The cell cycle of the hyperthermophilic archaeal genus Sulfolobus
Sammanfattning: The third domain of life, Archaea is one of the three main evolutionary lineages together with the Bacteria and the Eukarya domains. The archaea are, despite their prokaryotic cell organisation, more closely related to eukaryotes than to bacteria in terms of the informational pathways (DNA replication, transcription and translation). Organisms from the archaeal hyperthermophilic genus Sulfolobus thrives in a hot (80°C), acidic (pH 2-4) and sulphur-rich environment.In my thesis, I have used a variety of different approaches to study the Sulfolobus cell cycle. After dilution of a stationary phase cell culture with fresh medium, synchronous cell cycle progression was obtained. From the synchronised cell culture experiment we could conclude that the major cell cycle events (nucleoid segregation, cell division and chromosome replication) were tightly coupled to each other and to cellular mass increase. Inhibitors of the elongation stage of chromosome replication, and of cell division, as well as drugs arresting the cell cycle in the post-replicative phase, were found in an in vivo screening of a range of antibiotics. The cell cycle was found to be regulated such that the previous cell cycle step had to be successfully accomplished before the next could initiate, except for DNA replication which could occur without an intervening cell division event.The replication pattern of Sulfolobus solfataricus was analysed using a marker frequency assay. From the results, we were able to determine that a single origin is utilized in vivo, that the replication directionality is bidirectional, and also an approximate location of the replication origin within the genome.Intracellular virus production in vivo of SIRV2 (Sulfolobus islandicus rod-shaped virus2) in Sulfolobus islandicus was also analysed. The effects on the host cell were determined, including loss of cell viability, inhibited initiation of replication at virus infection and DNA degradation and loss of cell integrity at the time of virus release. Also, for the first time intracellular virus DNA was visualized with flow cytometry.
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