Transcriptome and Metabolome Profiling Unveil Pigment Formation Variations in Brown Cotton Lines (Gossypium hirsutum L.).

Naturally brown colored cotton (NBCC) is becoming increasingly popular due to its natural properties of coloration. However, poor fiber quality and color fading are key issues that are hindering the cultivation of naturally colored cotton. In this study, based on transcriptome and metabolome of 18 days post-anthesis (DPA), we compared the variations of pigment formation in two brown cotton fibers (DCF and LCF), with white cotton fiber (WCF) belonging to a near-isogenic line. A transcriptome study revealed a total of 15,785 differentially expressed genes significantly enriched in the flavonoid biosynthesis pathway. Furthermore, for flavonoid biosynthesis-related genes, such as flavonoid 3'5'-hydroxylase (F3'5'H), anthocyanidin synthase (ANS), anthocyanidin reductase (ANR), chalcone synthase (CHS), dihydroflavonol 4-reductase (DFR), and chalcone isomerase (CHI), their expressions significantly increased in LCF compared with DCF and WCF. Moreover, transcription factors MYB and bHLH were significantly expressed in LCF and DCF. Most flavonoid-related metabolites (myricetin naringenin, catechin, epicatechin-epiafzelechin, and epigallocatechin) were found to be more highly up-regulated in LCF and DCF than WCF. These findings reveal the regulatory mechanism controlling different brown pigmentation in cotton fibers and elucidate the need for the proper selection of high-quality brown cotton fiber breeding lines for promising fiber quality and durable brown color pigmentation.

Genome-wide identification of ABCC gene family and their expression analysis in pigment deposition of fiber in brown cotton (Gossypium hirsutum).

ABC (ATP-binding cassette) transporters are a class of superfamily transmembrane proteins that are commonly observed in natural organisms. The ABCC (ATP-binding cassette C subfamily) protein belongs to a subfamily of the ABC protein family and is a multidrug resistance-associated transporter that localizes to the tonoplast and plays a significant role in pathogenic microbial responses, heavy metal regulation, secondary metabolite transport, and plant growth. Recent studies have shown that the ABCC protein is also involved in the transport of anthocyanins/proanthocyanidins (PAs). To clarify the types and numbers of ABCC genes involved in PA transport in Gossypium hirsutum, the phylogenetic evolution, physical location, and structure of ABCC genes were classified by bioinformatic methods in the upland cotton genome, and the expression levels of these genes were analyzed at different developmental stages of the cotton fiber. The results showed that 42 ABCC genes were initially identified in the whole genome of upland cotton; they were designated GhABCC1-42. The gene structure and phylogenetic analysis showed that the closely related ABCC genes were structurally identical. The analysis of chromosomal localization demonstrated that there were no ABCC genes on the chromosomes of AD/At2, AD/At5, AD/At6, AD/At10, AD/At12, AD/At13, AD/Dt2, AD/Dt6, AD/Dt10, and AD/Dt13. Outside the genes, there were ABCC genes on other chromosomes, and gene clusters appeared on the two chromosomes AD/At11 and AD/Dt8. Phylogenetic tree analysis showed that some ABCC proteins in G. hirsutum were clustered with those of Arabidopsis thaliana, Vitis vinifera and Zea mays, which are known to function in anthocyanin/PA transport. The protein structure prediction indicated that the GhABCC protein structure is similar to the AtABCC protein in A. thaliana, and most of these proteins have a transmembrane domain. At the same time, a quantitative RT-PCR analysis of 42 ABCC genes at different developmental stages of brown cotton fiber showed that the relative expression levels of GhABCC24, GhABCC27, GhABCC28, GhABCC29 and GhABCC33 were consistent with the trend of PA accumulation, which may play a role in PA transport. These results provide a theoretical basis for further analysis of the function of the cotton ABCC genes and their role in the transport of PA.

Integrative Analysis of the Expression of SIGLEC Family Members in Lung Adenocarcinoma via Data Mining.

Background: Sialic acid-binding immunoglobulin-type lectin (SIGLEC) family members are involved in regulating immune-cell activation, proliferation, and apoptosis, and they play an important role in tumor development. However, their expression and correlation with immune molecules in lung adenocarcinoma (LUAD) remain unclear. Methods: We utilized Gene Expression Profiling Interactive Analysis, Kaplan-Meier analysis, the limma package in R/Bioconductor, the University of California Santa Cruz Cancer Genome Browser, cBioPortal, STRING, Cytoscape, DAVID, and the Tumor Immune Estimation Resource for gene and protein profiling and analyses. Results: The results showed that SIGLEC10 and SIGLEC15 levels were upregulated in LUAD, whereas SIGLEC1, CD22 (SIGLEC2), CD33, myelin-associated glycoprotein (SIGLEC4), SIGLEC5, SIGLEC6, SIGLEC7, SIGLEC8, SIGLEC11, and SIGLEC14 levels were significantly downregulated, with their low expression associated with poor overall survival. Moreover, we observed high SIGLEC-mutation rates (22%) in LUAD patients, with SIGLEC functions determined as primarily involved in regulating the immune response, signal transduction, inflammatory response, and cell adhesion. Furthermore, we found that SIGLEC expression was significantly correlated with immune-cell infiltration, especially macrophages, neutrophils, and dendritic cells, and highly associated with immune molecules such as CD80, CD86, CD28, B-cell-activating factor, programmed cell death 1 ligand 2, and colony stimulating factor 1 receptor. Conclusion: These results provide insight into the potential molecular mechanism associated with SIGLEC-related development of LUAD, as well as clues for screening biomarkers and therapeutic targets.

