The Nostoc Symbiont of Lichens : Diversity, Specificity and Cellular Modifications

Cyanobacteria belonging to the genus Nostoc have the capacity to form symbiotic associations with a wide range of organisms. Diversity, specificity and cellular modifications of the symbiosis between Nostoc and fungi in the formation of lichens were investigated in this thesis. The use of the tRNALeuUAA intron as a genetic marker for the subgeneric identification of Nostoc in complex field material was developed. Lichens belonging to the genera Peltigera and Nephroma show limited variability in their Nostoc symbionts. The in situ symbiont consists of a single strainn rather than a community of different Nostocs, and single thalli consistently contained the same symbiont. Patterns in symbiont identity were found in geographically remote populations and the lichen species, rather than growth locality, was shown to be important for the identity of the Nostoc symbiont. Examination of a P. aphthosa photosymbiodeme revealed that one Nostoc has the capacity to perform the physiological roles found in both bipartite and tripartite lichens. The symbiotic association between bryophytes and Nostoc on the other hand exhibited a much greater variation of Nostoc symbionts. Evolutionary patterns in the tRNALeuUAA intron were analyzed and it was shown that sequence variation was caused by several processes other than random mutations. Such evolutionary processes in genetic markers are crucial to consider, especially if phylogenetic reconstructions are attempted. Protein profiles of symbiotic and free living Nostoc were analyzed using 2-dimensional gel electrophoresis. One of the major proteins in the extracts from freshly isolated symbionts was partially sequenced and shown to contain a fasciclin domain. The corresponding ORF in N. punctiforme was homologous to symbiotically induced genes found in different symbiotic systems. This thesis gives new perspectives on lichens and pr for further exaovides a platform for further examiniations using tools provided by modern biology

Fasciclin Domain Proteins Are Present in Nostoc Symbionts of Lichens

Differences in the soluble protein fraction between the freshly isolated cyanobiont of lichen Peltigera membranacea, the corresponding free-living strain, and Nostoc punctiforme were analyzed. One protein, which was among the most prominent proteins of the freshly isolated cyanobiont, was expressed at a lower level in the corresponding free-living strain and was not detected at all on the two-dimensional gels of N. punctiforme. This protein was partially sequenced, and the corresponding open reading frame (ORF) in the N. punctiforme genome was identified. This ORF contains a fasciclin domain typical of a class of surface-associated proteins involved in cell adhesion. Similar fasciclin motif-containing genes have previously been shown to be symbiotically induced in other symbiotic systems

Sequence Variation of the tRNA ^Leu Intron as a Marker for Genetic Diversity and Specificity of Symbiotic Cyanobacteria in Some Lichens

ABSTRACT We examined the genetic diversity of Nostoc symbionts in some lichens by using the tRNA Leu (UAA) intron as a genetic marker. The nucleotide sequence was analyzed in the context of the secondary structure of the transcribed intron. Cyanobacterial tRNA Leu (UAA) introns were specifically amplified from freshly collected lichen samples without previous DNA extraction. The lichen species used in the present study were Nephroma arcticum , Peltigera aphthosa , P. membranacea , and P. canina . Introns with different sizes around 300 bp were consistently obtained. Multiple clones from single PCRs were screened by using their single-stranded conformational polymorphism pattern, and the nucleotide sequence was determined. No evidence for sample heterogenity was found. This implies that the symbiont in situ is not a diverse community of cyanobionts but, rather, one Nostoc strain. Furthermore, each lichen thallus contained only one intron type, indicating that each thallus is colonized only once or that there is a high degree of specificity. The same cyanobacterial intron sequence was also found in samples of one lichen species from different localities. In a phylogenetic analysis, the cyanobacterial lichen sequences grouped together with the sequences from two free-living Nostoc strains. The size differences in the intron were due to insertions and deletions in highly variable regions. The sequence data were used in discussions concerning specificity and biology of the lichen symbiosis. It is concluded that the tRNA Leu (UAA) intron can be of great value when examining cyanobacterial diversity. </jats:p