A novel role of the cotton calcium sensor CBL3 was involved in Verticillium wilt resistance in cotton.

BACKGROUND: Verticillium wilt, causes mainly by the soilborne pathogen Verticillium dahliae, is a devastated vascular disease resulting in huge financial losses in cotton, so research on improving V. dahliae stress tolerance in cotton is the utmost importance. Calcium as the second messenger acts as a crucial role in plant innate immunity. Cytosolic Ca2+during the pathogen infection is a significant increase in plant immune responses. Calcineurin B-like (CBL) proteins are widely known calcium sensors that regulate abiotic stress responses. However, the role of cotton CBLs in response to V. dahliae stress remains unclear. OBJECTIVE: To discover and utilize the gene to Verticillium wilt resistance and defense response mechanism of cotton. METHODS: Through screening the gene to Verticillium wilt resistance in cotton, four GhCBL3 copies were obtained from the current common cotton genome sequences. The protein domain and phylogenetic analyses of GhCBL3 were performed using NCBI Blast, DNAMAN, and MotifScan programs. Real-time RT-PCR was used to detect the expression of GhCBL3 gene in cotton seedlings under various stress treatments. The expression construct including GhCBL3 cDNA was transduced into Agrobacterium tumefaciens (GV3101) by heat shock method and transformed into cotton plants by Virus-Induced Gene Silencing (VIGS) method. The results of silencing of GhCBl3 on ROS accumulation and plant disease resistance in cotton plants were assessed. RESULTS: A member of calcineurin B-like proteins (defined as GhCBL3) in cotton was obtained. The expression of GhCBL3 was significantly induced and raised by various stressors, including dahliae, jasmonic acid (JA) and H2O2 stresses. Knockdown GhCBL3 in cotton by Virus-Induced Gene Silencing analysis enhanced Verticillium wilt tolerance and changed the occurrence of reactive oxygen species. Some disease-resistant genes were increased in GhCBL3-silencing cotton lines. CONCLUSION: GhCBL3 may function on regulating the Verticillium dahliae stress response of plants.

Ca2+-responsive phospholipid-binding BONZAI genes confer a novel role for cotton resistance to Verticillium wilt.

Verticillium wilt which produced by the soil-borne fungus Verticillium dahliae is an important biotic threat that limits cotton (Gossypium hirsutum) growth and agricultural productivity. It is very essential to explore new genes for the generation of V. dahliae resistance or tolerance cotton varieties. Ca2+ signaling as a secondary messenger is involved in pathogen stress response. Despite Ca2+-responsive phospholipid-binding BONZAI (BON) genes have intensively been investigated in Arabidopsis, their function has not still been characterized in cotton. Here, we showed that three copies of GhBON1, two copies of GhBON2 and GhBON3 were found from the genome sequences of upland cotton. The expression of GhBON1 was inducible to V. dahliae. Knocking down of GhBON1, GhBON2 and GhBON3 using virus induced gene silencing (VIGS) each increased up-regulation of defense responses in cotton. These GhBON1, GhBON2 and GhBON3-silenced plants enhanced resistance to V. dahliae accompanied by higher burst of hydrogen peroxide and decreased cell death and had more effect on the up-regulation of defense response genes. Further analysis revealed that GhBON1 could interacts with BAK1-interacting receptor-like kinase 1 (GhBIR1) and pathogen-associated molecular pattern (PAMP) receptor regulator BAK1 (GhBAK1) at plasma membrane. Our study further reveals that plant Ca2+ -responsive phospholipid-binding BONZAI genes negatively regulate Verticillium wilt with the conserved function in response to disease resistance or plant immunity.

SsPsaH, a H subunit of the photosystem I reaction center of Suaeda salsa, confers the capacity of osmotic adjustment in tobacco.

BACKGROUND: Abiotic stress effects agricultural production, so research on improving stress tolerance of crop is important. Suaeda salsa is a halophyte with high salt and drought tolerance and ability to desalinate saline soil and improve soil quality. OBJECTIVE: To discover and utilize of salt and drought tolerance-related genes, we further investigated the mechanisms of salt and drought tolerance. METHODS: Through screening a salt treated Suaeda salsa cDNA library and further cloning a H subunit of the photosystem I reaction center SsPsaH cDNA, and then the protein domain and phylogenetic analyses of PSI genes was conducted with the NCBI Blast, DNAMAN, and MotifScan programs. The S. salsa seedlings were subjected to various stress treatments and analyze expression of SsPsaH under these treatments by real-time RT-PCR. SsPsaH expression construct was introduced into S. pombe cells by electroporation and transformed into N. tabacum plants by the leaf disc transformation method. RESULTS: A member of the H subunit of the Photosystem I reaction center (defined as SsPsaH) was obtained. The expression of SsPsaH was up-regulated by abscisic acid (ABA), salt, and drought stress treatments. Over-expressing SsPsaH in recombinant yeasts enhanced high salinity tolerance and increased tolerance to sorbitol during seed germination and seedling root development in tobacco, respectively. Some stress-related mark genes such as a LEA family gene of NtLEA, a binding protein of a drought response element of NtDREB, the ascorbate peroxidase gene (NtAPX) were also up-regulated in SsPsaH overexpressing transgenic tobacco lines. CONCLUSIONS: These results show that SsPsaH may contribute to the salt and osmotic stress response of plants.

