The sensitivity of Arabidopsis thaliana auxin amidohydrolase mutants and overexpressors on salt, osmotic and temperature stress

Za optimalni učinak, koncentracija auksina u stanicama je strogo regulirana transportom i metabolizmom auksina. Konjugati auksina predstavljaju skladišnu ili razgradnu formu auksina. Auksinske amidohidolaze su enzimi koji hidroliziraju konjugate te oslobađaju aktivni oblik auksina. U ovom istraživanju, odredio sam relativnu ekspresiju gena za auksinske amidohidrolaze ILR1, ILL2 i IAR3 u nadekspresorskim linijama ILR1 13.3, ILR1 3.1, ILL2 25.6, ILL2 27.2, ILL2 8.2, IAR3 11.5 te IAR3 13.4 vrste Arabidopsis thaliana, ekotipa Wassilewskija. Istražio sam utjecaj solnog (150 mM NaCl), osmotskog (300 mM manitola) i temperaturnog stresa (38 °C) na produžni rast i razvoj korijena kod nadekspresora i mutanata (ilr1-2, ill2-1, iar3-2, ill2-1iar3-2 i ilr1-2ill2-1iar3-2) auksinskih amidohidrolaza. Uz to, analizirao sam lokalizaciju rekombinantnog proteina ILL2-GFP u korijenu biljke nadekspresora ILL2 8.2 uzgajanih u uvjetima solnog, osmotskog i temperaturnog stresa. Nadekspresorske linije ILR1 3.1, ILR1 13.3 i ILL2 27.2 imale su 731.17, 182.5 odnosno 350.93 puta veću ekspresiju odgovarajućeg gena auksinske amidohidrolaze u odnosu na divlji tip. Navedene linije su imale i značajnu promjenu u arhitekturi korijena u uvjetima solnog stresa u odnosu na divlji tip. U uvjetima solnog i osmotskog stresa nakupljanje proteina ILL2 bilo je povećano u zoni sazrijevanja korijena, dok se u vršku korijena nije mijenjalo u odnosu na kontrolne uvjete.Auxins act in concentration dependant manner, that is tightly controled by auxin metabolism as well as its transportation. Auxin conjugates are storage or degradable forms of auxins. Auxin amidohydrolases are enzymes that hydrolyse conjugates and release the active form of auxin. In this research, I measured relative expression of auxin amidohydrolases ILR1, ILL2 i IAR3 in Arabidopsis thaliana, ecotype Wassilewskija, overexpressing lines ILR1 13.3, ILR1 3.1, ILL2 25.6, ILL2 27.2, ILL2 8.2, IAR3 11.5 and IAR3 13.4. To reveal function of auxin amidohydrolases in plant stress response I investigated the influence of salt (150 mM NaCl), osmotic (300 mM mannitol) and temperature stress (38 °C) on growth and development of roots in overexpressing lines and auxin amidohydrolase mutants (ilr1-2, ill2-1, iar3-2, ill2- 1iar3-2 i ilr1-2ill2-1iar3-2). Further, I analysed localisation of recombinant protein ILL2-GFP in the overexpressing line ILL2 8.2, grown in conditions of salt, osmotic and temperature stress. Overexpressing lines ILR1 3.1, ILR1 13.3 and ILL2 27.2 had 731.17, 182.5 and 350.93 times higher expression of corresponding native gene compared to wild type, respectively. The root architecture of those overexpressing lines was significantly changed in salt stress condition, compared to wild type. In salt and osmotic stress, the accumulation of ILL2 was significanly increased in the root maturation zone and did not change in the root tip compared to the control conditions