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Vol. 16, No. 12, pp. 1555-1567, June 15, 2002
Biocenter of the Goethe University, D-60439 Frankfurt am Main, Germany
We generated transgenic tomato plants with altered expression of
heat stress transcription factor HsfA1. Plants with 10-fold overexpression of HsfA1 (OE plants) were characterized by a single HsfA1 transgene cassette, whereas plants harboring a tandem
inverted repeat of the cassette showed cosuppression (CS plants) by
posttranscriptional silencing of the HsfA1 gene connected with
formation of small interfering RNAs. Under normal growth conditions,
major developmental parameters were similar for wild-type (WT), OE, and
CS plants. However, CS plants and fruits were extremely sensitive to
elevated temperatures, because heat stress-induced synthesis of
chaperones and Hsfs was strongly reduced or lacking. Despite the
complexity of the plant Hsf family with at least 17 members in tomato,
HsfA1 has a unique function as master regulator for induced
thermotolerance. Using transient reporter assays with mesophyll
protoplasts from WT tomato, we demonstrated that plasmid-encoded HsfA1
and HsfA2 were well expressed. However, in CS protoplasts the
cosuppression phenomenon was faithfully reproduced. Only transformation
with HsfA2 expression plasmid led to normal expression of the
transcription factor and reporter gene activation, whereas even high
amounts of HsfA1 expression plasmids were silenced. Thermotolerance in CS protoplasts was restored by plasmid-borne HsfA2, resulting in
expression of chaperones, thermoprotection of firefly luciferase, and
assembly of heat stress granules.
[Key Words: Posttranscriptional gene silencing; heat stress transcription factors; chaperones; siRNA; tomato; thermotolerance]
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