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Autophagy-related gene, TdAtg8, in wild emmer wheat plays a role in drought and osmotic stress response

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Abstract

An autophagy-related gene Atg8 was cloned for the first time from wild emmer wheat, named as TdAtg8, and its role on autophagy under abiotic stress conditions was investigated. Examination of TdAtg8 expression patterns indicated that Atg8 expression was strongly upregulated under drought stress, especially in the roots when compared to leaves. LysoTracker® red marker, utilized to observe autophagosomes, revealed that autophagy is constitutively active in Triticum dicoccoides. Moreover, autophagy was determined to be induced in plants exposed to osmotic stress when compared to plants grown under normal conditions. Functional studies were executed in yeast to confirm that the TdATG8 protein is functional, and showed that the TdAtg8 gene complements the atg8∆::kan MX yeast mutant strain grown under nitrogen deficiency. For further functional analysis, TdATG8 protein was expressed in yeast and analyzed using Western immunoblotting. Atg8-silenced plants were exposed to drought stress and chlorophyll and malondialdehyde (MDA) content measurements demonstrated that Atg8 plays a key role on drought stress tolerance. In addition, Atg8-silenced plants exposed to osmotic stress were found to have decreased Atg8 expression level in comparison to controls. Hence, Atg8 is a positive regulator in osmotic and drought stress response.

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Abbreviations

API:

Aminopeptidase I

BSMV:

Barley stripe mosaic virus

CDS:

Coding sequence

CVT:

Cytoplasm to vacuole targeting

Dpi:

Days post-inoculation

ECL:

Enhanced chemiluminescence

GAL4 AD:

GAL4 activation domain

mAPI:

Mature API

MDA:

Malondialdehyde

ORF:

Open reading frame

PDS:

Phytoene desaturase

PE:

Phosphatidylethanolamine

PEG:

Polyethylene glycol

prAPI:

Precursor API

qRT-PCR:

Quantitative real-time PCR

RT-PCR:

Reverse-transcriptase PCR

SPAD:

Soil plant analysis development

VIGS:

Virus-induced gene silencing

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Acknowledgments

Yeast Atg8 mutant strain (BY4741, atg8Δ::kanMX, MATa; his3Δ1; leu2Δ0; met15Δ0; ura3ΔO) and pRS316 plasmid with yeast Atg8 gene were kindly provided by Nakatogawa laboratory. Polyclonal anti-API antibody and Arabidopsis thaliana anti-Atg8a antibody used in this study were kindly provided by Dr. Dan Klionsky and Dr. Richard D. Vierstra, respectively. BSMV vectors used in this study were obtained from Dr. S Scofield (USDA-Agricultural Research Service). Nullisomic–tetrasomic wheat lines were kindly provided by Prof. Bikram Gill from Kansas State University. G.K. is recipient of Yousef Jameel Scholarship for PhD studies. This project was partially supported by EU-FP6 COST Action, TUBITAK (The Scientific and Technological Research Council of Turkey).

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Authors and Affiliations

  1. Biological Sciences and Bioengineering Program, Sabanci University, Tuzla, Istanbul, Turkey

    Duygu Kuzuoglu-Ozturk, Ozge Cebeci Yalcinkaya, Bala Ani Akpinar, Geraldine Mitou, Gozde Korkmaz, Devrim Gozuacik & Hikmet Budak

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  1. Duygu Kuzuoglu-Ozturk
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  2. Ozge Cebeci Yalcinkaya
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  3. Bala Ani Akpinar
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  4. Geraldine Mitou
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  7. Hikmet Budak
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Correspondence to Hikmet Budak.

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The authors D. Kuzuoglu-Ozturk and O. Cebeci Yalcinkaya are equally contributed to this work.

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Kuzuoglu-Ozturk, D., Cebeci Yalcinkaya, O., Akpinar, B.A. et al. Autophagy-related gene, TdAtg8, in wild emmer wheat plays a role in drought and osmotic stress response. Planta 236, 1081–1092 (2012). https://doi.org/10.1007/s00425-012-1657-3

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