Date of Graduation

5-2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Entomology (PhD)

Degree Level

Graduate

Department

Entomology and Plant Pathology

Advisor/Mentor

Goggin, Fiona L.

Committee Member

Wiedenmann, Robert N.

Second Committee Member

Rojas, Clemencia M.

Third Committee Member

Szalanski, Allen L.

Fourth Committee Member

Korth, Kenneth L.

Keywords

Aphid resistance; Arabidopsis thaliana; Chloroplast; Fatty Acid Desaturases; Myzus persicae; Plant defense; Singlet oxygen

Abstract

Fatty Acid Desaturase 7 (FAD7) is a chloroplast-localized enzyme that alters the fatty acid content of photosynthetic membranes, and that negatively regulates plant defenses against aphids. Previous studies in the model organism Arabidopsis (Arabidopsis thaliana) have shown that loss-of-function mutations in FAD7 decrease population growth of the green peach aphid (GPA; Myzus persicae Sulzer). This study further characterized the effects of a fad7 null mutant on aphids, and investigated the role of reactive oxygen species (ROS), including singlet oxygen (1O2), in plant responses to aphid resistance in fad7 as well as in wild type plants and a mutant with heightened 1O2 accumulation (flu). Bioassays indicated that aphid resistance in fad7 is species-specific and impacts the generalist herbivore GPA but not a specialist herbivore, the cabbage aphid (Brevicoryne brassicae). The effects of fad7 on GPA appeared to be antibiotic but not antixenotic, and reduced population growth by decreasing adult fecundity and increasing juvenile development time. Compared to wild type controls, fad7 also has increased expression of an 1O2-responsive reporter gene (AAA-ATPase:Luc) both constitutively and in response to aphids, and aphid challenge upregulated AAA-ATPase:Luc in both fad7 and in wild type plants. These results suggest that 1O2 may contribute to plant defenses against aphids. A redox-sensitive green fluorescent protein (roGFP2) tagged to the chloroplast showed that GPA induce a rapid and sustained oxidative response in the chloroplast; furthermore, ROS accumulation in response to aphid feeding, as measured with luminol, was attenuated in transgenic plants with enhanced expression of an 1O2 scavenger in the chloroplast (SPS1oex). Together, these results suggest that ROS, including 1O2, are accumulating in the chloroplast in response to aphid challenge. To explore whether 1O2 impacts host suitability for aphids, aphid performance was also measured on the conditional fluorescence (flu) mutant, which has normal ROS levels when grown in continuous light but accumulates high 1O2 in the chloroplast when transferred from light to dark and back to light again (ie. a light/dark/light shift). The population growth of GPA was significantly lower on flu exposed to a light/dark/light shift than on flu grown in continuous light or on wild type plants in either light treatment; thus, high 1O2 conferred aphid resistance. Because 1O2 generated in the chloroplast is known to impact nuclear gene expression via a retrograde signaling pathway that requires the chloroplast-localized EXECUTER1/EXECUTER2 (EX1/EX2) proteins, additional experiments were conducted to determine if EX1/EX2 were required for aphid resistance in fad7 or flu. Null mutations in EX1 and EX2 partially reduced aphid resistance in flu, and eliminated resistance in the fad7. This indicates that 1O2 accumulation and EX1/EX2 signaling contribute to aphid resistance in both genotypes. Compared to wild type plants, fad7 had comparable chlorophyll levels and higher maximum potential quantum efficiency (Fv/Fm) of Photosystem II, which is typically the primary source of 1O2 in the chloroplast. Thus, enhanced aphid resistance associated with increased 1O2 accumulation did not compromise photosynthesis in fad7. This work demonstrates, for the first time, a role for 1O2-mediated chloroplast signaling in plant defense against herbivores.

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