Analysis of LE-ACS3, a 1-Aminocyclopropane-1-carboxylic Acid Synthase Gene Expressed during Flooding in the Roots of Tomato Plants

Abstract

The plant hormone ethylene is produced in response to a variety of environmental stresses. Previous work has shown that flooding or anaerobic stress in the roots of tomato plants caused an increase in the production of the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) in the roots, due to flooding-induced activity of ACC synthase (EC 4.4.1.14). RNA was extracted from roots and leaves of tomato plants flooded over a period of 48 h. Blot analysis of these RNAs hybridized with probes for four different ACC synthases revealed that the ACC synthase gene LE-ACS3 is rapidly induced in roots. LE-ACS2 is also induced, but at later times. The genomic clone for LE-ACS3 was isolated and sequenced. At all time points, the probe from the LE-ACS3 coding region hybridized to two bands in the RNA blots. Hybridization using the first and third introns of LE-ACS3 separately as probes indicate that flooding may inhibit processing of the LE-ACS3 transcript. Sequence homology analysis identified three putative cis-acting response elements in the promoter region, corresponding to the anaerobic response element from the maize adh1 promoter, the root-specific expression element from the cauliflower mosaic virus 35S promoter and a recognition element for chloroplast DNA binding factor I from the maize chloroplast ATP synthase promoter. The plant hormone ethylene is produced in response to a variety of environmental stresses. Previous work has shown that flooding or anaerobic stress in the roots of tomato plants caused an increase in the production of the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) in the roots, due to flooding-induced activity of ACC synthase (EC 4.4.1.14). RNA was extracted from roots and leaves of tomato plants flooded over a period of 48 h. Blot analysis of these RNAs hybridized with probes for four different ACC synthases revealed that the ACC synthase gene LE-ACS3 is rapidly induced in roots. LE-ACS2 is also induced, but at later times. The genomic clone for LE-ACS3 was isolated and sequenced. At all time points, the probe from the LE-ACS3 coding region hybridized to two bands in the RNA blots. Hybridization using the first and third introns of LE-ACS3 separately as probes indicate that flooding may inhibit processing of the LE-ACS3 transcript. Sequence homology analysis identified three putative cis-acting response elements in the promoter region, corresponding to the anaerobic response element from the maize adh1 promoter, the root-specific expression element from the cauliflower mosaic virus 35S promoter and a recognition element for chloroplast DNA binding factor I from the maize chloroplast ATP synthase promoter.