The slow road to the eukaryotic genome

The eukaryotic genome is a mosaic of eubacterial and archaeal genes in addition to those unique to itself. The mosaic may have arisen as the result of two prokaryotes merging their genomes, or from genes acquired from an endosymbiont of eubacterial origin. A third possibility is that the eukaryotic genome arose from successive events of lateral gene transfer over long periods of time. This theory does not exclude the endosymbiont, but questions whether it is necessary to explain the peculiar set of eukaryotic genes. We use phylogenetic studies and reconstructions of ancestral first appearances of genes on the prokaryotic phylogeny to assess evidence for the lateral gene transfer scenario. We find that phylogenies advanced to support fusion can also arise from a succession of lateral gene transfer events. Our reconstructions of ancestral first appearances of genes reveal that the various genes that make up the eukaryotic mosaic arose at different times and in diverse lineages on the prokaryotic tree, and were not available in a single lineage. Successive events of lateral gene transfer can explain the unusual mosaic structure of the eukaryotic genome, with its content linked to the immediate adaptive value of the genes its acquired. Progress in understanding eukaryotes may come from identifying ancestral features such as the eukaryotic splicesome that could explain why this lineage invaded, or created, the eukaryoticniche.

The Role of Splicing Factors and Small Nuclear RNAS in Spliceosomal Formation

Protein coding genes are comprised of protein-coding exons and non-protein-coding introns. The process of splicing involves removal of the introns and joining of the exons to form a mature messenger RNA, which subsequently undergoes translation into polypeptide. The spliceosome is a large, RNA/protein assembly of five small nuclear RNAs as well as over 300 proteins, which catalyzes intron removal and exon ligation. The selection of specific exons for inclusion in the mature messenger RNA is spatio-temporally regulated and results in production of an enormous diversity of polypeptides from a single gene locus. This phenomenon, known as alternative splicing, is regulated, in part, by protein splicing factors, which target the spliceosome to exon/intron boundaries. The first part of my dissertation (Chapters II and III) focuses on the discovery and characterization of the 45 kilodalton FK506 binding protein (FKBP45), which I discovered in the silk moth, Bombyx mori, as a U1 small nuclear RNA binding protein. This protein family binds the immunosuppressants FK506 and rapamycin and contains peptidyl-prolyl cis-trans isomerase activity, which converts polypeptides from cis to trans about a proline residue. This is the first time that an FKBP has been identified in the spliceosome. The second section of my dissertation (Chapters IV, V, VI and VII) is an investigation of the potential role of small nuclear RNA sequence variants in the control of splicing. I identified 46 copies of small nuclear RNAs in the 6X whole genome shotgun of the Bombyx mori p50T strain. These variants may play a role in differential binding of specific proteins that mediate alternative splicing. Along these lines, further investigation of U2 snRNA sequence variants in Bombyx mori demonstrated that some U2 snRNAs preferentially assemble into high molecular weight spliceosomal complexes over others. Expression of snRNA variants may represent another mechanism by which the cell is able to fine tune the splicing process.