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Cd-Induced Autophagy Responses in Pakchoi as Revealed by Transcriptome Analysis

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Abstract

Cadmium (Cd) is a toxic heavy metal element, which can cause oxidative stress in plant cells and induce cell damage. Currently, the research investigating autophagy function and Cd injury in plants is still limited. In our study, we exposed Pakchoi plants to Cd chloride at different concentrations (0, 2, 4, 6, and 8 mg·L−1) for 4 consecutive weeks. The results showed that toxic damage of Pakchoi was induced by Cd exposure and its Cd injury accumulation showed a positive dose relationship with metal concentrations. When Pakchoi was exposed to Cd, it generated excessive reactive oxygen species (ROS), especially H2O2. Our data revealed that LC3 upregulation and autophagosome formation were noted in H2O2-activated root cells, indicating that Cd damage caused autophagy in Pakchoi. Under Cd stress, the antioxidant enzyme activity of peroxidase (POD), ascorbate peroxidase (APX), catalase (CAT), and superoxide dismutase (SOD) were decreased as Cd concentration was increased, lowest in 8 mg·L−1 treatment. Using transcriptome analysis by RNA sequencing, we identified 860 significant differentially expressed genes (DEGs) with a fold change of no less than 2 between the Cd-treated and the control groups, among which 513 genes were upregulated and 347 genes were downregulated. The DEGs were mostly enriched in functions related to the hormonal regulation of digestion and absorption, transport processes, transporter activity, and the apical plasma membrane composition. A total of 6 autophagy-related genes, including 4 AMPK genes, 1 ATG13 gene, and 1 Beclin1 gene, were found to have significantly different expression levels in the Cd groups. The expression levels of these 6 genes were significantly different between the control and Cd groups (p < 0.05) when validated by real-time quantitative PCR. Furthermore, we found the ratio of p-AMPK and AMPK increased significantly in the Cd group (p < 0.05), indicating that Cd-induced autophagy may be mediated by AMPK phosphorylation. This research provides a better understanding of the stress effect of autophagy during Cd injury.

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Availability of Data and Materials

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Abbreviations

Cd:

Cadmium

ATGs:

Autophagy-related proteins

PI-3K:

Phosphatidylinositol 3-kinase

PI3P:

Phosphatidylinositol triphosphate

PAS:

Pre-autophagosomal structure/phagophore assembly site

PE:

Phosphatidylethanolamine

MDC:

Monodansylcadaverine

SOD:

Superoxide dismutase

APX:

Ascorbic acid peroxidase

CAT:

Catalase

POD:

Peroxidase

ICP-MS:

Inductively coupled plasma mass spectrometry

NBT:

Nitro blue tetrazolium

BCA:

Bicinchoninic acid

SDS-PAGE:

Sodium dodecyl sulfate–polyacrylamide gel electrophoresis

PVDF:

Polyvinylidene difluoride

CEL:

Chemiluminescence

DEGs:

Differentially expressed genes

Nt:

NCBI non-redundant nucleotide sequences

Pfam:

Protein family

KOG/COG:

Clusters of orthologous groups of proteins

Swiss-Prot:

A manually annotated and reviewed protein sequence database

KO:

KEGG ortholog database

GO:

Gene ontology

SD:

Standard deviation

qRT-PCR:

Quantitative real-time PCR

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Funding

This research was funded by the National Natural Science Foundation of China (NSFC) (31860560), the Jiangxi Natural Science Foundation (20224BAB205027), and Jiangxi Modern Agricultural Industry Technology System Construction Project (JXARS-16). The NSFC’s role in the study was to financially support the investigator and students who designed and performed experiments, collected and analyzed the data, and wrote the manuscript.

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Authors and Affiliations

  1. College of Agronomy, Jiangxi Agricultural University, Nanchang, 330045, Jiangxi, China

    L. I. Meng, S. I. Shucheng, W. U. Caijun & XIAO Xufeng

  2. Department of Animal Science, Jiangxi Biotech Vocational College, 1636 Liantang N Blvd, Nanchang County, Nanchang, 330200, Jiangxi, China

    Zhang Ming

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  1. L. I. Meng
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Contributions

X.X. designed the research. L.M., Z.M., S.S., and W.C. carried out the experiments. X.X., L.M., and S.S. analyzed the data. X.X., L.M., and Z.M. wrote the manuscript. All authors have read and approved the manuscript.

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Correspondence to XIAO Xufeng.

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Meng, L.I., Shucheng, S.I., Ming, Z. et al. Cd-Induced Autophagy Responses in Pakchoi as Revealed by Transcriptome Analysis. Plant Mol Biol Rep 42, 165–182 (2024). https://doi.org/10.1007/s11105-023-01403-8

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