Cloning and Characterization of <em>TPE4A</em>, a Thiol-Protease Gene Induced during Ovary Senescence and Seed Germination in Pea^{<a href="#FN1" rid="FN1" class=" fn">1</a>}

Abstract

Senescence is the last step of plant development, leading to the death of a plant tissue or organ or the whole plant. There is evidence to indicate that senescence is a genetically controlled process (Gan and Amasino, 1997; Noodén et al., 1997). One of the major events taking place during the senescence process is the ordered degradation of cell constituents, and the degradation of proteins is a characteristic of senescence. Protease genes have been cloned from several senescent plants (Jones et al., 1995; Smart et al., 1995; Drake et al., 1996), and thiol-proteases are the most common proteolytic enzymes induced in senescent plant cells (Granell et al., 1998).

The unpollinated pea (Pisum sativum) ovary is a convenient system in which to study the natural senescence of reproductive organs in plants. When pollination is prevented, the ovary stops growing about 3 d after anthesis; natural senescence then starts, leading to ovary death and abscission within 2 or 3 d (Carbonell and García-Martínez, 1980). This senescence process is accompanied by the loss of sensitivity to GA₃ treatments (García-Martínez and Carbonell, 1980) and by an increase in proteolytic activity at neutral pH (Carbonell and García-Martínez, 1985; Carrasco and Carbonell, 1988; Cercós et al., 1992). New proteases have been found associated with senescence in unpollinated pea ovaries (Granell et al., 1992; Cercós and Carbonell, 1993; Cercós et al., 1993). The senescence process is prevented when parthenocarpic fruit set is induced by treatment of the unpollinated ovaries with GA₃ and other plant-growth regulators (García-Martínez and Carbonell, 1980).

Like senescence, seed germination involves the degradation and remobilization of stored nutrients. Thiol-proteases also play a key role in this remobilization step (Granell et al., 1998). It is not known whether the same thiol-proteases are involved in senescence and germination or whether there are different thiol-proteases for each process. There are few reports addressing this question (Griffiths et al., 1997), and none of them reports a single-copy gene being expressed in both senescence and germination.

In this study, we isolated and characterized a cDNA clone encoding TPE4A, a thiol-protease induced during senescence of unpollinated pea ovaries. The induction of TPE4A transcription during senescence and its repression after GA₃ treatment were observed by northern-blot analysis and in situ hybridization. We also detected TPE4A transcription in other organs, including germinating seeds. We report evidence for a single-copy thiol-protease gene that is induced in both senescence and seed germination.

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