Developmental regulation of embryonic genes in plants

Abstract

Somatic embryogenesis from cultured carrot cells progresses through successive morphogenetic stages termed globular, heart, and torpedo. To understand the molecular mechanisms underlying plant embryogenesis, we isolated two genes differentially expressed during embryo development. The expression of these two genes is associated with heart-stage embryogenesis. By altering the culture conditions and examining their expressions in a developmental variant cell line, we found that these genes were controlled by the developmental program of embryogenesis and were not directly regulated by 2,4-dichlorophenoxyacetic acid, the growth regulator that promotes unorganized growth of cultured cells and suppresses embryo morphogenesis. These genes are also expressed in carrot zygotic embryos but not in seedlings or mature plants.Embryogenesis involves the commitment of cells to an embryogenic fate and the expression of the embryogenic program to produce multicellular forms. It is presumed that morphogenesis is specified by the selective expression of gene sets in a sequential manner (1). As a model system to identify genes participating in plant embryogenesis and to study their mechanisms of gene regulation, we chose carrot somatic embryogenesis. Cultured carrot cells proliferate as unorganized cell clusters in a chemically defined medium supplemented with an auxin, 2,4-dichlorophenoxyacetic acid. Upon removal of 2,4-dichlorophenoxyacetic acid, these cell clusters readily form embryos that progress from globular to heart and to torpedo stages (2, 3). It is generally believed that the cultured carrot cells possess embryogenic potential, but the execution of embryo morphogenesis is suppressed by 2,4-dichlorophenoxyacetic acid (4).The onset of embryogenesis involves new protein synthesis (5-7) that is followed by an organized pattern of rapid cell division (8), resulting in a globular-shaped structure. The development of the heart stage involves axis formation and polarized growth resulting in a structure with bilateral symmetry. The heart-stage embryos elongate to produce torpedostage embryos that consist of initial cells for shoot and root meristems (9, 10) and three tissue layers, the protoderm, the cortex, and the procambium.Because early plant embryogenesis does not involve changes in the levels of abundant gene products (5), it has been difficult to isolate differentially expressed genes by conventional methods (11). We, therefore, used a combined immunoadsorption and epitope selection method to isolate cDNA clones corresponding to genes preferentially expressed during embryogenesis (12). This paper describes the temporal expression of these genes at the mRNA and protein levels. (17) with 2,4-dichlorophenoxyacetic acid at 0.1 mg/liter. WOiC and HA were subcultured at a cell density of 8 x 105 cells per ml, and 10-day-old cultures were used to produce somatic embryos as follows. Cultures of WOO1C and HA were filtered through a 500-,um mesh nylon screen, and the filtrates were pelleted by low-speed centrifugation and...

Interaction of nuclear factors with upstream sequences of a lipid body membrane protein gene from carrot.

“…The transcription of DC 59 is under strict developmental control, and both mRNA and protein are detected only in embryonic tissues (Borkird et al, 1988). Structural analysis showed that the gene was composed of a single exon resulting in an mRNA and a polypeptide with sizes that are in good correlation with previously published results (Choi et al, 1987).…”

“…However, the following three results suggest that they may play a role in the coordination of DC 59 gene expression: (1) The level of the C1 complex was correlated with the accumulation of DC 59 mRNA in callus and in embryos. DC 59 is expressed, although at Tower levels, in callus (Borkird et al, 1988). (2) Three of the four complexes could be detected only in embryos where the gene is highly active.…”

“…To determine the mechanism of ABA-regulated RNA accumulation, we extracted RNA from young somatic embryos (8 days old). The expression of the DC 59 gene is very low in young somatic embryos (Borkird et al, 1988), but the addition of ABA had a dramatic effect ( Figure 6B). An increase of 50-fold to 100-fold of DC 59 mRNA accumulation was observed in 15 min (lane 2).…”

“…We have isolated cDNA clones that detect embryo-specific RNAs to investigate embryonic gene regulation (Choi et al, 1987). The expression patterns of the embryo-specific RNAs are similar during somatic and zygotic embryogenesis (Borkird et al, 1988). In this paper, we report the molecular characterization of a gene, DC 59, that corresponds to one of the cDNA clones, cDNA 59.…”

Expression of DC8 is associated with, but not dependent on embryogenesis

“…Some ABA-inducible LEA genes are inducible only in cells formed in the seed [ 14,32], while others appear to lack seed specificity [30,38]. Carrot LEA genes that encode hydrophilic proteins, such as DC8, are expressed in carrot zygotic embryo and endosperm tissues, in somatic embryos, and in callus [2,4,9,46]. The ABA treatment causes large increases in levels of mRNA and protein in cultured carrot cells, but ABA cannot induce DC8 expression in mature leaf or root tissues [14,18].…”