Genome-wide identification and expression analysis of the NCED family in cotton (Gossypium hirsutum L.).

Abscisic acid (ABA) is an important plant hormone that plays multiple roles in regulating growth and development as well as in stress responses in plants. The NCED gene family includes key genes involved in the process of ABA synthesis. This gene family has been found in many species; however, the function of the NCED gene family in cotton is unclear. Here, a total of 23 NCED genes (designated as GhNCED1 to GhNCED23) were identified in cotton. Phylogenetic analysis indicated that the identified NCED proteins from cotton and Arabidopsis could be classified into 4 subgroups. Conserved motif analysis revealed that the gene structure and motif distribution of proteins within each subgroup were highly conserved. qRT-PCR and ABA content analyses indicated that NCED genes exhibited stage-specific expression patterns at tissue development stages. GhNCED5, GhNCED6 and GhNCED13 expression was similar to the change in ABA content, suggesting that this gene family plays a role in ABA synthesis. These results provide a better understanding of the potential functions of GhNCED genes.

Vaccination with recombinant flagellar proteins FlgJ and FliN induce protection against Brucella abortus 544 infection in BALB/c mice.

Brucella has been considered as a non-motile, facultative intracellular pathogenic bacterium. However, the genome sequences of different Brucella species reveal the presence of the flagellar genes needed for the construction of a functional flagellum. Due to its roles in the interaction between pathogen and host, we hypothesized that some of the flagellar proteins might induce protective immune responses and these proteins will be good subunit vaccine candidates. This study was conducted to screening of protective antigens among these flagellar proteins. Firstly, according to the putative functional roles, a total of 30 flagellar genes of Brucella abortus were selected for in vitro expression. 15 of these flagellar genes were successfully expressed as his-tagged recombinant proteins in Escherichia coli ER2566. Then, these proteins were purified and used to analyze their T cell immunity induction activity by an in vitro gamma interferon (IFN-γ) assay. Five of the flagellar proteins could stimulate significantly higher levels of IFN-γ secretion in splenocytes from S19 immunized mice, indicating their T cell induction activity. Finally, immunogenicity and protection activity of these 5 flagellar proteins were evaluated in BALB/c mice. Results showed that immunization with FlgJ (BAB1_0260) or FliN (BAB2_0122) plus adjuvant could provide protection against B. abortus 544 infection. Furthermore, mice immunized with FlgJ and FliN developed a vigorous immunoglobulin G response, and in vitro stimulation of their splenocytes with immunizing proteins induced the secretion of IFN-γ. Altogether, these data suggest that flagellar proteins FlgJ and FliN are protective antigens that could produce humoral and cell-mediated responses in mice and candidates for use in future studies of vaccination against brucellosis.

Identification of Brucella abortus virulence proteins that modulate the host immune response.

Brucellosis is an important zoonotic disease of almost worldwide distribution. One significant immune phenomenon of this disease is the ability of the pathogen to hide and survive in the host, establishing long lasting chronic infections. Brucella was found to have the ability to actively modulate the host immune response in order to establish chronic infections, but the mechanism by which the pathogen achieves this remains largely unknown. In our screening for protective antigens of Brucella abortus, 3 proteins (BAB1_0597, BAB1_0917, and BAB2_0431) were found to induce significantly higher levels of gamma interferon (IFNγ) in splenocytes of PBS immunized mice than those immunized with S19. This finding strongly implied that these three proteins inhibit the production of IFNγ. Previous studies have shown that LPS, PrpA, and Btp1/TcpB are three important immunomodulatory molecules with the capacity to interfere with host immune response. They have been shown to have the ability to inhibit the secretion of IFNγ, or to increase the production of IL-10. Due to the role of these proteins in virulence and immunomodulation, they likely offer significant potential as live, attenuated Brucella vaccine candidates. Understanding the mechanisms by which these proteins modulate the host immune responses will deepen our knowledge of Brucella virulence and provide important information on the development of new vaccines against Brucellosis.

Immunization of mice with recombinant protein CobB or AsnC confers protection against Brucella abortus infection.

Due to drawbacks of live attenuated vaccines, much more attention has been focused on screening of Brucella protective antigens as subunit vaccine candidates. Brucella is a facultative intracellular bacterium and cell mediated immunity plays essential roles for protection against Brucella infection. Identification of Brucella antigens that present T-cell epitopes to the host could enable development of such vaccines. In this study, 45 proven or putative pathogenesis-associated factors of Brucella were selected according to currently available data. After expressed and purified, 35 proteins were qualified for analysis of their abilities to stimulate T-cell responses in vitro. Then, an in vitro gamma interferon (IFN-γ) assay was used to identify potential T-cell antigens from B. abortus. In total, 7 individual proteins that stimulated strong IFN-γ responses in splenocytes from mice immunized with B. abortus live vaccine S19 were identified. The protective efficiencies of these 7 recombinant proteins were further evaluated. Mice given BAB1_1316 (CobB) or BAB1_1688 (AsnC) plus adjuvant could provide protection against virulent B. abortus infection, similarly with the known protective antigen Cu-Zn SOD and the license vaccine S19. In addition, CobB and AsnC could induce strong antibodies responses in BALB/c mice. Altogether, the present study showed that CobB or AsnC protein could be useful antigen candidates for the development of subunit vaccines against brucellosis with adequate immunogenicity and protection efficacy.