In this work, we have performed comparative evolutionary analysis, functional genomics analysis, and machine learning analysis to identify the molecular factors that discriminate between multipartite and unipartite bacteria, with the goal to decipher taxon-specific factors and those that are prevalent across the taxa underlying the these traits. We assessed the roles of evolutionary mechanisms, namely, horizontal gene transfer and gene gain, in driving the divergence of bacteria with single and multiple chromosomes. In addition, we performed functional genomic analysis to garner support for our findings from comparative evolutionary analysis. We found genes such as those encoding conserved hypothetical protein DR_A0179 and hypothetical protein DR_A0109 in Deinococcus radiodurans R1, and Putative phage phi-C31 gp36 major capsid-like protein and hypothetical protein RSP_3729 in Rhodobacter sphaeroides 2.4.1, which are located on accessory chromosomes in both bacteria and were not found in the inferred ancestral sequences, and on the primary chromosomes, as well as were not found in their closest relatives with single chromosome within the same clade. These genes emphasize the important potential roles of the secondary chromosomes in helping multipartite bacteria to adapt to specialized environments or conditions. In addition, we applied machine learning algorithms to predict multipartite genomes based on gene content of multipartite genomes and their unipartite relatives, and leveraged this to identify genes that are deemed important by machine learning in discriminating between multipartite and unipartite genomes. This approach led to the identification of marker genes that could be used in discriminating between bacteria with multipartite genomes and. bacteria with single chromosome genomes Furthermore, we examined modules in gene co-expression networks of multipartite Rhodobacter sphaeroides 2.4.1 and its close unipartite relative Rhodobacter capsulatus SB 1003 that were enriched in genes differentially expressing under stressful conditions representing different experiments. This led to the identification of 6 modules in the Rhodobacter sphaeroides 2.4.1 network and 3 modules in the Rhodobacter capsulatus SB 1003 network, which were significantly enriched (2-fold or more) in differentially expressing genes, revealing the vital roles of these gene modules representing different pathways or networks of pathways (known or unknown) in enabling the bacteria to adapt to stressful conditions. Overall, our study highlights genetic factors that may be driving the evolution of multipartite bacterial genomes; future studies may focus on unraveling the specific roles of these genes in the adaptation and maintenance of multipartite genomes.

Please read the abstract in the section 00front of this document / Dissertation (MSc (Botany))--University of Pretoria, 2005. / Plant Science / unrestricted

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Thesis (Ph. D.)--University of Hong Kong, 2007. / Title proper from title frame. Also available in printed format.

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Integrases (Int) are enzymes that mediate the integration and excision of viral DNA into or out of their hosts‟ chromosomes and can therefore be exploited to integrate or delete genes in a precise way. In order to establish lysogeny, integrase mediates recombination between the bacterial and phage attachment sites, attB and attP respectively to generate an integrated prophage flanked by attL and attR. This reaction occurs in vitro without any need for accessory proteins prompting the question, how does the prophage excise? Phages use accessory proteins, Recombination Directionality Factors, RDFs to control the directionality of integrase. For the serine integrase family, RDFs have been identified for three phages, TP901-1, φRv1 and Bxb1, and there is no detectable sequence conservation between them. This work has identified the φC31 early protein gp3 as the RDF. Gp3 acts stoichiometrically to activate excision and binds to Int in solution and in complex with DNA. Insight into the mechanism of gp3 action has revealed that it is at the synapse level that gp3 switches the directionality of Int. The properties of the gp3+Int driven reaction was found to be similar to that mediated by a previously characterised mutated Int, IntE449K that triggers gp3 independent excision (Rowley et al., 2008). Despite φC31 and φBT1 Ints being only 21% identical in sequence, the gp3 homologues from these phages cross-react. Both the gp3s bind to the last 200 amino acids of C-terminal domain of φC31 Int to activate excision and inhibit integration. Evidence is presented that gp3, on binding to Int, overcomes an innate mechanism that normally prevents synapsis of the excision substrates. These observations could lead to further exploitation of φC31 system as a tool for genome engineering.

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