Novel Molecular Markers for Late Phases of the Growth Cycle of Arabidopsis thaliana Cell-Suspension Cultures Are Expressed during Organ Senescence

Abstract

In an Arabidopsis thaliana T87-C3 cell-suspension culture, entry into the growth-arrest phase is rapidly followed by a loss of cell viability. Three cDNA clones, SRGI, SRG2, and SRG3, corresponding to genes with transcripts that accumulate during these late phases, were isolated by the mRNA differential display method. Amino acid sequence analysis shows that the putative SRCl protein is a new member of the Fe(ll)/ascorbate oxidase superfamily, and that SRG2 codes for a protein with significant homology to P-glucosidases. Significantly, all three SRG genes are expressed in senescing organs of Arabidopsis plants. Two previously characterized genes, SAG2 and SAG4, induced during natural senescence in Arabidopsis, were also found to be expressed in cell-suspension cultures and have expression kinetics similar to those observed for the SRGl gene. Taken together these findings suggest that certain molecular events are common to both plant senescence and growth arrest in Arabidopsis cell suspensions. Both internucleosomal cleavage of nDNA and an apparent compaction of chromatin, two characteristic features of programmed cell death in animal cells, have been observed in Arabidopsis cell cultures at a stage corresponding to loss of cell viability. ~~ ~Cultured cells are now universally recognized as appropriate model systems with which to investigate the molecular responses of plants to stimuli that affect growth, such as nutrient availability (e.g. Roitsch et al., 1995) and plant growth factor requirements (e.g. Teyssendier de la Serve et al., 1985;Dominov et al., 1992;Lee and Chen, 1993). For example, limitations in phosphate concentration (Kodama et al., 1990;Kock et al., 1995; Malboobi and Lefebvre, 1995), nitrogen supply (Kodama et al., 1990), carbon source (Kodama et al., 1990;Sheu et al., 1994;Hsieh et al., 1995;Tseng et al., 1995), or hormone depletion (Crowell and Amasino, 1991) result in modifications in gene expression, leading to changes in the cellular mRNA pattern.After nutrient exhaustion, certain cell cultures undergo rapid growth arrest, followed by a dramatic loss of viabil- ity. We are currently studying these late phases of the cell culture growth cycle, which exhibit a number of morphological symptoms similar to those that occur in senescing plant tissues, which have recently been described in sugarstarved, cultured rice cells (Chen et al., 1994).As a first step toward characterizing the molecular events associated with growth arrest and viability loss, we have attempted to select genes with a relative expression that is enhanced at the end of the growth phase of the cell culture. For this purpose, we chose an Arabidopsis fkaliana cell line (Axelos et al., 1992) with a particularly short growth-arrest period. Transcripts that accumulated at the end of the growth phase of the culture were characterized by the mRNA differential display method (Liang and Pardee, 1992). Three cDNA clones were isolated and the expression patterns of the corresponding genes were analyzed both throughout the cell culture gr...

Functional genomics of the Dof transcription factor family genes in suspension-cultured cells of Arabidopsis thaliana

“…Among them, a unique cell line, namely, T87, which has photosynthetic ability under light irradiation (Axelos et al 1992), have been widely used for biochemical and molecular biological analyses with respect to metabolic regulation, circadian rhythm, and stress response (e.g. Callard et al 1996;Uno et al 2000;Takahashi et al 2001;Nakamichi et al 2004;Stolc et al 2005;Nakamura et al 2007). Thus, this culture system is suitable for functional genomics study of transcription factors in regulation of metabolic pathway.…”

Cell Death Processes during Expression of Hybrid Lethality in Interspecific F1 Hybrid between Nicotiana gossei Domin and Nicotiana tabacum

“…Thus, none of the features of apoptosis-like cell death are observed in the TE of zinnia, although it is a developmentally regulated form of PCD (Obara et al, 2001). On the other hand, typical apoptotic features, including chromatin condensation and DNA fragmentation into nucleosomes, were found in pollination-induced petal senescence of petunia (Petunia hybrida; Xu and Hanson, 2000), old cultured cells of Arabidopsis (Callard et al, 1996), aleurone layers of germinating barley (Hordeum vulgare) seed (Wang et al, 1998) and carpel senescence in pea (Pisum sativum; Orzaez and Granell, 1997). These findings indicate that PCD in plants has diverse features of expression.…”

“…DNAaddering is widely recognized in plant PCD (Callard et al, 1996;Wang et al, 1998;Xu and Hanson, 2000). In the interspecific hybrids of N. suaveolens ϫ N. tabacum and cultured cells of N. glutinosa ϫ N. repanda, a DNA ladder was detected Yamada et al, 2001b).…”

Culture senescence and abscisic acid induce saporin production in cultured roots of Saponaria officinalis

“…By contrast with other enzymatic activities, such as those of malate dehydrogenase and glyoxalase I, which mark precisely the metabolic state of the culture, saporin activity is specifically enhanced during the latestationary phase, suggesting that it might have a role in the arrest of culture growth. Interesting molecular parallels between growth arrest in cell cultures and physiological senescence of plant tissues have recently been reported (Callard et al, 1996), envisaging a partial coincidence of the molecular events associated with these phenomena. These results are consistent with previous observations on the higher level of dianthins present in old, compared with young, carnation leaves (Reisbig & Bruland, 1983), with the reported increase of RIP activity (on both ribosomal and DNA substrates) in senescent and stressed leaves of Hura crepitans and Phytolacca americana (Stirpe et al, 1996), with the appearance of a RIP activity in barley senescent leaves (Chaundry et al, 1994) and, finally, with the saltstress-induced expression of RIP-related activities in the ice plant Mesembryanthemum crystallinum (Rippmann et al, 1997).…”