Light induced accumulation of a polyubiquitin-encoding RNA in plumules
of five day seedlings
--Robert A. Bouchard, Priti Krishna and David B. Walden
It has previously been shown that RNA accumulation from specific members of the gene families of the small heat-shock protein (shsp) genes and the hsp83 genes can also be induced by light in the cotyledons of Pharbitis nil (Krishna, Felsheim, Larkin and Das, Plant Physiol. 100: 1772-1779, 1992; Felsheim and Das, Plant Physiol. 100: 1764-1771, 1992). We have now detected light-induced accumulation of RNA detected by a gene-specific probe for the maize polyubiquitin encoding gene uwo6 (MubC1), already shown to be a heat-shock gene, in plumules of five-day, dark-grown maize seedlings.
Seeds were surface-sterilized and the seedlings were grown in the dark on sterile wet filter paper in foil-wrapped pans in a 25 C incubator for five days. On day five, sampling was done as follows: 0-time control seedling plumules were removed and immediately placed in liquid nitrogen; heat shock seedlings were placed at 41 C in the dark for 1 hour, then plumules were harvested and placed in liquid nitrogen; pans containing time course (30, 60, 105, and 150 minute) seedlings were unwrapped and covered with transparent plastic, then returned to the 25 C incubator for the designated times with the lights on, after which plumules were harvested and placed in liquid nitrogen. Whole-cell RNA was prepared and RNA-dots were probed with a PCR-generated fragment specific for maize polyubiquitin clone MubC1. As previously reported (Liu, Walden, and Atkinson, MNL 67:72, 1993), RNA detected by this probe is up-regulated under heat shock; in the present experiments, accumulation under heat shock was over 300% of the level seen in control plumules. RNA up-regulation to 140% of the level found in the control was detectable after 30 minutes of light-induction, and RNA accumulation reached a level comparable to that seen under heat shock by 150 minutes. In contrast, little or no detectable light-induced accumulation was observed using probes specific for transcripts of two maize small heat-shock genes: uwo9 (Mhsp18-1) and uwo10 (Mhsp18-3), ruling out a general stress response. We are currently examining light-induced accumulation of RNAs for the additional maize ubiquitin genes and other heat-shock genes for which we have specific probes in plumule and radicle RNA.
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