Nicotiana tabacum cDNAs encoding α and β subunits of a heterotrimeric GTP-binding protein isolated from hairy root tissues

Abstract

Heterotrimeric GTP-binding proteins (G-proteins) play important roles in signal transduction pathways in eukaryotic cells. Through differential screening of a hairy root cDNA library of tobacco (Nicotiana tabacum L.) against transcripts from non-root tissues of normal cuttings, we obtained a partial cDNA clone that showed abundant expression and high homology to the α subunit gene of plant G-protein. After RACE-PCR, a full-length cDNA clone was obtained, which was 1,677-bp in length and contained an open reading frame encoding a protein of 384 amino acids. A cDNA clone encoding a β subunit of G-protein was also isolated from the same cDNA library based on PCR amplification and library screening. The clone was 1,600-bp in length and contained an open reading frame encoding 377 amino acids. The deduced amino acid sequences of these clones showed high homology (75.5 to 99.8% amino acid identity) with α and β subunits of other plant G-proteins. Genomic Southern blot analysis showed that the amphidiploid tobacco genome possessed two major copies of both α and β subunit genes and some minor homologous copies. Northern blot analysis showed that the transcript of α subunit gene was abundant in the root tissues, particularly in the hairy root tissues. In contrast, the level of expression of the β subunit gene was equivalent in all the tissues studied. Possible function of tobacco G-protein was discussed.