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A proteomic study of cysteine protease induced cell death in anthers of male sterile tobacco transgenic plants

Abstract

Manifestation of male sterility in plants is an important requirement for hybrid seed production. Tapetum cell layer of anther is a primary target for genetic manipulation for male sterility. In our previous report, the targeted expression of Arachis cysteine protease in tapetum led to premature degeneration of tapetal layer that resulted in complete male sterility in transgenic tobacco plants. To correlate cysteine protease mediated cell death of tapetum, transmission electron microscopy (TEM) and proteomic pattern of anthers of cysteine protease induced male sterile plant were compared with the untransformed control plant. TEM study revealed the abnormal growth of tapetal cells exhibiting excessive vacuolization that synchronized with irregular exine wall formation of the microspores. In anther proteome, a total 250 protein spots were detected that were reproducible and exhibited similar distribution pattern. Further, anther proteome of male sterile plant showed the significant upregulation (≥ 1.5) of 56 protein spots. Using Mass spectroscopy (MALDI TOF/TOF), we have identified 14 protein spots that were involved in several processes such as energy metabolism, protein synthesis, plastid protein, lipid metabolism, and cell wall assembly. Upregulation of patatin-like protein-2 homolog, carboxylesterase 17 and dicer like protein-4 in male sterile anthers that have been demonstrated to induce cell death, suggesting that cysteine protease mediated premature tapetal cell death might involve the lipid peroxidation pathway in coordination with gene silencing mechanism.

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Abbreviations

AdCP :

Arachis cysteine protease

TEM:

Transmission electron microscopy

DTT:

Dithiothreitol

IEF:

Isoelectric focusing

PI:

Isoelectric point

PCD:

Programmed cell death

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Acknowledgements

The authors are grateful to Council of Scientific and Industrial Research, Government of India for a Research Grant [38 (1393/EMR-II)] to one of the authors (PBK), DST-FIST, UGC-SAP, Government of India, for the facilities provided to the Department of Plant Sciences, University of Hyderabad.

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Correspondence to Pawan Shukla or Pulugurtha Bharadwaja Kirti.

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Shukla, P., Gautam, R., Singh, N.K. et al. A proteomic study of cysteine protease induced cell death in anthers of male sterile tobacco transgenic plants. Physiol Mol Biol Plants 25, 1073–1082 (2019). https://doi.org/10.1007/s12298-019-00642-y

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Keywords

  • Cysteine protease
  • Anther proteome
  • Cell death
  • Male sterility
  • Patatin
  • Dicer like protein