Scientia Agricultura Sinica ›› 2016, Vol. 49 ›› Issue (3): 429-442.doi: 10.3864/j.issn.0578-1752.2016.03.003

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Responses of Endoplasmic Reticulum Stress-Related Genes in Maize Embryo to Artificial Aging Treatment

CAO Guang-can, LIN Yi-xin, XUE Mei-zhen, XING Lu-man, LÜ Wei-zeng, YANG Wei-fei, CHEN Jun-ying

 
  

  1. College of Agronomy, Henan Agricultural University, Zhengzhou 450002
  • Received:2015-07-23 Online:2016-02-01 Published:2016-02-01

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Abstract: 【Objective】 Endoplasmic reticulum (ER) stress and unfolded protein response (UPR) are involved in plant responses to environmental stresses. However, the expression of ER stress-related genes during maize seed aging has not been reported. In this study, the expression of ER stress-related genes during maize seed aging was investigated by Digital Gene Expression Profile (DGE) to provide theoretical support for clarifying the molecular mechanism of seed deterioration.【Method】 Hybrid maize (Zea mays L.) cultivar Zhengdan 958 seeds were used as experimental material and treated by artificial aging treatment (45℃, 100% relative humidity). DGE analysis was carried out on the Illumina Hiseq 2000 platform using total RNA extracted from 3 d artificial aging treatment and the untreated embryos (CK) of maize seeds. The reads with adaptor and ambiguous sequences, and the low-quality reads were filtered out to obtain the high quality clean reads. Clean reads were mapped to the maize reference genome and genes database using SOAPaligner/SOAP2. The gene expression level was calculated by the RPKM (Reads Per kb per million reads) method. A combination of FDR ＜0.001 and the absolute value of |log2 ratio (T/CK)|≥1 was used as the threshold to determine the significance of gene expression difference. All differentially expressed genes (DEGs) were assigned to the pathways in KEGG (Kyoto Encyclopedia of Genes and Genomes) database and searched for the differentially expressed genes related to ER stress. Quantitative real-time PCR was performed to analyze the expression patterns of ER stress-related genes in the different artificial aging times.【Result】 Analysis of the DGE revealed that 104 DEGs were relevant to the protein processing in ER during the process of artificial aging treatment. A total of 97 DEGs related to ER stress including 81 and 16 genes respectively up- and down-regulated were screened out. The expression levels of ER stress marker BiP gene, as well as ER chaperones, such as CRT, CNT, GRP94 genes were considered to be significantly up-regulated. In particular, 83 DEGs were involved in ER-associated degradation (ERAD) pathway, including 70 up-regulated and 13 down-regulated DEGs. Among them, gene encoding EDEM (ER degradation enhancing mannosidase I-like protein) which is a rate-limiting enzyme of ERAD pathway was down-regulated. Genes involved in protein ubiquitination were altered in expression, including E2 ubiquitin-conjugating enzyme UbcH5, E3 ubiquitin-ligases Hrd1 and Doa10. The results of qRT-PCR analysis validated the diversity and complexity of ER stress-related genes expression in different artificial aging times.【Conclusion】 Artificial aging treatment can cause endoplasmic reticulum stress in maize embryo. The cell can respond to ER stress by inducing up-regulation of ER chaperones genes and activation of the ERAD pathway. Inhibition of ERAD pathway resulted in the accumulation of misfolded proteins in the ER with these leading to further cell damage and subsequently accelerating the loss of seed vigor.

Key words: maize, seed aging, differential expressed genes, ER stress, ERAD

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