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Transcriptome analysis and phenotyping of walnut seedling roots under nitrogen stresses

ABSTRACT: Nitrogen is an essential core element in walnut seedling growth and development. However, nitrogen starvation and excessive nitrogen stress can cause stunted growth and development of walnut seedlings, and environmental pollution is also of concern. Therefore, it is necessary to study the mechanism of walnut seedling resistance to nitrogen stress. In this study, morphological and physiological observations and transcriptome sequencing of walnut seedlings under nitrogen starvation and excess nitrogen stress were performed. The results showed that walnut seedlings under nitrogen starvation and excess stress could adapt to the changes in the nitrogen environment by changing the coordination of their root morphology and physiological indexes. Based on an analysis of transcriptome data, 4911 differential genes (DEGs) were obtained (2180 were upregulated and 2731 were downregulated) in a comparison of nitrogen starvation and control groups. A total of 9497 DEGs (5091 upregulated and 4406 downregulated) were obtained in the comparison between the nitrogen overdose and control groups. When these DEGs were analysed, the differential genes in both groups were found to be significantly enriched in the plant’s circadian pathway. Therefore, we selected the circadian rhythm as the focus for further analysis. We made some discoveries by analysing the gene co-expression network of nitrogen metabolism, circadian rhythm, and hormone signal transduction. (a) Nitrite nitrogen (NO2−) or Glu may act as a nitrogen signal to the circadian clock. (b) Nitrogen signalling may be input into the circadian clock by regulating changes in the abundance of the CRY1 gene. (c) After the nitrogen signal enters the circadian clock, the expression of the LHY gene is upregulated, which causes a phase shift in the circadian clock. (d) The RVE protein may send information about the circadian clock’s response to nitrogen stress back to the nitrogen metabolic pathway via the hormone transduction pathway. In conclusion, various metabolic pathways in the roots of walnut seedlings coordinated with one another to resist the ill effects of nitrogen stress on the root cells, and these coordination relationships were regulated by the circadian clock. This study is expected to provide valuable information on the circadian clock regulation of plant resistance to nitrogen stress.

SUBMITTER: Song Y

PROVIDER: S-EPMC9283388 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Potato crop requires high dose of nitrogen (N) to produce high tuber yield. Excessive application of N causes environmental pollution and increases cost of production. Hence, knowledge about genes and regulatory elements is essential to strengthen research on N metabolism in this crop. In this study, we analysed transcriptomes (RNA-seq) in potato tissues (shoot, root and stolon) collected from plants grown in aeroponic culture under controlled conditions with varied N supplies i.e. low N (0.2 milli molar N) and high N (4 milli molar N). High quality data ranging between 3.25 to 4.93 Gb per sample were generated using Illumina NextSeq500 that resulted in 83.60-86.50% mapping of the reads to the reference potato genome. Differentially expressed genes (DEGs) were observed in the tissues based on statistically significance (p ≤ 0.05) and up-regulation with ≥ 2 log2 fold change (FC) and down-regulation with ≤ -2 log2 FC values. In shoots, of total 19730 DEGs, 761 up-regulated and 280 down-regulated significant DEGs were identified. Of total 20736 DEGs in roots, 572 (up-regulated) and 292 (down-regulated) were significant DEGs. In stolons, of total 21494 DEG, 688 and 230 DEGs were significantly up-regulated and down-regulated, respectively. Venn diagram analysis showed tissue specific and common genes. The DEGs were functionally assigned with the GO terms, in which molecular function domain was predominant in all the tissues. Further, DEGs were classified into 24 KEGG pathways, in which 5385, 5572 and 5594 DEGs were annotated in shoots, roots and stolons, respectively. The RT-qPCR analysis validated gene expression of RNA-seq data for selected genes. We identified a few potential DEGs responsive to N deficiency in potato such as glutaredoxin, Myb-like DNA-binding protein, WRKY transcription factor 16 and FLOWERING LOCUS T in shoots; high-affinity nitrate transporter, protein phosphatase-2c, glutaredoxin family protein, malate synthase, CLE7, 2-oxoglutarate-dependent dioxygenase and transcription factor in roots; and glucose-6-phosphate/phosphate translocator 2, BTB/POZ domain-containing protein, F-box family protein and aquaporin TIP1;3 in stolons, and many genes of unknown function. Our study highlights that these potential genes play very crucial roles in N stress tolerance, which could be useful in augmenting research on N metabolism in potato.

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Transcriptome analysis and phenotyping of walnut seedling roots under nitrogen stresses

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