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Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (3): 472-480.doi: 10.3724/SP.J.1006.2021.03027


Functional analysis of plasma membrane intrinsic protein ZmPIP1;1 involved in drought tolerance and photosynthesis in maize

ZHOU Lian, LIU Chao-Xian, XIONG Yu-Han, ZHOU Jing, CAI Yi-Lin*()   

  1. Maize Research Institute / Academy of Agricultural Sciences / State Cultivation Base of Crop Stress Biology for Southern Mountainous Land, Southwest University, Chongqing 400715, China
  • Received:2020-05-14 Accepted:2020-09-13 Online:2021-03-12 Published:2020-10-06
  • Contact: CAI Yi-Lin E-mail:caiyilin1789@163.com
  • Supported by:
    National Natural Science Foundation of China(31601312)


Plasma membrane intrinsic protein (PIP) is one of the main subfamily of aquaporin regulates diverse physiology functions during plant growth and development. In previous research, the expression of ZmPIP1;1 was induced by osmosis or salt stress. However, the biological function of ZmPIP1;1 was still unclear in maize. In this study, ZmPIP1;1 overexpressed transgenic plants were obtained and exhibited less water loss rate and enhanced drought tolerance compared to wild type. Transcriptome sequencing indicated significant changes in the expression levels of genes involved in ABA biosynthesis and its signaling pathways. Photosynthetic activity, kernel width and kernel weight was increased in ZmPIP1;1 overexpression maize, but not growth under normal condition compared with wild type. Moreover, interaction between ZmPIP1;1 and ZmPIP2;6 was observed by bimolecular fluorescence complementation (BiFC) experiment, resulting in re-localized on plasma membrane and chloroplast in maize mesophyll protoplast. Our study laid an important foundation for understanding the molecular mechanism of ZmPIP1;1, and provided a new method of molecular breeding for high photosynthetic efficiency.

Key words: maize, ZmPIP, drought stress, photosynthesis, re-localization

Fig. 1

Identification of ZmPIP1;1 overexpressed transgenic maize A: schematic illustration of T-DNA sequence of ZmPIP1;1 overexpression vector. B: PCR detection of T1 generation of ZmPIP1;1 overexpression transgenic lines; 1-5: five independent transgenic lines; MK: DNA marker; PC: positive control; WT: wild type. C: qRT-PCR analysis of four representative ZmPIP1;1 overexpressed transgenic lines."

Fig. 2

Phenotypes and water loss rate of ZmPIP1;1 overexpressed transgenic lines under drought treatment A: phenotype of WT and ZmPIP1;1 overexpressed transgenic maize under drought stress treatment for 15 days. B: relative shoot biomass of different genotypes grown in soil with the indicated treatments, compared with normal conditions. Values are means of four biological replicates with error bars indicating standard derivations (SD). C: determination of water loss from detached leaves of WT and ZmPIP1;1 overexpressed transgenic line. Values followed by * are significantly different at P < 0.01."

Fig. 3

Clusters of orthologous groups (COG) function classification of consensus sequences T and G columns represent the functions of “signal transduction mechanisms” and “carbohydrate transport and metabolism”, respectively."

Fig. 4

Relative expression level of key genes involved in ABA biosynthesis and its signaling pathway The relative expression level of (A) ZmNCED9-1, ZmNCED9-2, ZmABA1, and ZmAAO3 involved in ABA biosynthesis (B) ZmABF1, ZmPYL2, ZmABI5, and ZmSnRK3 involved in ABA signaling pathway in leaves of the WT and ZmPIP1;1 overexpressed lines under normal and drought stress treatments. Data are means of three biological replicates with error bars indicating standard deviations (SD). Values followed by asterisk are significantly different at P < 0.01. ZmACT1 was used as the internal control."

Fig. 5

AN and Ci of WT and ZmPIP1;1 overexpressing transgenic maize plants All data are means ± standard deviations (SD) (n = 4). Values followed by asterisk are significantly different at P < 0.01."

Fig. 6

Phenotypic characterization of ZmPIP1;1 overexpressed transgenic maize kernel Phenotypic determination (A) and photograph (B) of ZmPIP1;1 overexpressed transgenic maize kernel. Values followed by asterisk are significantly different at P < 0.01. Bar = 1 cm."

Fig. 7

Observation of BiFC experiment of ZmPIP1;1 and ZmPIP2;6 in maize protoplasts pCAMBIA-ZmPIP2;6-GFP, pDOE-ZmPIP1;1-ZmPIP2;6, and pDOE- ZmPIP2;6-ZmPIP1;1 were co-localized with a mCherry-labeled plasma membrane marker (mCherry-PM; CD3-1007). Bar = 20 μm."

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