Study of novel d1 alleles, defective mutants of the α subunit of heterotrimeric G-protein in rice

It has been shown that the disruption of the alpha-subunit gene of heterotorimeric G-proteins (G alpha) results in dwarf traits, the erection of leaves and the setting of small seeds in rice. These mutants are called d1. We have studied the expression profiles of the transcripts and translation products of rice Ga in ten alleles of d1 including five additional alleles newly identified. By RT-PCR, the transcripts of the G alpha gene were detected in the all d1 alleles. By western blot, the Ga proteins were not detected in the plasma membrane fractions of the d1 alleles with the exception of d1-4. In d1-4, one amino acid change in the GTP-binding box A of the G alpha protein was occurred and even in this case the G alpha protein was only just detectable in the plasma membrane fraction. Given that the G alpha protein did not accumulate in the plasma membrane fraction in d1-8 which has a deletion of just a single amino acid in the G alpha protein, it is likely that a proper conformation of the G alpha is necessary for accumulation of G alpha protein in the plasma membrane. Nine alleles of d1 showed a severer phenotype whilst d1-4 exhibited a mild phenotype with respect to seed size and elongation pattern of internodes. As brassinosteroid signaling was known to be partially impaired in d1s, the sensitivity to 24-epibrassinolide (24-epiBL) was compared among d1 alleles in a T65 genetic background. Only d1-4 showed responses similar to wild type rice. The results show that the d1-4 mutant is a mild allele in terms of the phenotype and mild hyposensitivity to the exogenously applied 24-epiBL.