Identification of genes involving in chitosan response in rice Oryza sativa L. and Arabidopsis Thaliana L.

Abstract

Chitosan is a natural degradable biopolymer that has a potential in agricultural utilization including plant growth induction. This study aimed to investigate the chitosan response in plants at the molecular level by proteomics, transcriptomics and mutagenesis techniques. Proteomic analysis of ‘LPT123’ rice seedlings treated with foliar application of chitosan at 40 gm/L revealed significant changes of 105 proteins mainly in metabolic process and signaling. Chitosan activated oxidative burst and ROS scavenger to balance ROS homeostasis in the level of signaling molecule rather than toxicity It also enhanced key enzymes in glycolysis and Calvin cycle as well as photosynthetic components. In addition, two receptors involving in plant growth, PSKB and BRL1 were induced Contrastly, application of chitosan at 80 mg/L in Arabidopsis illustrated that chitosan retarded seedlings growth both shoot and root development. Transcript profiling revealed significant change of 1,557 genes mainly in metabolic process and stress responses. High dose chitosan (HDC) inhibited plant growth by disturbing several growth-related genes and receptors. Besides, HDC adversely influenced on photosynthesis by disturbing photosystem I and II, iron homeostasis and promoting chlorophyll breakdown. Chitosan was recognized as PAMPs (pathogen-associated molecular patterns) to activate defense and stress responses. HDC triggered pattern recognition receptors (PRRs), pathogen-related (PR) genes, transcription factors, hormone signaling, and abiotic stress responsive genes. Abscisic acid (ABA) and jasmonic acid (JA) appeared to play a prominent role in HDC response. Taken together, both omic approaches implied that chitosan significantly changed plant metabolism and chloroplast was one of the chitosan target organelles. EMS-induced mutagenized Arabidopsis was achieved to screen for chitosan-resistant mutants in HDC treatment. There were 350 mutants that showed the chitosan resistant phenotypes such as larger in plant size or longer roots compared to wild-type plants. After multiple confirmation tests, there were only 5 candidate mutants that consistently resisted to HDC. Finally. only two of five mutants seemed to possess recessive single gene mutation, which were 106A and 161A mutant lines. these two mutants were subjected to sequencing analysis and mutation identification. The T-DNA insertion mutants carrying putative mutated gene were ordered from ABRC stock center and were subjected to characterize in HDC treatment. The ABRC mutant carrying insertion in TOM3 or OPT4 gene showed the best chitosan-resistant phenotypes. Lastly, the identified genes from transcriptomic data and none of them from mutagenesis analysis. The identification of chitosan responsive genes suggested the better understanding of plant molecular responses to chitosan. It also provided basic information for the improvement of chitosan utilization for plant growth enhancement.