Differential display-mediated isolation of a genomic sequence for a putative mitochondrial LMW HSP specifically expressed in condition of induced thermotolerance in Arabidopsis thaliana (L.) Heynh.

Differential display-mediated isolation of a genomic sequence for a putative mitochondrial LMW... Plants of Arabidopsis thaliana pre-treated at 37 °C for 2 h can survive an otherwise lethal heat shock at 45 °C. Differential display reverse transcriptase-PCR (DDRT-PCR) was utilized to clone DNA fragments corresponding to mRNAs specifically expressed in conditions of induced thermotolerance or of expression of thermotolerance. One of these DDRT-PCR fragments enabled the isolation of a genomic clone pAt1.3EX, containing the sequence Athsp23.5, the gene for a low-molecular-weight (LMW) heat shock protein (HSP), AtHSP23.5. Athsp23.5 is low- or single-copy in the Arabidopsis genome and its open reading frame is interrupted by a 137 bp intron. Analysis of the sequence suggests AtHSP23.5 is targeted to the mitochondrion. The steady-state level of the AtHSP23.5 mRNA varied significantly according to the heat treatment, increasing on heat shock (transfer from 22 °C to 37 °C), with a further increase during expression of thermotolerance (transfer from 22 °C to 37 °C and then to 45 °C). Expression was low after an abrupt stress (from 22 °C to 45 °C). This behaviour was different from that observed for other LMW HSP mRNAs that were present at high level at 37 °C, but did not increase significantly in condition of expression of thermotolerance, and reached a considerable steady-state level also during the abrupt stress at 45 °C. The retrotranscription of AtHSP23.5 mRNA followed by amplification with two primers encompassing the intron allowed for the isolation of an almost full-length cDNA sequence. The sequence analysis of the two cDNAs obtained from condition 22 °C→37 °C and condition 22 °C→37 °C→45 °C suggested that in both cases the intron had been correctly spliced. The importance of correct intron splicing in survival at high temperatures and the role of mitochondrial HSP in induction and expression of thermotolerance are discussed. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Plant Molecular Biology Springer Journals

Differential display-mediated isolation of a genomic sequence for a putative mitochondrial LMW HSP specifically expressed in condition of induced thermotolerance in Arabidopsis thaliana (L.) Heynh.

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Publisher
Springer Journals
Copyright
Copyright © 1997 by Kluwer Academic Publishers
Subject
Life Sciences; Biochemistry, general; Plant Sciences; Plant Pathology
ISSN
0167-4412
eISSN
1573-5028
D.O.I.
10.1023/A:1005824314022
Publisher site
See Article on Publisher Site

Abstract

Plants of Arabidopsis thaliana pre-treated at 37 °C for 2 h can survive an otherwise lethal heat shock at 45 °C. Differential display reverse transcriptase-PCR (DDRT-PCR) was utilized to clone DNA fragments corresponding to mRNAs specifically expressed in conditions of induced thermotolerance or of expression of thermotolerance. One of these DDRT-PCR fragments enabled the isolation of a genomic clone pAt1.3EX, containing the sequence Athsp23.5, the gene for a low-molecular-weight (LMW) heat shock protein (HSP), AtHSP23.5. Athsp23.5 is low- or single-copy in the Arabidopsis genome and its open reading frame is interrupted by a 137 bp intron. Analysis of the sequence suggests AtHSP23.5 is targeted to the mitochondrion. The steady-state level of the AtHSP23.5 mRNA varied significantly according to the heat treatment, increasing on heat shock (transfer from 22 °C to 37 °C), with a further increase during expression of thermotolerance (transfer from 22 °C to 37 °C and then to 45 °C). Expression was low after an abrupt stress (from 22 °C to 45 °C). This behaviour was different from that observed for other LMW HSP mRNAs that were present at high level at 37 °C, but did not increase significantly in condition of expression of thermotolerance, and reached a considerable steady-state level also during the abrupt stress at 45 °C. The retrotranscription of AtHSP23.5 mRNA followed by amplification with two primers encompassing the intron allowed for the isolation of an almost full-length cDNA sequence. The sequence analysis of the two cDNAs obtained from condition 22 °C→37 °C and condition 22 °C→37 °C→45 °C suggested that in both cases the intron had been correctly spliced. The importance of correct intron splicing in survival at high temperatures and the role of mitochondrial HSP in induction and expression of thermotolerance are discussed.

Journal

Plant Molecular BiologySpringer Journals

Published: Sep 29, 2004

References

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