This thesis is focused on the diversity of lichen photobionts and its relation to the mycobionts and environmental factors. The key objective is to map the diversity of lichen photobionts of the genus Stereocaulon, which has been insufficiently examined in comparison with related lichens Cladonia and Lepraria. In total, 92 samples belonging to 12 species of genus Stereocaulon were examined, being found in four continents, at the altitude of 50 to 2 900 metres a. s. l., and variety of natural and artificial substrata. Phylogenetic analyses were inferred on the basis of ITS rDNA, actin type I, 18S rDNA and rbcL sequences. Photobionts belonging to two unrelated genera within Trebouxiophyceae, Asterochloris and Chloroidium, were found in thalli of Stereocaulon lichens. While Asterochloris represents one of the most frequent photobionts of various lichens, Chloroidium has been until now recorded as a lichen photobiont very rarely. The content of heavy metals in substratum can be considered as a determinative factor for its occurrence. A new lineage affiliated to genus Asterochloris was discovered on the island La Palma (the Canary Islands). Interestingly, none of common, widespread Asterochloris lineages has been found in this locality. The lichen photobionts of the genus Stereocaulon were discovered in 12 of...

This diploma thesis has focused on the diversity and biogeography of Asterochloris photobionts. Since no study so far has been published on the biogeography of symbiotic microorganisms, the presented thesis is the first attemp to trace the biogeographic distribution and endemism of symbionts. By gathering 121 Asterochloris sequences obtained from lichen thalli sampled outside Europe and America, the diversity within the genus increased dramatically. The phylogenetic analysis based on the concatenated alignment of ITS rDNA and actin sequences obtained from Cladonia and Stereocaulon photobionts revealed 28 differently supported clades. Of them, eight lineages were newly discovered. Three environmental factors explaining the best the distribution pattern of Asterochloris photobionts were selected according to the statistical tests of the phylogenetic signal: two different types of biogeographical ecoregions and the substrate type. In general, the genus Asterochloris is distributed cosmopolitally, with a very low rate of endemism. Newly obtained data indicate that the restricted distribution of any photobiont clade is not caused by either historic or biological factors, but more likely by specific climatic or habitat preferences.

<p>Lichen-forming fungi employ a successful mode of nutrition as symbiotic partners with green algae and/or cyanobacteria (the photobiont). Nearly one fifth of all known fungi are obligate lichen formers, yet we know little of how they find compatible partners and establish long-lived symbiotic relationships. The combined growth of these symbionts forms a body (thallus) with emergent properties unlike either of the symbionts individually grown. Based on other well-studied eukaryotic systems, the development of a lichen thallus must rely upon the successful identification and collaboration of these two very different organisms. Identifying the molecular basis of microbe recognition and interactions remains one of the greatest challenges in studying symbiotic systems. </p><p>In this thesis, I determine the stage in which to begin looking for lichen symbiosis specific genes, and then examine mycobiont and photobiont genes that, when compared to the aposymbiotic state, are upregulated in the symbiotic state. Using the symbiosis between the mycobiont <italic>Cladonia grayi</italic> and the photobiont <italic>Asterochloris</italic> sp., as well as scanning electron microscopy observations of the earliest stages of contact between <italic>C. grayi</italic> and <italic>Asterochloris</italic> sp., I determined that the mycobiont undergoes a specific change in phenotypic growth in response to <italic>Asterochloris</italic> sp. This change is particular to the lichen symbiosis, and is not observed with algal shaped inanimate objects or algae other than <italic>Asterochloris</italic>. I then used this phenotypically defined stage that is exclusive to lichen symbiosis to begin studying the the genetic and molecular mechanisms underlying the development of a stratified lichen thallus. Using suppression subtractive hybridization to determine differential gene expression, fungal and algal libraries were made of upregulated genes in the first 2 stages of lichen symbiosis. The symbiotic expression levels of select genes were then verified using quantitative PCR. Lastly, a candidate gene for involvement in lichen symbiosis was transformed into <italic>Saccharomyces cerevisiae</italic> to test for protein function.</p><p>Further results of this study show that the fungal protein products of genes upregulated in lichen symbiosis show significant matches to proteins putatively involved in fungal self and non-self recognition, lipid metabolism, negative regulation of glucose repressible genes, an oxidoreductase, a dioxygenase, and a conserved hypothetical protein. Algal genes that are upregulated in lichen symbiosis include a chitinase-like protein, an amino acid metabolism protein, a dynein related protein, and a protein arginine methyltransferase. Furthermore, genes that are expressed in the early stages of lichen symbiosis are common varying metabolic pathways. Furthermore stages 1 and 2 of development are marked not by a drastic change in transcriptional products, but instead by an overall change in genes that are already expressed. Finally, the <italic>Cladonia~grayi Lip3</italic>was cloned in its entirety from genomic DNA and cDNA, was predicted to be secreted using signal peptide prediction software, and shown to be a functioning secreted extracellular lipase in yeast.</p><p>I conclude that many genes are involved in the interactions of symbionts and the development of a stratified lichen thallus, and that many more genes remain to be discovered. Furthermore, the possibility that genes exist in either symbiont that are specific to lichen symbiosis remains, and that their discovery awaits the creation of better genomic tools for \textit{Cladonia~grayi} and <italic>Asterochloris</italic>.</p> / Dissertation

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