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Transcriptome Analysis of Tolerant and Susceptible Maize Genotypes Reveals Novel Insights about the Molecular Mechanisms Underlying Drought Responses in Leaves - PubMed

  • ️Fri Jan 01 2021

Transcriptome Analysis of Tolerant and Susceptible Maize Genotypes Reveals Novel Insights about the Molecular Mechanisms Underlying Drought Responses in Leaves

Joram Kiriga Waititu et al. Int J Mol Sci. 2021.

Abstract

Maize (Zea mays L.) is the most essential food crop in the world. However, maize is highly susceptible to drought stress, especially at the seedling stage, and the molecular mechanisms underlying drought tolerance remain elusive. In this study, we conducted comparative transcriptome and physiological analyses of drought-tolerant (CML69) and susceptible (LX9801) inbred lines subjected to drought treatment at the seedling stage for three and five days. The tolerant line had significantly higher relative water content in the leaves, as well as lower electrolyte leakage and malondialdehyde levels, than the susceptible line. Using an RNA-seq-based approach, we identified 10,084 differentially expressed genes (DEGs) with 6906 and 3178 DEGs been annotated and unannotated, respectively. Two critical sets of drought-responsive DEGs, including 4687 genotype-specific and 2219 common drought-responsive genes, were mined out of the annotated DEGs. The tolerant-line DEGs were predominantly associated with the cytoskeleton, cell wall modification, glycolysis/gluconeogenesis, transport, osmotic regulation, drought avoidance, ROS scavengers, defense, and transcriptional factors. For the susceptible line, the DEGs were highly enriched in the photosynthesis, histone, and carbon fixation pathways. The unannotated DEGs were implicated in lncRNAs, including 428 previously reported and 22% putative TE-lncRNAs. There was consensus on both the physiological response and RNA-seq outcomes. Collectively, our findings will provide a comprehensive basis of the molecular networks mediating drought stress tolerance of maize at the seedling stage.

Keywords: RNA sequencing; drought stress; lncRNA; maize; transcriptome.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1

Phenotypic and physiological responses of drought-tolerant line CML69 and drought-susceptible line LX9801. The seedling phenotypic response of CML69 and LX9801 at; (A) control (C)-well-watered plants; (B) three days stress (3D); (C) five days stress (5D). Physiological effects of drought stress on; (D) relative water content (%), (E) relative electrolyte leakage (%), and (F) MDA content (nmol g−1 FW) at C, 3D, and 5D. Bars with two stars (**) are significantly different at p ≤ 0.01.

Figure 2
Figure 2

Differentially expressed genes (DEGs) in CML69 and LX9801 during drought conditions. (A) Graphic presentation of DEGs in 3D and 5D stress treatment of both CML69 and LX9801. (B) Venn diagram showing DEGs’ profile in both inbred lines after drought treatment. (C) Heatmap showing the clustering analysis of 2219 common drought-responsive genes. (D) Venn diagram showing the comparison of DEGs expressed at 3D drought stress in both inbred lines. (E) Venn diagram showing the comparison of DEGs expressed at 5D drought stress in both inbred lines. Drought treatments are labeled as control (C), three days (3D), and five days (5D).

Figure 3
Figure 3

Gene Ontology enrichment analysis of the common drought-responsive genes. (A) DEGs expressed at 3D drought stress. (B) DEGs expressed at 5D drought stress. The GO terms shown here are the topmost biological process (BP), molecular functions (MF), and cellular component (CC) categories from the tolerant line (CML69) and susceptible line (LX9801). Drought treatments are labeled as control (C), three days (3D), and five days (5D).

Figure 4
Figure 4

KEGG pathway enrichment analysis of the common drought-responsive genes. (A) DEGs expressed at 3D drought stress. (B) DEGs expressed at 5D drought stress. The experimental comparisons were based on the hypergeometric test, while the significance of the enrichment of the KEGG pathway is based on the q value (q < 0.05). The “rich factor” shows the DEGs’ ratio to the total gene number in specific pathways. Drought treatments are labeled as control (C), three days (3D), and five days (5D).

Figure 5
Figure 5

KEGG map of the photosynthetic antenna proteins. It is an analysis of DEGs, comparing drought-treated and control samples in both CML69 and LX9801. Boxes in a red frame indicate that the corresponding DEGs were downregulated in the drought-treated samples, and the boxes with a green frame suggest that the expression levels of the related genes were not changed as determined by our RNA-seq. Drought treatments are labeled as control (C), three days (3D), and five days (5D), and all genes’ abbreviations are defined in Table S9.

Figure 6
Figure 6

KEGG map of the photosynthesis pathway. It is an analysis of DEGs, comparing drought-treated and control samples in both CML69 and LX9801. Boxes in a red frame indicate that the corresponding DEGs were downregulated in the drought-treated samples, and the boxes with a green frame suggest that the expression levels of the related genes were not changed as determined by our RNA-seq. Drought treatments are labeled as control (C), three days (3D), and five days (5D), and all genes’ abbreviations are defined in Table S9.

Figure 7
Figure 7

Correlation analysis between RNA-seq and qRT-PCR methods. Log2fold values of RNA-seq data (x-axis) are plotted against log2fold values of qRT-PCR (y-axis) data.

Figure 8
Figure 8

The schematic molecular model describing the main pathways involved in the acquisition of drought tolerance in maize seedling. The model was constructed based on our main common drought-responsive genes identified in this report, as well as plant abiotic stress pathway schemes previously described. The black and red pointing arrows display the main pathways which were enriched in CML69 and LX9801, respectively. The blue pointing arrows display the common pathways in both drought-tolerant and sensitive inbred lines. Abbreviation’s key: RWC, relative water content; LEA, late embryogenesis abundant; CL, coumarate-CoA ligase; TFs, transcription factors; ncRNAs, noncoding RNAs; lncRNAs, long noncoding RNAs; TE-lncRNAs, transposable elements long noncoding RNAs; RBCs, ribulose carboxylase.

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