plays an important role in genetic improvement and leads to genetic gains. We found that the K+ content of tobacco showed significant heterosis, which is highly significant for cultivating tobacco varieties with high K+ content. However, the mechanism by which K+ content heterosis occurs in tobacco leaves is not clear. In this study, a comprehensive comparative transcriptome sequencing analysis of root samples from the hybrid G70 × GDH11 and its parental inbred lines G70 and GDH11 was performed to elucidate the importance of the root uptake capacity of K+ in the formation of heterosis. The results showed that 29.53% and 60.49% of the differentially expressed genes (DEGs) exhibited dominant and over-dominant expression patterns, respectively. These non-additive upregulated DEGs were significantly enriched in GO terms, such as metal ion transport and reaction, ion balance and homeostasis, ion channel activity, root meristem growth, and regulation of root hairs. The KEGG annotation results indicated that these genes were mainly involved in the pathways such as energy metabolism, carbohydrate formation, amino acid metabolism, and signal transduction. Further analysis showed that probable potassium transporter 17 (NtKT17) and potassium transporter 5-like (NtKT5), associated with potassium ion absorption, glutamate receptor 2.2-like and glutamate receptor 2.8-like, associated with ion channel activity, LOC107782957, protein detoxification 42-like, and probable glutamate carboxypeptidase 2, associated with root configuration, showed a significantly higher expression in the hybrids. These results indicated that the over-dominant expression pattern of DEGs played a key role in the heterosis of K+ content in tobacco leaves, and the overexpression of the genes related to K+ uptake, transport, and root development in hybrids helped to improve the K+ content of plants, thus showing the phenomenon of heterosis.

Nicotine is a unique alkaloid present in tobacco that is widely used in cigarettes and in the agricultural, chemical, and pharmaceutical industries. However, the research on nicotine is mostly limited to its synthesis pathways, and only a few studies have explored the effects of other metabolic pathways on nicotine precursors. Regulating the nicotine content in tobacco can greatly promoting the application of nicotine in other fields. In this study, we performed global data-independent acquisition proteomics analysis of four tobacco varieties. Of the four varieties, one had high nicotine content and three had a low nicotine content. A total of 31,259 distinct peptides and 6,018 proteins across two samples were identified. A total of 45 differentially expressed proteins (DEPs) co-existed in the three comparison groups and were mainly involved in the transport and metallic processes of the substances. Most DEPs were enriched in the biosynthesis of secondary metals, glutathione metabolism, carbon metabolism, and glycolysis/gluconeogenesis. In addition, the weighted gene co-expression network analysis identified an expression module closely related to the nicotine content (Brown, r = 0.74, P = 0.006). Gene Ontology annotation and Kyoto Encyclopaedia of Genes and Genomes enrichment analysis showed that the module proteins were mainly involved in the synthesis and metabolism of nicotine precursors such as arginine, ornithine aspartate, proline, and glutathione. The increased levels of these precursors lead to the synthesis and accumulation of nicotine in plants. More importantly, these proteins regulate nicotine synthesis by affecting the formation of putrescine, which is the core intermediate product in nicotine anabolism. Our results provide a reference for tobacco variety selection with a suitable nicotine content and regulation of the nicotine content. Additionally, the results highlight the importance of other precursor metabolism in nicotine synthesis.

Background Potassium(K+) plays a vital role in improving the quality of tobacco leaves. However, how to improve the potassium content of tobacco leaves has always been a difficult problem in tobacco planting. K+ content in tobacco hybrid is characterized by heterosis, which can improve the quality of tobacco leaves, but its underlying molecular genetic mechanisms remain unclear. Results Through a two-year field experiment, G70×GDH11 with strong heterosis and K326×GDH11 with weak heterosis were screened out. Transcriptome analyses revealed that 80.89% and 57.28% of the differentially expressed genes (DEGs) in the strong and weak heterosis combinations exhibited an overdominant expression pattern, respectively. The genes that up-regulated the overdominant expression in the strong heterosis hybrids were significantly enriched in the ion homeostasis. Genes involved in K+ transport (KAT1/2, GORK, AKT2, and KEA3), activity regulation complex (CBL-CIPK5/6), and vacuole (TPKs) genes were overdominant expressed in strong heterosis hybrids, which contributed to K+ homeostasis and heterosis in tobacco leaves. Conclusions K+ homeostasis and accumulation in tobacco hybrids were collectively improved. The overdominant expression of K+ transport and homeostasis-related genes conducted a crucial role in the heterosis of K+ content in tobacco leaves.