We have isolated a homeobox-containing gene from Arabidopsis thaliana using a degenerate oligonucleotide probe corresponding to the most conserved region of the homeodomain. This strategy has been used previously to isolate homeobox-containing genes from Caenorhabditis, and recently from A. thaliana. The Arabidopsis genes have an unusual structure in that they have a leucine zipper motif adjacent to the carboxy terminal region of the homeo domain, a feature not found in homeobox-containing genes isolated from animals. We report the isolation and primary structure of a new member of this Arabidopsis homeobox-leucine zipper gene family. This new member has the homeodomain and leucine-zipper motif similar to the two genes previously identified, but differs from these genes in the part corresponding to the carboxy terminus of the polypeptide, as well as in size and isoelectric point of the protein.

Genes specifically induced during somatic embryogenesis may play key roles in plant embryo develop-ment. An antiserum against an extract of carrot somatic embryos revealed a few rare antigens induced at the onset of embryogenesis. Through differential immunoadsorption techniques, we purified antibodies against the embryo-specific antigens and probed a phage X gtll library of cDNA from carrot somatic embryos. This paper describes three distinct cDNA clones that hybridize to embryo-specific RNAs. Monospecific antibodies, purified by affinity to the recombinant phage fusion proteins, confirm that the doned cDNAs encode unique embryo-specific 0eptide antigens. One 50NkDa protein correlates with embryogenic ability in cultures of other plant species, incuding cereals.

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...

A recently discovered class of genes in Arabidopsis thaliana encode putative transcription factors which contain a homeodomain closely linked to a leucine zipper motif. We have previously reported on the cloning and cDNA sequence of one gene of this class, Athb-3. In this article we show this gene to be expressed predominantly in the cortex of the root and the stem. Using the Athb-3 clone as a probe we have isolated cDNA clones corresponding to three novel homeodomain-leucine zipper proteins. These clones, Athb-5, Athb-6 and Athb-7, hybridized to transcripts that were relatively abundant in the leaf, but also present in other vegetative organs, as well as in the flower. Only weak hybridization was observed to seed pod samples. These observations indicate that these Athb genes have major functions in the mature plant, and therefore, in contrast to homeobox genes in other eukaryotes and to the kn-1 gene in maize, are unlikely to function in the primary control of developmental processes during embryogenesis or organogenesis. The deduced amino acid sequences of Athb-5, Athb-6 and Athb-7 are highly similar to the previously isolated Athb-1, Athb-2 and Athb-3 in the homeodomain and leucine-zipper parts of the proteins, whereas the similarities to homeodomain proteins from other eukaryotes are limited. The Athb proteins thus constitute a new and well defined class of homeodomain proteins, apparently unique to plants.