Cytokinin levels and signaling respond to wounding and the perception of herbivore elicitors in Nicotiana attenuata

Nearly half a century ago insect herbivores were found to induce the formation of green islands by manipulating cytokinin （CK） levels. However, the response of the CK pathway to attack by chewing insect herbivores remains unclear. Here, we characterize the CK pathway of Nicotiana attenuata （Torr. ex S. Wats.） and its response to wounding and perception of herbivoreassociated molecular patterns （HAMPs）. We identified 44 genes involved in CK biosynthesis, inactivation, degradation, and signaling. Leaf wounding rapidly induced transcriptional changes in multiple genes throughout the pathway, as well as in the levels of CKs, including isopentenyladenosine and cis-zeatin riboside; perception of HAMPs present in the oral secretions （OS） of the specialist herbivore Manduca sexta amplified these responses. The jasmonate pathway, which triggers many herbivore-induced processes, was not required for these HAMP-triggered changes, but rather suppressed the CK responses. Interestingly CK pathway changes were observed also in systemic leaves in response to wounding and OS application indicating a role of CKs in mediating long distance systemic processes in response to herbivory. Since wounding and grasshopper OS elicited similar accumulations of CKs in Arabidopsis thaliana L., we propose that CKs are integral components of wounding and HAMP-triggered responses in many plant species.

Shifting Nicotiana attenuata's diurnal rhythm does not alter its resistance to the specialist herbivore Manduca sexta

Arabidopsis thaliana plants are less resistant to attack by the generalist lepidopteran herbivore Trichoplusia ni when plants and herbivores are entrained to opposite, versus identical diurnal cycles and tested under constant conditions. This effect is associated with circadian fluctuations in levels of jasmonic acid, the transcription factor MYC2, and glucosinolate contents in leaves. We tested whether a similar effect could be observed in a different plant-herbivore system： the wild tobacco Nicotiana attenuata and its co-evolved specialist herbivore, Manduca sexta. We measured larval growth on plants under both constant and diurnal conditions following identical or opposite entrainment, profiled the metabolome of attacked leaf tissue, quantified specific metabolites known to reduce M. sexta growth, and monitored A/I. sexta feeding activity under all experimental conditions. Entrainment did not consistently affect NI. sexta growth or plant defense induction. However, both were reduced under constant dark conditions,as was M. sexta feeding activity. Our data indicate that the response induced by NI. sexta in N. attenuata is robust to diurnal cues and independent of plant or herbivore entrain- ment. We propose that while the patterns of constitutive or general damage-induced defense may undergo circadian fluctuation, the orchestration of specific induced responses is more complex.