Functional analysis of PAT1

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Phytochromes are red/far-red photoreceptors, which play a key role throughout the life of plants. Phytochrome A (phyA) is the only far-red light receptor and required for germination and de-etiolation in far-red light enriched environments. phyA localises to the cytosol in the dark but translocates into the nucleus after activation by light. We have previously shown that nuclear transport of phyA is essential for far-red light perception and that it depends on the two homologous proteins FHY1 (far-red elongated hypocotyl 1) and FHL (FHY1-like). Both FHY1 and FHL contain an NLS (nuclear localisation sequence) and interact with phyA in a light regulated manner. The phyA-binding site at the C-terminus of FHY1/FHL is highly conserved and indispensable for FHY1/FHL function. Remarkably, in far-red light the pat1-1 mutant exhibits a strong dominant negative phenotype, which is caused by the expression of a C-terminally truncated PAT1. Our data show that this truncated version of PAT1 interacts with the C-terminal phyA-binding site of FHY1/FHL and that it almost completely blocks phyA nuclear transport. Both fusing an NLS directly to phyA and increasing the amount of FHY1 by overexpression rescues the pat1-1 phenotype, which identifies the pat1-1 mutant as a phyA nuclear transport mutant. According to our working model PAT1 regulates the amount of FHY1/FHL available for phyA nuclear transport and, as a consequence, sensitivity to far-red light. Thus, any signal regulating PAT1 would modulate far-red light signalling. In the pat1-1 mutant this regulation may be disturbed. Using a combined molecular, cell biological and physiological approach I want to identify factors interacting with the regulatory domain of PAT1 as well as the signal/signalling pathway that employs PAT1 for cross talk with far-red light perception.