Analýza interakčních partnerů Lipid Transfer Proteinů (LTPs) v listech tabáku

Bc. Zuzana Durajová

Bachelor's thesis

Analysis of Lipid Transfer Protein (LTPs) interaction partners in tobacco leaves

Abstract:

Non-specific lipid transfer proteins (nsLTPs) are specific for plants in which they are associated with the transfer of lipids in the synthesis of leaf wax layers, cell wall remodelling, plant sexual reproduction, and plant responses to stress. Due to their involvement in signalling pathways involved in plant systemic resistance, they have been classified as pathogenesis-related proteins (PR proteins). Previous in vitro studies have also identified nsLTPs as potential partners of elicitins, protein elicitors secreted by phytopathogenic oomycetes of the genus Phytophthora, inducing a state of systemic resistance in plants. The goal of the presented bachelor thesis deals with the confirmation of the possible interaction of nsLTPs with elicitins in an in vivo model of tobacco leaves. Selected nsLTPs showing structural similarity to elicitins were cloned into the binary expression vector of Agrobacterium tumefaciens containing green fluorescent protein (GFP). After the expression of the given nsLTPs in the leaves of Nicotiana benthamiana transformed by A. tumefaciens, their extracellular localization was verified and subsequently their potential interaction with elicitin β-cryptogein secreted by the oomycete Phytophthora cryptogea and inducing systemic resistance in tobacco plants was monitored by the method of immunoprecipitation. In contrast to the measured interaction of LTP1 with elicitin β-cryptogein under in vitro conditions, here we didn´t observe any interaction of selected LTPs with β-cryptogein.

Abstract:

Nešpecifické lipid transfer proteíny (nsLTPs) sú typické pre rastliny, u ktorých sa spájajú s prenosom lipidov v rámci tvorby voskových vrstiev na povrchu listov, remodeláciou bunkovej steny, sexuálnou reprodukciou rastlín a reakciami rastlín na stres. Na základe ich zapojenia sa do signálnych dráh podieľajúcich sa na systémovej rezistencii rastlín ich dnes radíme medzi s patogenézou súvisiace proteíny (PR proteíny). V rámci predchádzajúcich in vitro štúdií boli nsLTPs označované aj ako potenciálni partneri elicitínov, proteínových elicitorov sekretovaných fytopatogénnymi oomycétami rodu Phytophthora schopnými navodiť u rastlín stav systémovej rezistencie. Daná bakalárska práca sa zaoberá potvrdením možnej interakcie nsLTPs s elicitínmi v in vivo modeli listov tabaku. V rámci praktickej časti boli naklonované vybrané nsLTPs vykazujúce štruktúrnu podobnosť s elicitínmi do expresného vektoru Agrobacterium tumefaciens obsahujúceho zelený fluorescenčný proteín (GFP). Po expresii daných nsLTPs v listoch Nicotiana benthamiana po transformácii A. tumefaciens bola overená ich extracelulárna lokalizácia. Následne pomocou metódy imunoprecipitácie bola sledovaná ich potenciálna interakcia s elicitínom, β-kryptogeínom, sekretovaným oomycétou Phytophthora cryptogea a indukujúcim systémovú rezistenciu u rastlín tabaku. V tomto prípade, na rozdiel od in vitro podmienok, kedy bola pozorovaná interakcia LTP1 s elicitínom β-kryptogeínom, žiadna interakcia zaznamenaná nebola.

Keywords

lipid transfer proteiny, rastlinní imunita, systémová rezistence, elicitiny, Agrobacterium tumefaciens, rastlinná imunita, systémová rezistencia, lipid transfer proteins, plant immunity, systemic resistance, elicitins

Language used: Slovak

Date on which the thesis was submitted / produced: 1. 6. 2021

Thesis defence

• Date of defence: 7. 7. 2021
• Supervisor: doc. Mgr. Jan Lochman, Ph.D.
• Reader: Mgr. Markéta Pernisová, Ph.D.

Masaryk University

Faculty of Science

Bachelor programme / field:
Biochemistry / Biochemistry