Following the infection of the potato, with the potato virus Y (PVY), transcription factor Pti5 translocates into nucleus. Pti5 belongs to ERF familiy (ethylene response factor), which have the main role in activation and silencing of targeted genes important in plant response to stress. In this master thesis, we were studying which mechanism is responsible for translocation of Pti5 into the nucleus. Using transient transformation of Nicotiana benthamiana, with the construct, which has gene Pti5 fused with yellow fluorescent protein (YFP) under constitutive 35S promoter and then treated the plants with kinase inhibitor. We followed the expression of YFP under confocal microscope. Our results show that translocation of Pti5 is not caused by phosphorylation. Additionally, we found, that only one protein P19, viral supressor of RNA silencing from tombusviruses, is enough for translocation of Pti5 to the nucleus.
We knew that Pti5 is in the crossroad between signaling pathways of salycilic acid (SA), ethylen (ET) and jasmonic acid (JA). We were interested in activity of promoter Pti5, in comparison to promoter PR1b, which is marker of SA signaling pathway, following treatment with hormones and their inhibitors mentioned above. Our results show that both, analog of SA INA and precursor of ET ACC, increase the induction of promoter Pti5 and PR1b in N. benthamiana leaves. We also can conclud that PR1b is regulated by SA independantly of ET, while induction of Pti5 by SA is most likely regulated through ET signaling. With Western blot we wanted to found out, if the accumulation of Pti5 is increased in the plants infected with PVY. We proved that only with the results obtained by confocal microscopy.
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