Guannan Liu:
Studying Extrachromosomal Genetic Elements in Sulfolobus

Date: 15-02-2015    Supervisor: Roger A. Garrett

Archaea constitute a separate domain in the universal tree of life. They exhibit exceptional biological properties and provide important insights into the origin of cellular life. Rapid advances in DNA sequencing and bioinformatical methods as well as the development of versatile genetic tools have facilitated the characterization of viruses, plasmids and membrane vesicles. Studying the interactions between Sulfolobus and extrachromosomal genetic elements has provided many new insights into basic molecular processes.

Secreted membrane vesicle seems to be a common characteristic for Sulfolobus. In order to study the biochemical compositions and the genetic functions of these membrane vesicles, production of membrane vesicles in Sulfolobus was optimized, and the membrane vesicles were shown to contain cellular DNA. Furthermore, DNA sequencing revealed that the DNA bound to membrane vesicles consisted of random chromosomal fragments, including IS elements. The results suggest that membrane vesicles could serve as vehicles for the inter-cellular transport of genetic material.

A variant of ATV, ATV2, was isolated that infected a newly isolated Sulfolobus solfataricus P3 strain. Comparative genomics of three closely related viruses (ATV, ATV2, ATVv) revealed a conserved genome organization, but many differences in gene size and content. Comparison of the CRISPR loci in S. solfataricus P3 with those of three published S. solfataricus strains showed many shared spacers, as well as different spacers, especially those adjoining the leader region. Several spacers of the newly isolated S. solfataricus P3 had significant sequence matches to ATV and ATV2 genomes, indicating S. solfataricus P3 has been a host for ATV viruses previously.

Finally, interactions between pKEF9 and Sulfolobus hosts were studied to gain a better understanding of the interactions between conjugative plasmids and hosts. The result also demonstrated why certain archaeal conjugative plasmids are gradually lost during continuous growth. Whereas loss of pKEF9 in S. islandicus was due to interference from the host CRISPR-Cas system, whereas the deactivation of pKEF9 in S. solfataricus was caused by  mobile elements after it had integrated into the host genome.