A novel class of plant proteins containing a homeodomain with a closely linked leucine zipper motif.

The homeobox, a 183 bp DNA sequence element, was originally identified as a region of sequence similarity between many Drosophila homeotic genes. The homeobox codes for a DNA-binding motif known as the homeodomain. Homeobox genes have been found in many animal species, including sea urchins, nematodes, frogs, mice and humans. To isolate homeobox-containing sequences from the plant Arabidopsis thaliana, a cDNA library was screened with a highly degenerate oligonucleotide corresponding to a conserved eight amino acid sequence from the helix-3 region of the homeodomain. Using this strategy two cDNA clones sharing homeobox-related sequences were identified. Interestingly, both of the cDNAs also contain a second element that potentially codes for a leucine zipper motif which is located immediately 3'' to the homeobox. The close proximity of these two domains suggests that the homeodomain-leucine zipper motif could, via dimerization of the leucine zippers, recognize dyad-symmetrical DNA sequences.     

Early steps in the evolution of multicellularity: deep structural and functional homologies among homeobox genes in sponges and higher metazoans

The sponge homeobox gene EmH-3 had not been attributed to any homeobox family. Comparative promoter and homeodomain sequence analyses suggest that it is related to the Hox11 gene, which belongs to the Tlx homeobox family. Hox11 is highly expressed in proliferating progenitor cells, but expression is downregulated during cell differentiation. Using reporter gene methodology, we monitored function of the sponge EmH-3 promoter transfected into human erythroleukemia K562 cells. These cells express the Tlx/Hox11 gene constitutively, and downregulate its expression upon differentiation. The same pattern of expression and downregulation was observed for the sponge reporter construct. We propose that Tlx/Hox11 genes have structural and functional homologies conserved in phylogenetically distant groups, that represent a deep homology in the regulation of cell proliferation, commitment and differentiation.     

A new homeobox-leucine zipper gene from Arabidopsis thaliana

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.     

<Emphasis Type="Italic">ASYMMETRIC LEAVES2-LIKE1</Emphasis>gene,a member of the AS2/LOB family,controls proximal–distal patterning in <Emphasis Type="Italic">Arabidopsis</Emphasis> petals

The formation and the development of the floral organs require an intercalate expression of organ-specific genes. At the same time, meristem-specific genes are repressed to complete the differentiation of the organs in the floral whorls. In an Arabidopsis activation tagging population, a mutant affected in inflorescence architecture was identified. This gain-of-function mutant, designateddownwards siliques1 (dsl1-D), has shorter internodes and the lateral organs such as flowers are bending downwards, similar to the loss-of-function brevipedicellus (bp) mutant. The affected gene in dsl1-D appeared to be ASYMMETRIC LEAVES2-LIKE1 (ASL1)/LATERAL ORGAN BOUNDARIESdomain gene 36 (LBD36), which is a member of the ASYMMETRIC LEAVES2 (AS2)/LATERAL ORGAN BOUNDARIES (LOB) domain gene family. Analysis of the loss-of-function mutant asl1/lbd36 did not show morphological aberration. Double mutant analysis of asl1/lbd36 together with as2, the ASL1/LBD36 closest homologue, demonstrates that these two members of the AS2/LOB family act partially redundant to control cell fate determination in Arabidopsis petals. Moreover, molecular analysis revealed that overexpression of ASL1/LBD36 leads to repression of the homeobox gene BP, which supports the model that an antagonistic relationship between ASL/LBD and homeobox members is required for the differentiation of lateral organs.     

DNA-binding specificity of the S8 homeodomain.

The murine S8 homeobox gene is expressed in a mesenchyme-specific pattern in embryos, as well as in mesodermal cell lines. The S8 homeodomain is overall similar to paired type homeodomains, but at position 50, which is crucial for specific DNA recognition, it contains a Gln, as is found in Antennapedia (Antp)-type homeodomains. We determined the DNA-binding specificity of the purified S8 homeodomain by in vitro selection of random oligonucleotides. The resulting 11-bp consensus binding site, ANC/TC/TAATTAA/GC resembles, but subtly differs from, the recognition sequences of Antp-type homeodomains. Equilibrium binding constants of down to 6.0 x 10(-10) M were found for binding of the S8 homeodomain to selected oligonucleotides. Using specific antibodies and an oligonucleotide containing an S8-site, we detected by band-shift two abundant DNA binding activities in mesodermal cell lines that correspond to S8 and two more that correspond to its close relative MHox. These S8 protein forms are differentially expressed in retinoic acid-treated P19 EC cells.     

Increase in expression of the homeobox gene, GBX1, in retinol-induced epidermal mucous metaplasia

Using a degenerate RT-PCR-based screening method, we isolated the homeobox gene, Gbx1, from the shank skin of 13-day-old chick embryos. By in situ hybridization analysis we showed that the Gbx1 was expressed in the epidermis of the skin and the mucous epithelium of the intestine, and that among many homeobox genes isolated, expression of the Gbx1 strongly increased in the epidermis when the skin was cultured with 20 microM retinol, which induces epidermal mucous metaplasia. The Gbx1 expression in the epidermis was increased by interaction with the retinol-pretreated dermal fibroblasts, resulting in mucous metaplasia. These results suggest that the Gbx1 regulates the differentiation and transdifferentiation of the epithelium and controls the morphology of the epithelium. We isolated the chick Gbx1 cDNA clones. The amino acid sequences in homeodomain and its downstream encoded by human and chick Gbx1 cDNA were almost the same, but those upstream of the homeodomain were rather different.