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Ice recrystallization inhibition proteins of perennial ryegrass enhance freezing tolerance

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Abstract

Ice recrystallization inhibition (IRI) proteins are thought to play an important role in conferring freezing tolerance in plants. Two genes encoding IRI proteins, LpIRI-a and LpIRI-b, were isolated from a relatively cold-tolerant perennial ryegrass cv. Caddyshack. Amino acid alignments among the IRI proteins revealed the presence of conserved repetitive IRI-domain motifs (NxVxxG/NxVxG) in both proteins. Quantitative reverse transcriptase PCR (qRT-PCR) analysis indicated that LpIRI-a was up-regulated approximately 40-fold while LpIRI-b was up-regulated sevenfold after just 1 h of cold acclimation, and by 7 days of cold acclimation the transcripts had increased 8,000-fold for LpIRI-a and 1,000-fold for LpIRI-b. Overexpression of either LpIRI-a or LpIRI-b gene in Arabidopsis increased survival rates of the seedlings following a freezing test under both cold-acclimated and nonacclimated conditions. For example, without cold acclimation a −4°C treatment reduced the wild type’s survival rate to an average of 73%, but resulted in survival rates of 85–100% for four transgenic lines. With cold acclimation, a −12°C treatment reduced the wild type’s survival rate to an average of 38.7%, while it resulted in a survival rate of 51–78.5% for transgenic lines. After cold acclimation, transgenic Arabidopsis plants overexpressing either LpIRI-a or LpIRI-b gene exhibited a consistent reduction in freezing-induced ion leakage at −8, −9, and −10°C. Furthermore, the induced expression of the LpIRI-a and LpIRI-b proteins in transgenic E. coli enhanced the freezing tolerance in host cells. Our results suggest that IRI proteins play an important role in freezing tolerance in plants.

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Abbreviations

AFP:

Antifreeze proteins

CA:

Cold-acclimated

Ct:

Threshold cycle

DHFR:

Dihydrofolate reductase

EV:

Empty vector

G:

Glycine

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

IRI:

Ice recrystallization inhibition

Lp :

Lolium perenne

LRR:

Leucine-rich repeat

N:

Asparagine

NA:

Nonacclimated

ORF:

Open reading frame

qRT-PCR:

Quantitative reverse transcriptase PCR

Ta :

Triticum aestivum

V:

Valine

WT:

Wild type

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Acknowledgments

This project was supported by grants to S. F. by the United States Golf Association and the Iowa Turfgrass Institute. We wish to thank The Jacklin Seed for providing the Caddyshack perennial ryegrass seed.

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Authors and Affiliations

  1. Department of Horticulture, Iowa State University, Ames, IA, 50011-1100, USA

    Chunzhen Zhang, Shui-zhang Fei, Rajeev Arora & David J. Hannapel

  2. Interdepartmental Graduate Program in Plant Biology, Iowa State University, Ames, IA, 50011-1100, USA

    Shui-zhang Fei, Rajeev Arora & David J. Hannapel

  3. Interdepartmental Graduate Program in Genetics, Iowa State University, Ames, IA, 50011-1100, USA

    Shui-zhang Fei & David J. Hannapel

Authors
  1. Chunzhen Zhang
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  2. Shui-zhang Fei
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  3. Rajeev Arora
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  4. David J. Hannapel
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Correspondence to Shui-zhang Fei.

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Zhang, C., Fei, Sz., Arora, R. et al. Ice recrystallization inhibition proteins of perennial ryegrass enhance freezing tolerance. Planta 232, 155–164 (2010). https://doi.org/10.1007/s00425-010-1163-4

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