The Gene Expression And Regulation Responding To Low Nitrogen Stress In Root Systems Of Maize Mutant RTCS

Nitrogen, one of the three plant nutrients, directly affects crop yield and protein content which has an important role during the plant growth and development. Nitrogen content either too high or too low will affect the growth of plants. Reducing the nitrogen content of the plant will destroy some important biological processes, such as photosynthesis and nitrogen metabolism. Plant roots are the most directly the organ to absorb nitrogen and subject to nitrogen change of the environment, so root structure of growth and development may affect the absorption of nitrogen,so identification of the related genes that control root growth-related and deciphering of molecular regulatory networks will facilitate efficient nitrogen ultilization-efficiency. In this study, glutamine synthetase activity and determination of chlorophyll contect were detedced between maize mutant (rtcs) and wild-type (B73) after processing 12h,24h,48h,96h to test total protein,under low nitrogen (0.04mM) and normal nitrogen (4mM) condition,respectively. Additionaly, the transcripts of maize root of the four time points were analyzed by RNA-seq to compare gene expression profile between the mutant and wild-type under normal and low nitrogen condition. The main results obtained are as following:1.Mutant roots showed obvious morphological differences compared with wild-type after approximately eight days. There are primary root and seed roots in wild-type, while only primary root root seed in mutant RTCS. which also was observed previously by other group. There were longer main root, more lateral root both mutant and wild type in lower nitrogen conditions compared with normal condition, and the difference becomes more significant as the increasing of time-point. The content of total protein and glutamine synthetase activity were lower both mutant and wild type under low nitrogen condition than normal conditions; Obviously, the total protein and glutamine synthetase enzyme activity of mutant were lower than the wild type under normal conditions and low nitrogen condition. At both nitrogen levels, total chlorophyll content and chlorophyll a content of wild-type and mutant showed a significant increasing trend as time going on, while it is not the same case for, Chlorophyllb content. Those results suggest that the difference of total chlorophyll content between two stains is mainly caused by differences of chlorophyll a content. Chlorophyll content in the leaves was decreased under low nitrogen stress compared with the normal nitrogen condition, and the low nitrogen stress significantly affect chlorophyll content of the mutant than that of wild-type.2. RNA-seq results of mutant and wild-type seedlings roots show that 32-46M clean reads were obtained in 32 samples. There are 78.82%-85.57% of the reads can be mapped to genome sequence of maize B73. And annotated to 27921-28547 known protein-coding genes, in which has 25,285 genes are expressed at all experimental time points and experimental conditions. To explore the related mechanism in root development, the transcriptome were contrasted bettween wild-type and mutant under normal nitrogen conditions, and 7418 differentially expressed genes were identified, Only 10.5%(786 genes) DEGs at four time points showing differential expression. Functional annotation analysis found that these co-modulated DEGs mainly involved in seventy-nine biological pathways, including plant signal transduction, phenylalanine biosynthesis, biological process phenylalanine metabolism, glutathione metabolism, plant and pathogen interaction, starch and sucrose metabolism. Gene Ontology functional enrichment analysis revealed these genes are significantly enriched in 29 GO team, including a biological regulation of primary metabolism regulation, transcriptional regulation, regulation of gene expression, abiotic stress response and so on. K-means clustering analysis showed that co-modulated DEGs are classified into eight groups.3. To explore whether the roots of corn under low nitrogen stress response to gene expression regulation mechanism, gene expression difference of wild type and mutant were compared in normal nitrogen and low nitrogen stress. There are 7977 and 7762 differentially expressed genes related to low nitrogen stress were detected in wild-type and mutant, respectively. Under low nitrogen stress there are 593 genes showed differential expression in four time points, including 194 differentially upregulated expressed genes, 399 differentially down-regulated expressed genes in wild-type. In the mutant under low nitrogen stress condition, there are 804 genes showed differential expression in four time points, including 543 differentially upregulated expressed genes, and 261 differentially down-regulated expressed genes. There are 403 co-modulated DEGs participate in low nitrogen stress response in both stains. These co-modulated DEGs annotated to ninety biological processes, seventy-two molecular function and tweenty-three categories of cell component. Through KOBAS enriched-analysis, two major pathways enriched differentially expressed genes were identified, i.e.,Glycerides metabolism and galactose metabolic. Additional, there are seven and five co-modulated DEGs were annoted to phenylalanine biosynthesis pathway, respectively.4. The changes of genotype-specific transcription levels at four time points caused by low nitrogen stress were analyzed. The results showed there are one hundred and ninety and four hundred and one differentially expressed genes in wild-type and mutant, respectively. A large number of genotype-specific DEGs in wild-type presented in the following GO team:cold stress response, response to abiotic stress, balance multicellular organisms, copper ions, and biological quality of multicellular organisms regulate. Many mutant specific DEGs are classified into subcategories cellular components (membrane, cells, and extracellular ubiquitin ligase complex).