FUNCTION OF RICE NUCLEOLIN1 IN SALT-RESISTANT ABILITY OF ARABIDOPSIS

Abstract

The objectives of this dissertation have two main goals; the first is to compare the LPT123 and its salt-tolerant mutation line, LPT123-TC171, at the genomic level to indicate the variations that contribute to stress-tolerant character. Second objective is to characterize the function of Nucleolin1 gene from rice (OsNUC1) on salt-adaptive mechanism. OsNUC1 presents in two splicing forms which are OsNUC1-S and OsNUC1-L.To elucidate the function of OsNUC1, I generated the transgenic Arabidopsis, with OsNUC1 over-expression under the control of constitutive promoter. LPT123-TC171 is the salt and drought tolerant rice line which was selected from somaclonal variation of the original Thai rice, LPT123. Genome of LPT123 and LPT123-TC171 were comparatively studied in 3 levels which are chromosome level (polyploidy, aneuploidy), chromosome structure and DNA sequence changes by whole genome sequencing analysis. The results showed that LPT123-TC171 did not show the changes in the set of chromosome (polyploidy) or number of chromosome (aneuploidy). Interestingly, in the study of chromosome structure, the significant deficiency of chromosome ends (telomeres) in the mutant line were found. This change could be resulted from the oxidative stress in the screening process. In addition, the functional genomic approach or exome sequencing gave the new aspects of genome response to in vitro condition. Exome sequencing revealed the molecular spectrum and pattern of change of somaclonal variant. The major nucleotide substitutions in somaclonal variant were A↔G and T↔C transitions. Moreover, the total point mutation were found which are 493 positions within the known genes responding to both 17 drought and salt stress genes and 100 positions within 6 salt-responsive genes. NUC is the multifunctional nucleolar protein found in all eukaryotes that involved in RNA metabolism, mRNA stability and ribosome biogenesis process. The recent study in Arabidopsis showed that NUC has functions in plant growth and development. In rice, OsNUC1 presents in two mRNA species, OsNUC1-S and OsNUC1-L, which were increased the expression after salt treatment. By using transgenic Arabidopsis, it revealed the new role of OsNUC1-S. The transgenic Arabidopsis with the overexpression of OsNUC1-S showed hypersensitive to ABA during seed germination. Based on the transcriptome analysis, 9-cis-epoxycarotenoid dioxygenase (NCED), the gene encoding the key enzyme in ABA biosynthesis, was dramatically up-regulated in transgenic Arabidopsis up to 22 fold, suggesting that OsNUC1-S may increase the salt-tolerant ability via the increase of ABA production. These were also supported with the increase of the ABA inducible gene expression in transgenic Arabidopsis grown in salt condition The OsNUC1-L gene enhanced salt-tolerant ability in transgenic Arabidopsis. Besides, they also displayed some developmental changes such as early flowering and short life-cycle. Based on transcriptomic study, the transgenic Arabidopsis dominantly enriched group of genes in photosynthetic process, ribosome and plasma membrane component leading to the role of OsNUC1-L in salt stress adaptation. These results suggested that overexpression of the OsNUC1 gene in transgenic Arabidopsis improved salt stress tolerance by regulating expression the genes in some specific pathways.