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Cysteine Protease 51 (CP51), an anther-specific cysteine protease gene, is essential for pollen exine formation in Arabidopsis

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Abstract

Cysteine proteases play important roles in intracellular protein degradation, programmed cell death and responses to environmental stimuli in plant. Although subclassification and biochemical analysis of major plant papain-like cysteine proteases (PLCPs) have been studied, the biological function of many PLCPs remained unknown. In this study, we identified a PLCP gene Cysteine Protease 51 (CP51) which participates in exine formation and anther development in Arabidopsis thaliana. Promoter-GUS fusion detection showed its specific expression in anthers at stages 9–12. RNA interference (RNAi) transgenic plants with reduced CP51 transcriptional levels exhibited a male sterile phenotype with aborted microspores, shortened siliques and fewer or no seeds. Cytological analysis indicated that pollen abortion occurred due to defective pollen exine and the tapetum degraded earlier during the transition from the uninucleated stage to the binucleated stage. Scanning electron microscopy demonstrated that aborted microspores lacked complete or normal reticulate exine, and the intine membrane was extruded in the pollens of CP51-RNAi plants. Transmission electron microscopy further revealed that the tapetum degeneration was initiated early and that normal tectum connections to the bacula were missing in anthers of CP51-RNAi plants. Taken together, these results suggested that CP51 critically mediates tapetum stability and pollen exine formation.

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Abbreviations

PLCPs:

Papain-like cysteine proteases

qRT-PCR:

Quantitative real-time polymerase chain reaction

PCD:

Programmed cell death

WT:

Wild-type

GUS:

β-Glucuronidase

Kan:

Kanamycin

NPTII :

Neomycin phosphotransferase

HPT :

Hygromycin B phosphotransferase

DAPI:

4′,6′-Diamidino-2-phenylindole

ORF:

Open reading frame

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Acknowledgments

We thank Jianbo Cao (Huazhong Agricultural University, China) for help with scanning electron microscopy and transmission electron microscope. This work was supported by the National Basic Research Program of China (2011CB109305), the State High-Tech Development Plan of China (2013AA102602), Commonweal Specialized Research Fund of China Agriculture (201103016), Innovation Program of Wuhan Academy of Agricultural Science and Technology (CX201242) and the Hubei Agriculture Science and Technology Innovation Center.

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Authors and Affiliations

  1. Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, No. 2 Xudong 2nd Road, Wuhan, 430062, China

    Yongxue Yang, Caihua Dong, Jingyin Yu, Chaobo Tong, Junyan Huang & Shengyi Liu

  2. Wuhan Institute of Agricultural Science, Wuhan Academy of Agricultural Science and Technology, Wuhan, 430345, China

    Yongxue Yang

  3. Key Laboratory of Oil Crops in Huanghuaihai Plains, Ministry of Agriculture, P China, Henan Provincial Key Laboratory for Oil Crops Improvement, Industrial Crops Research Institute, Henan Academy of Agricultural Sciences, Zhengzhou, 450002, China

    Lei Shi

  4. College of Life Science, South-Central University for Nationalities, Wuhan, 430074, China

    Zhenbo Li

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  1. Yongxue Yang
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Correspondence to Shengyi Liu.

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Yongxue Yang and Caihua Dong have contributed equally to this work.

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Yang, Y., Dong, C., Yu, J. et al. Cysteine Protease 51 (CP51), an anther-specific cysteine protease gene, is essential for pollen exine formation in Arabidopsis . Plant Cell Tiss Organ Cult 119, 383–397 (2014). https://doi.org/10.1007/s11240-014-0542-0

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