This study is focused on changes in protein expression in a glioblastoma cell line during infection with tick-borne encephalitis virus. Newly synthesized proteins were distinguished from previously synthesized proteins using bioorthogonal chemistry (BONCAT method) to observe changes in protein synthesis. Labelled proteins were visualized using two-dimensional PAGE and western blotting followed by Click reaction on membrane. Differences in protein pattern between control and infected cells were observed.

京都大学 / 新制・課程博士 / 博士(農学) / 甲第24669号 / 農博第2552号 / 新制||農||1099(附属図書館) / 学位論文||R5||N5450(農学部図書室) / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 菅瀬 謙治, 教授 小川 順, 教授 森 直樹 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

In the field of marine microbial ecology, many questions remain unanswered with regards to the physiological trade-offs made by phytoplankton to maximize growth (e.g., nutrient acquisition) and minimize loss (e.g., predation defenses). These tradeoffs, which occur at the cellular level, have wide reaching impacts on food web dynamics and global biogeochemical cycles. In the first chapter, we explored the use of a non-canonical amino acid (NCAA) technique, bioorthogonal non-canonical amino-acid tagging (BONCAT), in phytoplankton model systems. This technique has potential to work well in natural systems by enabling isolation of only newly synthesized proteins during an incubation period with the NCAA, reducing the complexity of natural proteomics and easing the elucidation of patterns. However, in testing BONCAT across several groups of cultured phytoplankton, we discovered that the NCAA molecule induced a stress response in the globally ubiquitous marine picocyanobacteria, Synechococcus sp. Therefore, in addition to confirming the uptake of modified amino acids by phytoplankton, chapter one investigated the implications of this stress response and limitations when using this technique to study marine microbial communities. In chapter two, we addressed our initial question by exploring tradeoffs at the protein level in a simplified culture system. This approach revealed insights into metabolic tradeoffs in response to predation pressure and nutrient stress. These insights into how phytoplankton negotiate these physiological tradeoffs at the protein level could ultimately allow for targeted proteomic studies in natural systems.