Genome-wide Identification And Structure-function Analysis Of CDPK And FAD Gene Families In Two Diploid Cotton Species

Cotton is one of the important economic crops in China. Cotton fiber is the main natural textile fiber, and cottonseed, which is rich in fat and protein, is now an important source of edible oil, industrial oil and animal feedstuff. Cotton is mostly grown in tropical and subtropical regions of the world, and its cultivation has been achieved even in relatively cold regions. Low temperature (under15℃) can adversely affect plant development, resulting in poor germination, infection by fungi and other disease-causing organisms, which ultimately cause significant losses in yield. Adaptation of plants to the environmental stresses includes the perception of stress signals and subsequent signal transduction, leading to the activation of various physiological and metabolic responses, which allow plants to set up new balance of material and energy metabolism, to maintain their steady growth under low temperature. In these physiological and metabolic processes, changes in plasma membrane occupy an extremely important position, because the plant cell membrane interfaces to the outside surroundings. The stresses impact in the plasma membrane firstly, and the irreversible damage on membrane system caused by low temperature stress occurs on the phase changes of biological membrane lipid molecules. Calcium-dependent protein kinases (CDPKs), which just rely on Ca2+, participate in the regulation of many signal transduction pathways. Membrane-bound fatty acid desaturases (membrane-bound FADs) are key enzymes that catalyze desaturation of fatty acids.These two enzymes play very important roles in signal transduction and maintenance of plasma membrane properties under the stress of low temperature, respectively. Based on genome sequences of two diploid cotton species, Gossypium raimondii and Gossypium arboreum, CDPK and membrane-bound FAD gene families were analyzed at the genome-wide level, and expression profiles of these genes under low temperature were also studied in this reseach. The main results were as follows:1. A genome-wide identification of CDPK genes in G. raimondii was carried out. 41CDPK genes were identified and distributed unevenly among13G. raimondii chromosomes. All these CDPK proteins were divided into four subgroups. Segmental duplication events led to CDPK gene family expansion in G. raimondii, and the duplicated CDPK genes might have experienced strong purifying selection pressure with limited functional divergence. The majority of GrCPKs contained the low temperature-responsive elements (LTREs) in the promoter regions, and the expression profiling of all selected11CDPK genes response to cold stress showed that eight genes were induced by cold treatment, indicating that CDPK genes were involved in regulating cotton response to cold stress.2. A comparative analysis of CDPK genes in G. arboreum and G. raimondii was performed.43CDPK genes were identified in G. arboreum, which was two more than in G. raimondii.42CDPK genes were located on12G. arboreum chromosomes and one gene was mapped on the scaffold. There were41orthologous gene pairs between G. arboreum and G. raimondii, which shared highly conserved gene structures and protein domains. The expression patterns of the orthologous gene pairs between two diploid cotton genomes were divergent, which might be caused by adapting to their surrounding environment.3. A systematic analysis of membrane-bound FAD gene family in G. raimondii was conducted. The identified19membrane-bound FAD genes were mapped on11G. raimondii chromosomes and categorized into four subgroups. Tandem duplication might have led to the increasing size of the FAD2cluster in the Omega Desaturase subfamily, whereas segmental duplication appeared to be the dominant mechanism for the expansion of the Sphingolipid and Front-end Desaturase subfamilies. Expression profiles of membrane-bound FAD genes in different tissues of G. raimondii showed that GrFAD2.2, GrFAD3.1, GrFAD8.2and GrDSD2shared high expression levels in ovule, especially GrFAD2.2, which could be a candidate gene for the gene engineering to improve the quality of cottonseed oil. Seven membrane-bound FAD genes were significantly up-regulated and five genes were greatly suppressed in G. raimondii leaves exposed to low temperature conditions, which indicated that membrane-bound FAD genes might participate in low temperature stress response. For this important enzyme, the existence of a pseudogene GrFAD2.1was revealed in G. raimondii.4. A comparative analysis of membrane-bound FAD genes in G. arboreum and G. raimondii was performed.20membrane-bound FAD genes were identified in G. arboreum and mapped on12chromosomes. Compared with G. raimondii, G. arboreum contained two more FAD3genes and one less FAD2gene. There were18orthologous gene pairs between G. arboreum and G. raimondii, and their gene structures and protein domains were highly conserved. GrFAD2.2and GaFAD2.1exhibited highest transcript abundance in ovule compared with other tissues examined, which suggested that GrFAD2.2and GaFAD2.1might contribute to the high content of linoleic acid in cottonseed. GaFAD3.3and GaSLDl showed the highest expression level in fiber, which indicated that these two genes might play important roles during fiber elongation. The expression patterns of the orthologous gene pairs between two diploid cotton genomes were different, which suggested that the function of these genes have been divergent during the process of evolution. There was also a pseudogene GaFAD3.2in G. arboreum.In summary, the systematic analyses of CDPK and membrane-bound FAD genes were performed in G. arboreum and G. raimondii through applying comparative genomics approach. The phylogenetic analysis showed that both two gene families underwent specific expansion in cotton genomes. Expression patterns under low temperature indicated that CDPK and membrane-bound FAD genes participated in cotton response to low temperature stress. Different expression patterns of the orthologous gene pairs between two diploid cotton genomes implied that the function of these genes have been divergent during the process of evolution. These results will provide valuable information for breeding stress-resistant cotton and further studying on the biological function and evolutionary relationship of the two gene families in Gossypium.