BIOLOGY & BIOTECHNOLOGY SEMINAR:
"Death & Taxus: A tale of plant cell mortality"
~ Michelle McKee, BBT Graduate Student ~
Advisor: Professor Susan Roberts
Abstract: Production of specialized metabolites in plant cell culture is often enhanced through elicitation in combination with metabolic engineering approaches. These elicitation factors create a fight or flight response in the culture, which can lead to specialized metabolite biosynthesis and/or cell death, depending on the factor and dose. Past studies of viability in relation to paclitaxel production in Taxus cell culture are conflicting as to whether cell death is due to redirection of flux between conserved and specialized metabolism or the cytotoxic compound itself. This work investigates different elicitation methods and culture conditions that have been shown to induce cell death in Taxus cell culture, particularly programmed cell death (PCD). Investigation methods include viability assays, genomic DNA smears, dry weight measurements, and UPLC-quantified metabolite levels. Results indicate that paclitaxel levels increase under methyl jasmonate elicitation and cell viability decreases quickly after exposure to salicylic acid. However, according to Coulter Counter data, these drastically different outcomes result in the same growth curves as the control cultures. Further analyses demonstrate that particle number and size-based dry weight calculations are inaccurate measurements of growth in non-viable cultures. However, data suggest that cell and aggregate volume may be a rapid and straightforward indicator of PCD in Taxus cell culture. Using temperature-treated cell death controls for necrosis and apoptosis, we have confirmed cell volume increase as a consequence of PCD in Taxus. Ongoing work is exploring the underlying mechanisms and the use of different inhibitors of PCD to understand and control for this type of culture death, which is often concurrent with elicitation for paclitaxel production. This research provides a foundation for understanding the identified, but not understood mechanisms of PCD in Taxus and how it relates to paclitaxel production. This work could identify potential molecular targets for engineering Taxus cultures to prevent PCD following certain elicitation techniques. Application of this knowledge could increase the yields of paclitaxel, a valuable pharmaceutical, and other metabolites produced using plant cell culture.
Tuesday, April 2, 2019 @4:15 p.m. - GP 1002