Oxidative enzyme activities and protein profile of transgenic rice Oryza sativa L. overxpressing calmodulin gene OsCam1-1

Abstract

Rice (Oryza sativa L.) in which a large family of CaM and CaM-related proteins has been found, is one of the most important crops in the world and is the staple food for about three billion people. Thailand is one of the most important rice-exporter and rice-producer countries in the rice-world market. However, rice production is confronting the saline soil problem and recently coping with the low yielding. Of five OsCam genes, OsCam1-1 was shown to be rapidly and strongly increased in leaves under osmotic stress and is shown to play important roles in salt-stress responses. A proteomic study was performed to characterize the responses of transgenic rice Oryza sativa L. cv. KDML105, harboring OsCam1-1 gene to simulated salt stress. The emphasis of the present study was to investigate the overall protein expression changes when exposed to salt. Three sets of rice seeds (wild-type, transgenic line with vector alone and transgenic line with OsCam1-1 gene) were germinated in MS media for 3 weeks and stressed by 150 mM NaCl. Proteins in the shoot and the root of the stressed rice plants along with their controls were extracted and in-gel digested. Peptides of these samples were analysed by mass spectrometry. Each identified protein was functionally classified according to their known and putative functions by using STRAP software program. These proteins were categorized in to 7 groups based on their biological functions including cellular process (42%), metabolic process (14%), regulation (12%), developmental process (2%), localization (2%), response to stimulus (2%) while the rest are proteins in other processes or of unknown function. Many of the proteins in shoot and root responded to the induced salt stress were localized in nucleus (24%) and also in chromosome (5%). Significant fractions were also located in mitochondria (5%), ER (3%), and other intracellular organelles (5%) while only a small fraction was located in cytoplasm. Some are part of macromolecular complex (8%). Comparison of protein expression profiles among the three rice lines in both shoots and roots by using Venn diagram revealed the unique salt-responsive proteins in transgenic rice lines that constitutively over-express the OsCam1-1 gene when compared to the wild type and the control transgenic lines without OsCam1-1 over-expression. In addition, the complementary data on enzyme activity analysis (Peroxidase, Ascorbate peroxidase, Glutathione-S-transferase, and Adenosine Triphosphatase) in the the wild-type and the transgenic rice lines were also investigated. These results suggested that these proteins could play important roles in mediating plant response to salt stress and indicated that OsCam1-1 overexpression probably contributes to salt resistance in rice through these proteins.