Characterization of CRT1 family members in root-induced basal resistance, SAR, and ISR of Arabidopsis and barley

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Plants are continuously exposed to biotic, as well as abiotic, stress factors, such as pathogenic microbes. Interaction with and recognition of microbes leads to production of signals at the site(s) of contact that are transmitted to the surrounding and distal tissues of the plant. This signal transduction network following non-self-recognition has been extensively studied in the aerial parts of the plant. Despite the fact that the rhizosphere is populated by a disproportionately higher number of microbes, to date relatively little is known about signaling in the roots upon contact with microbial pathogens and mutualists.A genetic screen in Arabidopsis thaliana led to the discovery of CRT1 (Compromised for Recognition of Turnip Crinkle Virus 1). CRT1 belongs to the MORC-type subclade of GHKL ATPases and was shown to play an important role in Arabidopsis resistance to viral, bacterial, and oomycete pathogens, acting at five levels of immunity including effector-triggered immunity (ETI), pattern-triggered immunity (PTI), basal resistance, non-host resistance, and systemic acquired resistance (SAR). Previous collaborative work of the applicant together with the Mercator fellow has shown that a subpopulation of CRT1 i) resides in the nucleus, ii) binds DNA, iii) exhibits endonuclease activity, and iv) affects tolerance to a DNA-damaging agent, arguing that it has (an) important nuclear function(s) during the activation of immune responses.CRT1 family members have been identified in all plant species, including monocots, sequenced to date, suggesting they have an evolutionary conserved function. Five CRT1 family proteins, named HvCRT1, HvCRH1, HvCRH3, HvCRH6, and HvCRH7, have been identified by the applicants within the not yet fully annotated barley (Hordeum vulgare) genome. Altering expression of these genes either by RNAi-mediated gene silencing or overexpression changes the resistance of barley to root rot caused by Fusarium graminearum and leaf powdery mildew caused by Blumeria graminis. Given these results, we strongly suspect that CRT1 and/or its homologs participate in root-microbe interactions in Arabidopsis, barley, and other plant species. Since silencing of CRT1 family members in barley leads to higher resistance against necrotrophic (Fusarium) and biotrophic (Blumeria) pathogens, it is anticipated that these genes may have high potential for agronomical applications.To elucidate the mode of action for CRT1 family members and to reveal the underlying mechanisms for the apparent species-specific differences between CRT1 function in barley and Arabidopsis, recombinant AtCRT1 and HvCRT1 will be characterized using genetic, molecular, biochemical, biophysical, and structural methods. Furthermore, stable transgenic barley lines with altered expression of CRT1 family genes will be generated and characterized with respect to their response to root-initiated pathogen infection. The long-term goal of these studies is crop plant improvement.