Proteomic profiling of cotton fiber developmental transition from cell elongation to secondary wall deposition.

Cotton fiber developmental transition from elongation to secondary cell wall biosynthesis is a critical growth shifting phase that affects fiber final length, strength, and other properties. Morphological dynamic analysis indicated an asynchronous fiber developmental pattern between two most important commercial cotton species, Gossypium hirsutum (Gh) and G. barbadense (Gb). Using isobaric tags for relative and absolute quantitation techniques, we examined the temporal changes of protein expression at three representative development periods (15-19, 19-23, and 23-27 dpa) in both species. Strikingly, a large proportion of differentially expressed proteins (DEPs) were identified at 19-23 dpa in Gh and at 23-27 dpa in Gb, corresponding to their fiber developmental transition timing. To better understand fiber transitional development, we comparatively analyzed those DEPs in 19-23 dpa of Gh vs. in 23-27 dpa of Gb, and noted that these cotton species indeed share fundamentally similar fiber developmental features under the biological processes. We also showed that there are limited overlaps in both specific upregulated and downregulated proteins between the two species, suggesting species-specific protein regulations in the development process. Proteomic profiling results revealed dynamic changes of several key proteins and biological processes that are potentially correlated with fiber developmental transition. During the transition, upregulated proteins are mainly involved in carbohydrate/energy metabolism, oxidation-reduction, cytoskeleton, protein turnover, Ca2+ signaling, etc., whereas important downregulated proteins are mostly involved in phenylpropanoid and flavonoid secondary metabolism pathways. The gene expressions of several changed proteins in this key stage were also examined by quantitative reverse transcription polymerase chain reaction. Overall, the present study provides accurate pictures of the regulatory networks of functional proteins during the fiber developmental transition, therefore highlighting candidate genes/proteins and related pathways for the cotton fiber improvement.

Genome-Wide Characterization and Expression Profiles of the Superoxide Dismutase Gene Family in Gossypium.

Superoxide dismutase (SOD) as a group of significant and ubiquitous enzymes plays a critical function in plant growth and development. Previously this gene family has been investigated in Arabidopsis and rice; it has not yet been characterized in cotton. In our study, it was the first time for us to perform a genome-wide analysis of SOD gene family in cotton. Our results showed that 10 genes of SOD gene family were identified in Gossypium arboreum and Gossypium raimondii, including 6 Cu-Zn-SODs, 2 Fe-SODs, and 2 Mn-SODs. The chromosomal distribution analysis revealed that SOD genes are distributed across 7 chromosomes in Gossypium arboreum and 8 chromosomes in Gossypium raimondii. Segmental duplication is predominant duplication event and major contributor for expansion of SOD gene family. Gene structure and protein structure analysis showed that SOD genes have conserved exon/intron arrangement and motif composition. Microarray-based expression analysis revealed that SOD genes have important function in abiotic stress. Moreover, the tissue-specific expression profile reveals the functional divergence of SOD genes in different organs development of cotton. Taken together, this study has imparted new insights into the putative functions of SOD gene family in cotton. Findings of the present investigation could help in understanding the role of SOD gene family in various aspects of the life cycle of cotton.

Molecular and biochemical evidence for phenylpropanoid synthesis and presence of wall-linked phenolics in cotton fibers.

The mature cotton (Gossypium hirsutum L.) fiber is a single cell with a typically thickened secondary cell wall. The aim of this research was to use molecular, spectroscopic and chemical techniques to investigate the possible occurrence of previously overlooked accumulation of phenolics during secondary cell wall formation in cotton fibers. Relative quantitative reverse transcription-polymerase chain reaction analysis showed that GhCAD6 and GhCAD1 were predominantly expressed among seven gene homologs, only GhCAD6 was up-regulated during secondary wall formation in cotton fibers. Phylogenic analysis revealed that GhCAD6 belonged to Class I and was proposed to have a major role in monolignol biosynthesis, and GhCAD1 belonged to Class III and was proposed to have a compensatory mechanism for monolignol biosynthesis. Amino acid sequence comparison showed that the cofactor binding sites of GhCADs were highly conserved with high similarity and identity to bona fide cinnamyl alcohol dehydrogenases. The substrate binding site of GhCAD1 is different from GhCAD6. This difference was confirmed by the different catalytic activities observed with the enzymes. Cell wall auto-fluorescence, Fourier transform infrared spectroscopy (FTIR), high-performance liquid chromatography (HPLC) and chemical analyses confirmed that phenolic compounds were bound to the cell walls of mature cotton fibers. Our findings may suggest a potential for genetic manipulation of cotton fiber properties, which are of central importance to agricultural, cotton processing and textile industries.