Proteomic Analysis Of Maize Axial-roots Elongation Induced By Low Phosphorus Stress

As an essential macronutrient for plant growth and development, phosphorus (P) not only is important component of many compounds in plants, but also plays important roles in photosynthesis, energy metabolism, enzyme activation/inactivation, signal transduction etc.. P is involved in almost all life activities, so its deficiency seriously affects the growth and yield of crops. Due to the low soil P content and easily with other elements to form insoluble P, the concentration of available phosphate (Pi) by roots in soil is very low, which can not satisfy the demand of plants to acquire sufficient Pi. Maize (Zea mays L.) is an important food and cash crops, occupies a pivotal position in the national economy. Lack of available Pi by roots in soil has become an important factor that restricting yield of corn. The plants mainly absorb soil Pi by roots which is of high plasticity. Roots development and morphology is particularly important for the absorption of Pi. Therefore, exploiting the regulatory mechanism of root morphogenesis mediated by Low-P stress is of important theoretical significance and potential practical value.Growth and development changes of maize axile roots with the extension of the time for low-P treatmentIn the study, the hydroponic cultured seedlings of maize inbred line, endosperms of which had been removed, were used as a rapidly response system of P deficiency, to observe and analyze the changing of growth rate and length of axis roots, phosphorus concentration and biomass of plant etc. with the extension of the time for low-P treatment. These results showed that P concentration in vivo continued to reduce, the growth and development of shoots was inhibited, while the growth of axis roots was induced at a certain extent during the treatment of low-P. At the first three days of low-P treatment, the growth rate of maize axis roots was not significantly different, compared to+P condition; to the7th day of low-P treatment, the growth rate of maize axis roots was accelerated, compared to+P condition, to the11th day the difference became more apparent between-P and+P. As the above causes, axis roots length of maize showed elongation trend at the7th day of-P treatment, and to the1lth day of-P treatment, the axis roots length was significantly longer than that of+P. At the cellular level, the increase of cell number and/or length is two important reasons which may lead to the elongation of axis roots. To clear the main reason of maize axis roots elongation under low-P stress, the cell morphology of the maturation zone and meristem zone in the maize axis roots were observed by staining paraffin sections with hematoxylin. The results showed that the cell size in maturation zone of axis roots was not significantly different between-P and+P plants during low-P treatment. However, with the extension of time for low-P treatment, cell division capacity in meristem zone of axis roots had been enhanced gradually and showed obvious differences at the11th day of low-P treatment. The above results indicated that under low-P stress the growth acceleration of axis roots and eventually leading to its length increase are the results of increased activity of apical cell proliferation.Comparative proteomic analysis of the apical fragments in axis roots under different low-P treatmentsThe regulatory mechanism of growth acceleration and elongation in maize axis roots mediated by low-P stress is still quite unclear by far. To better understand the regulatory mechanism of maize axis roots responsive to low-P environment, we conducted the comparative proteome analysis for proteins isolated from apical fragments of axis roots treated with1000μM (control) or5μM KH2PO4for1d,3d,7d and11d. At low-P treatment for1d,3d,7d and11d, approximate924,936,947and962protein spots were detected by2-DE, respectively. And48,71,80and70protein spots of which changed significantly in amount (P<0.05;=2folds). The results showed that under low-P stress the proteome of maize axis roots had a large degree of changes. Further analysis of these differentially expressed proteins, we found that under different degree of low-P stress there are great difference between the proteins which participated in P-starvation response of maize axis roots, only a few differentially expressed proteins were involved in the different degree low-P stress processes. Therefore, under different P-deficiency level maize root system will start different P-stress adaptive regulatory mechanism, to ensure normal growth and development of roots. The results indicated that the growth acceleration and elongation of axis roots induced by low-P stress is a complicated process of dynamic regulation at proteome level.At the1st,3rd,7th and11th day of low-P treatment, we identified42,56,62and53differentially accumulated proteins by MALDI-TOF MS, respectively. These identified proteins represented a large range of functional categories, including metabolism, protein fate and synthesis, secondary metabolism, cellular organization, defense/interaction with environment, transport, transcription/cell cycle/signal transduction and unclassified or unknown proteins. The results indicated that the changes of growth and low-P tolerance of maize axis roots are involved in a dynamic regulation process related with multiple metabolic pathways.Several differently accumulated proteins may play important roles in the changes of growth and development of maize axis roots induced by low-P stress. Among them, a large number of proteins are involved in glycolysis, citric acid cycle, pentose phosphate pathway and amino acid metabolism, such as sucrose synthase2, pyruvate dehydrogenase El component subunit beta,3-phosphate dehydrogenase, UDP-glucose pyrophosphorylase, triose phosphate isomerase, enolase,6-phosphogluconate dehydrogenase, ATP-citrate synthase, malate dehydrogenase, etc. In other words, some carbon metabolism related proteins participated in the process of perception low-P environment and metabolic regulation of maize axis roots. Combined with previous reports, it could be speculated that sugar not only as metabolites, but also as signal molecule participates in low-P response process of maize axis roots. Further analysis of these carbon metabolism-related proteins indicated that during the process of low-P stress most of the amino acid metabolism-related proteins abundance decreased, while most of the glycolysis, pentose phosphate and citric acid cycle pathways proteins abundance relatively increased. The results indicated that under low-P stress more carbon sources flow to glycolysis, pentose phosphate and tricarboxylic acid cycle pathways to produce more energy, reducing power, and the intermediate sugar molecules to meet the needs of maize axis root growth and development. This may also be an important reason for growth acceleration and elongation of maize axis roots under low-P stress. In addition, several phytohormone metabolism and signal transducti on-associated proteins were also involved in the low-P stress process of maize axis roots, such as beta-glucosidase, importin a subunit protein, SRC2, serine threonine kinase receptor associated protein, type Ⅰ protein phosphatases, transcription factors BTF3etc.. These proteins were likely to regulate the process of low-P-mediated growth and development of maize axis roots by participating in the regulation of cell cycle and intracellular signal transduction etc..In the work, we observed the growth changes of maize axis roots and its cell morphology of the maturation zone and meristem zone, and conducted on the dynamic proteomic analysis of apical fragments of axis roots under low-P treatment. The results cleared the inherent relationship between growth changes of maize axis roots and phosphorus deficiency; revealed that under low-P stress the increase of meristematic cell proliferation activity is the main reason of maize axis roots growth acceleration and elongation; revealed that the regulations of carbon metabolism, cell cycle and cell signal transduction play important roles in growth and development of maize axis roots mediated by low-P stress at the proteome level; showed that the growth changes of maize axis roots mediated by low-P stress is a complicated and dynamic regulation process related with multi-gene and multi-pathway, which involved the changes of cellular metabolic state and gene expression. At the same time, the results also promoted our understanding on regulation mechanisms of maize P nutrition, and provided ideas and objects for establishing the experiment designs for improving low-P tolerance of crops.