Plant-symbiotic fungi as chemical engineers: multi-genome analysis of the Clavicipitaceae reveals dynamics of alkaloid Loci

The fungal family Clavicipitaceae includes plant symbionts and parasites that produce several psychoactive and bioprotective alkaloids. The family includes grass symbionts in the epichloae clade (Epichloë and Neotyphodium species), which are extraordinarily diverse both in their host interactions and in their alkaloid profiles. Epichloae produce alkaloids of four distinct classes, all of which deter insects, and some—including the infamous ergot alkaloids—have potent effects on mammals. The exceptional chemotypic diversity of the epichloae may relate to their broad range of host interactions, whereby some are pathogenic and contagious, others are mutualistic and vertically transmitted (seed-borne), and still others vary in pathogenic or mutualistic behavior. We profiled the alkaloids and sequenced the genomes of 10 epichloae, three ergot fungi (Claviceps species), a morning-glory symbiont (Periglandula ipomoeae), and a bamboo pathogen (Aciculosporium take), and compared the gene clusters for four classes of alkaloids. Results indicated a strong tendency for alkaloid loci to have conserved cores that specify the skeleton structures and peripheral genes that determine chemical variations that are known to affect their pharmacological specificities. Generally, gene locations in cluster peripheries positioned them near to transposon-derived, AT-rich repeat blocks, which were probably involved in gene losses, duplications, and neofunctionalizations. The alkaloid loci in the epichloae had unusual structures riddled with large, complex, and dynamic repeat blocks. This feature was not reflective of overall differences in repeat contents in the genomes, nor was it characteristic of most other specialized metabolism loci. The organization and dynamics of alkaloid loci and abundant repeat blocks in the epichloae suggested that these fungi are under selection for alkaloid diversification. We suggest that such selection is related to the variable life histories of the epichloae, their protective roles as symbionts, and their associations with the highly speciose and ecologically diverse cool-season grasses

Cross-species transferability and mapping of genomic and cDNA SSRs in pines

Two unigene datasets of Pinus taeda and Pinus pinaster were screened to detect di-, tri- and tetranucleotide repeated motifs using the SSRIT script. A total of 419 simple sequence repeats (SSRs) were identified, from which only 12.8% overlapped between the two sets. The position of the SSRs within their coding sequences were predicted using FrameD. Trinucleotides appeared to be the most abundant repeated motif (63 and 51% in P. taeda and P. pinaster, respectively) and tended to be found within translated regions (76% in both species), whereas dinucleotide repeats were preferentially found within the 5'- and 3'-untranslated regions (75 and 65%, respectively). Fifty-three primer pairs amplifying a single PCR fragment in the source species (mainly P. taeda), were tested for amplification in six other pine species. The amplification rate with other pine species was high and corresponded with the phylogenetic distance between species, varying from 64.6% in P. canariensis to 94.2% in P. radiata. Genomic SSRs were found to be less transferable; 58 of the 107 primer pairs (i.e., 54%) derived from P. radiata amplified a single fragment in P. pinaster. Nine cDNA-SSRs were located to their chromosomes in two P. pinaster linkage maps. The level of polymorphism of these cDNA-SSRs was compared to that of previously and newly developed genomic-SSRs. Overall, genomic SSRs tend to perform better in terms of heterozygosity and number of alleles. This study suggests that useful SSR markers can be developed from pine EST