THE INDUCTION OF SEED-GERMINATION IN ARABIDOPSIS-THALIANA IS REGULATED PRINCIPALLY BY PHYTOCHROME B AND SECONDARILY BY PHYTOCHROME-A

We examined whether spectrally active phytochrome A (PhyA) and phytoch rome B (PhyB) play specific roles in the induction of seed germination in Arabidopsis thaliana (L.) Heynh., using PhyA- and PhyB-null mutant s, fre1-1 (A. Nagatani, J.W. Reed, J. Chory [1993] Plant Physiol 102: 269-277) and hy3-Bo64 (J. Reed, P. Nagpal, D.S. Poole, M. Furuya, I. C hory [1993] Plant Cell 5: 147-157). When dormant seeds of each genotyp e imbibed in the dark on aqueous agar plates, the hy3 (phyB) mutant di d not germinate, whereas the fre1 (phyA) mutant germinated at a rate o f 50 to 60%, and the wild type (WT) germinated at a rate of 60 to 70%. By contrast, seeds of all genotypes germinated to nearly 100% when pl ated in continuous irradiation with white or red light. When plated in continuous far-red tight, however, frequencies of seed germination of the WT and the fre1 and hy3 mutants averaged 14, nearly 0, and 47%, r espectively, suggesting that PhyB in the red-absorbing form prevents P hyA-dependent germination under continuous far-red light. When irradia ted briefly with red or far-red light after imbibition for 1 h, a typi cal photoreversible effect on seed germination was observed in the fre 1 mutant and the WT but not in the hy3 mutant. In contrast, when allow ed to imbibe in the dark for 24 to 48 h and exposed to red light, the seed germination frequencies of the hy3 mutant were more than 40%. Imm unoblot analyses of the mutant seeds showed that PhyB apoprotein accum ulated in dormant seeds of the WT and the fre1 mutant as much as in th e seeds that had imbibed. In contrast, PhyA apoprotein, although detec ted in etiolated seedlings grown in the dark for 5 d, was not detectab le in the dormant seeds of the WT and the hy3 mutant. The above physio logical and immunochemical evidence indicates that PhyB in the far-red -absorbing form was stored in the Arabidopsis seeds and resulted in ge rmination in the dark. Hence, PhyA does not play any role in dark germ ination but induces germination under continuous irradiation with far- red light. Finally, we examined seeds from a signal transduction mutan t, det1, and a det1/hy3 double mutant. The det1 seeds exhibited photor eversible responses of germination on aqueous agar plates, and the det 1/ hy3 double mutant seeds did not. Hence, DET1 is likely to ad in a d istinct pathway from PhyB in the photoregulation of seed germination.