Identification of thaumatin-like protein family in Saccharum spontaneum and functional analysis of its homologous gene in sugarcane cultivar

Thaumatin like protein (TLP) plays an important role in plant growth and development and resistance to environmental stress. In order to fully understand the structure and functional characteristics of sugarcane TLP gene family, 122 SsTLP family genes were screened and analyzed for their physicochemical properties, gene chromosomal distribution, gene structure, and phylogenetic evolution using the genomic database of Saccharum spontaneum and bioinformatics methods. In addition, one homologous gene of SsTLP87, ScTLP1, was cloned from a major sugarcane cultivar ROC22 in mainland China. Gene expression pattern, subcellular localization, transient expression, and transcriptional self-activation activity of ScTLP1 were identified. The results showed that 122 SsTLP family genes were distributed on 28 chromosomes, and there was a gene clustering with 1-13 SsTLP genes on each chromosome. Most SsTLP proteins were predicted to be acidic, hydrophobic, and unstable proteins which located in cytoplasm. There were many types and distributions of intron number (1-30), intron phase (0-2), and motif (1-5) in SsTLP family genes. Phylogenetic tree analysis indicated that the coding proteins of SsTLP genes were clustered in the I-X branches of TLP family, among which, the number of the class V SsTLPs with antibacterial potential was the most (36). ScTLP1 gene obtained from sugarcane cultivar ROC22 encoded 227 amino acid residues, belonging to the class V member. ScTLP1 was located in the cell membrane and had no transcriptional self-activation activity. The similarity of amino acid sequence between ScTLP1 and SsTLP87 was as high as 99.56%. qRT-PCR demonstrated that ScTLP1 gene was constitutively expressed in different tissues of sugarcane with the highest expression level in the bud. The expression level of ScTLP1 gene had no significant change under the treatments of abscisic acid (ABA) and salicylic acid (SA), but it was up-regulated by Sporisorium scitaminea, methyl jasmonate (MeJA), sodium chloride (NaCl), polyethylene glycol (PEG), and low temperature of 4℃. After transient overexpression of ScTLP1 gene, DAB staining in Nicotiana benthamiana leaves was deepened, and the expression level of NtEFE26 and NtPR3 was up-regulated. Furthermore, the incidence of transient overexpression of ScTLP1 gene in N. benthamiana leaves inoculated with Pseudomonas solanacearum and Fusarium solani var. coeruleum was lighter than that of the control, suggesting that ScTLP1 gene could induce an allergic reaction of N. benthamiana, participate in the ethylene and jasmonic acid signal transduction pathways, and enhance the defense effect of N. benthamiana against pathogens. These results laid a good foundation for further study on the structure and function of the TLP gene family in sugarcane.