NPK15, a tobacco protein-serine/threonine kinase with a single hydrophobic region near the amino-terminus.
A cDNA clone (cNPK15) was isolated from tobacco cells in suspension culture, which encodes a predicted protein kinase of 422 amino acids. The predicted NPK15 protein consists of a hydrophobic region near the amino-terminus, a linker domain and the catalytic domain of a protein-serine/threonine kinase in the carboxyl-half. NPK15 was not found to be closely related to any reported protein, but its putative catalytic domain shares some structural similarity with those of receptor-like protein kinases of plants, such as ZmPK1 from Zea mays and TMK1 from Arabidopsis, even though no receptor-like domain is found in NPK15. Recombinant NPK15 expressed in Escherichia coli as a fusion protein was found capable of autophosphorylation and of phosphorylation of the histone H1 protein on both serine and threonine residues. Upon overexpression of cNPK15 under control of the promoter of cauliflower mosaic virus 35S RNA in tobacco cells, into which it had been introduced by Agrobacterium-mediated transformation, the NPK15 gene acted as a "suicide" gene and blocked proliferation of the host cells. By contrast, such a suicide effect was not observed with the gene for a kinase-negative mutant protein in which the nucleotide sequence for the ATP-binding site had been mutated or with a mutant derivative encoding a protein in which the hydrophobic region had been deleted. Thus, the protein kinase activity of NPK15 and the hydrophobic region of the protein are responsible for the suicide effect. The NPK15 protein kinase seems to be associated with specific cellular functions. Southern blot analysis with cNPK15 as the probe detected several fragments in restriction digests of genomic DNAs from both tobacco and other members of the Solanaceae. This results suggests that NPK15-related genes constitute a small gene family in the genomes of Solanaceae.
Department of Agronomy, Faculty of Agriculture, Tohoku University, Sendai, Japan.
Y
Y
Ito
H
H
Banno
T
T
Moribe
K
K
Hinata
Y
Y
Machida
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D018076:DNA, Complementary; D018744:DNA, Plant; D017343:Genes, Plant; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D010947:Plants, Toxic; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics
D000596:Amino Acids; D018076:DNA, Complementary; D018744:DNA, Plant; D010940:Plant Proteins; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases