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Functional analysis of MAP kinase-dependent elicitor response signaling in tobacco Hall, Hardy Craig


While the MAP kinase, SIPK, is rapidly activated by stress in tobacco (Nicotiana tabacum), its role in mediating stress response outcomes, particularly in its relation to phytohormones and second messengers is unclear. I investigated the tissue and transcriptional responses of tobacco to bacterial and oomycete elicitors, by comparing SIPK-silenced plant, and cell suspension culture lines to WT. SIPK-silencing led to increased sensitivity of tobacco leaf tissue to treatment with bacterial harpin (hrpZ[sub Psph]) (Pseudomonas syringae pv. phaseolicola), indicating that SIPK plays a negative regulatory role in the hrpZ[sub Psph]-induced hypersensitive response. HrpZ[sub Psph]-induced reactive oxygen species (ROS) burst was enhanced in the SIPK-silenced plants compared to the WT plants, indicative of a negative regulatory role for SIPK in harpin-induced ROS accumulation. Strong hrpZ[sub Psph]-induced activation of SIPK (and WIPK) in the NahG line demonstrates that SIPK and WIPK activation are largely SA-independent. SIPK-silenced tobacco plants hyper-accumulated SA following harpin treatment, suggesting that SIPK acts upstream as a negative regulator of harpin-induced SA accumulation. To address the functional roles of SIPK in mediating transcriptional responses induced by hrpZ[sub Psph] and the oomycetes elicitor β-megaspermin (Phytophthora megasperma H20), I monitored the transcriptional responses of WT and SIPK-Ri tobacco cell suspension cultures to each elicitor at 4 and 8-hours post-treatment, using TIGR potato cDNA microarrays (10K element). SIPK-affected transcriptional responses were analyzed in the context of the WT responses to these elicitors in order to identify a set of genes associated with each elicitor that respond in a SIPK-dependent manner. SIPK-silencing was found to have a substantial impact on both hrpZ[sub Psph]- and β-megaspermin-induced transcriptional responses. Hierarchical clustering of genes that responded significantly to either elicitor (but not both) revealed striking functional differences between the two SIPK-dependent responses, suggesting that signal transduction through SIPK plays an important role in conditioning elicitor-specific transcriptional responses.

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