Takefumi Kawata

Nucleotide sequences of the 5.8S rRNA gene and internal transcribed spacer regions in carrot and broad bean ribosomal DNA.

SummaryNucleotide sequences of the first and second internal transcribed spacers (ITS1 and ITS2, respectively) of ribosomal DNA (rDNA) from two dicot plants, carrot and broad bean, were determined. These sequences were compared with those of rice, a monocot plant, and other eukaryotic organisms. Both types of ITS region in some species of Angiospermae were the shortest among all eukaryotes so far examined and showed a wide range of variation in their G+C content, in contrast to a general trend toward very high G+C content in animals. Phylogenetic relationships of plants with animals and lower eukaryotes were considered using the nucleotide sequences of carrot and broad bean 5.8S rDNA that were determined in the present study, together with that of wheat 5.8S rRNA, which has been reported previously.

Nuclear protein(s) binding to the conserved DNA hexameric sequence postulated to regulate transcription of wheat histone genes

Nuclear protein(s) that specifically bind(s) to the upstream hexamer motif, ACGTCA, of wheat histone H3 and H4 genes has (have) been identified. Sequences homologous to this hexamer are found to be conserved in the upstream region of not only wheat histone genes but also other plant and animal histone genes. This suggests a possible role(s) for the hexamer and the nuclear protein(s) in the transcriptional regulation of the wheat histone genes. This hexamer is homologous to the upstream core sequence, TGACGTCA, which is highly conserved in some animal genes whose expression is regulated by cAMP.

DNA-binding protein(s) interacts with a conserved nonameric sequence in the upstream regions of wheat histone genes

A nuclear protein(s), HBP‐2, that binds to the upstream region of the wheat histone H4 gene was identified from a fractionated nuclear extract of wheat germ by DNase I footprinting. The DNase I‐protected region contained the conserved nonameric motif, CATCCAACG. Cross‐competition experiments that used the mobility shift assay showed that this nuclear protein(s) binds specifically to the upstream sequence that has been postulated to be a cis element of the wheat H3 gene. Our findings suggest that this DNA‐binding protein(s) may be a trans‐acting factor in the regulation of the transcription of wheat histone genes.

Isolation and characterization of a cDNA clone encoding the TATA box-binding protein (TFIID) from wheat

We isolated a complementary DNA (cDNA) encoding the TATA-binding factor ‘TFIID’ from a wheat seedling cDNA library. The wheat TFIID transcript of 1.2 kb poly(A)+ RNA was expressed at a low level early in germination, but gradually increased as the seedlings developed. In vitro binding experiments showed that the bacterially expressed wheat TFIID protein could specifically bind to the TATA boxes of the cauliflower mosaic virus (CaMV) 35S, wheat histone H3 and adenovirus major late genes with different affinity. A comparison with Arabidopsis TFIID showed the presence of a plant-specific region consisting of 13 amino acids at the divergent amino terminus and a conserved region (182 amino acids) at the carboxy terminus longer than that observed in yeasts (180 amino acids) and animals (181 amino acids).

Function of the hexameric sequence in the cauliflower mosaic virus 35S RNA promoter region

The hexameric sequence ACGTCA functions in transcriptional regulation of wheat histone genes. The cauliflower mosaic virus (CaMV) 35S RNA promoter has the same hexameric sequence, and mutation analyses confirmed that the hexamer contributed greatly to transcription from the 35S promoter when a test gene with this promoter was introduced into sunflower cells. Electrophoretic mobility shift assays revealed the existence of a nuclear protein(s) in sunflower cells which is homologous to the HBP-1b that has been identified as binding to the 35S promoter in wheat. These results provide evidence of the involvement of the hexameric sequence and the HBP-1b-like DNA binding protein(s) in transcription from the 35S promoter.