The goal of the fungal mitochondrial genome project (FMGP) is to sequence complete mitochondrial genomes for a representative sample of the major fungal lineages; to analyze the genome structure, gene content, and conserved sequence elements of these sequences; and to study the evolution of gene expression in fungal mitochondria. By using our new sequence data for evolutionary studies, we were able to construct phylogenetic trees that provide further solid evidence that animals and fungi share a common ancestor to the exclusion of chlorophytes and protists. With a database comprising multiple mitochondrial gene sequences, the level of support for our mitochondrial phylogenies is unprecedented, in comparison to trees inferred with nuclear ribosomal RNA sequences. We also found several new molecular features in the mitochondrial genomes of lower fungi, including: (1) tRNA editing, which is the same type as that found in the mitochondria of the amoeboid protozoan Acanthamoeba castellanii; (2) two novel types of putative mobile DNA elements, one encoding a site-specific endonuclease that confers mobility on the element, and the other constituting a class of highly compact, structured elements; and (3) a large number of introns, which provide insights into intron origins and evolution. Here, we present an overview of these results, and discuss examples of the diversity of structures found in the fungal mitochondrial genome.

Detailed knowledge of gene maps or even complete nucleotide sequences for small genomes leads to the feasibility of evolutionary inference based on the macrostructure of entire genomes, rather than on the traditional comparison of homologous versions of a single gene in different organisms. The mathematical modeling of evolution at the genomic level, however, and the associated inferential apparatus are qualitatively different from the usual sequence comparison theory developed to study evolution at the level of individual gene sequences. We describe the construction of a database of 16 mitochondrial gene orders from fungi and other eukaryotes by using complete or nearly complete genomic sequences; propose a measure of gene order rearrangement based on the minimal set of chromosomal inversions, transpositions, insertions, and deletions necessary to convert the order in one genome to that of the other; report on algorithm design and the development of the DERANGE software for the calculation of this measure; and present the results of analyzing the mitochondrial data with the aid of this tool.Evolutionary inference based on DNA sequences traditionally compares homologous versions of a single gene in different organisms. These comparisons are generally reliable indicators of phylogenetic relationships, even for very divergent organisms, but are limited in being based on point mutations only. In particular, homology between related mitochondrial genes may become difficult to distinguish from noise levels due to rapid nucleotide substitution (1), and this is not the only context in which the degree of sequence homology between genes having common origin is not a useful measure. Availability ofcomplete nucleotide sequence for organellar genomes suggests the possibility of inferring phylogenetic distances from their gene orders instead offrom sequences of individual genes (2). Analyses ofevolution at the genome level necessarily differ from sequence comparisons of individual genes. Though the processes of insertion and deletion of sequence elements have direct counterparts at the genomic level, the predominant process, nucleotide substitution, does not, whereas other processes assume major importance, such as the transposition of a segment from one region of a chromosome to another or the inversion of a chromosomal segment. Here we propose a quantitative analysis of transposition, inversion, and insertion/deletion, leading to the reconstruction of a mitochondrial phylogeny.Though the inference of evolutionary history through genomic rearrangements is well-established (3-5), it has been the goal of our work to define a general edit distance that combines a variety of order-disrupting events, to devise and implement a combinatorial algorithm capable of estimating this distance, and to apply these tools in a uniform way across a wide spectrum of eukaryotic organisms to generate input suitable for phylogenetic tree construction methods. Our results generally agree with evolutionary relationships inferred from gene...

Genome-wide association (GWA) studies offer a powerful unbiased method for the identification of multiple susceptibility genes for complex diseases. Here we report the results of a GWA study for Crohn's disease (CD) using family trios from the Quebec Founder Population (QFP). Haplotype-based association analyses identified multiple regions associated with the disease that met the criteria for genome-wide significance, with many containing a gene whose function appears relevant to CD. A proportion of these were replicated in two independent German Caucasian samples, including the established CD loci NOD2 and IBD5. The recently described IL23R locus was also identified and replicated. For this region, multiple individuals with all major haplotypes in the QFP were sequenced and extensive fine mapping performed to identify risk and protective alleles. Several additional loci, including a region on 3p21 containing several plausible candidate genes, a region near JAKMIP1 on 4p16.1, and two larger regions on chromosome 17 were replicated. Together with previously published loci, the spectrum of CD genes identified to date involves biochemical networks that affect epithelial defense mechanisms, innate and adaptive immune response, and the repair or remodeling of tissue.haplotype ͉ complex disease ͉ IL23R C rohn's disease (CD) is a chronic inflammatory bowel disease characterized by transmural inflammatory lesions that can affect the entire gastrointestinal tract (1). The lifetime prevalence is 0.5-1% in Caucasian populations (2) and reflects the combined effects of genetic predisposition and environmental factors (3). Genetic linkage and candidate gene approaches (4-14) have contributed to the elucidation of loci influencing genetic susceptibility to CD. More recently, genome-wide association (GWA) studies (15-21) have provided further insight into the molecular pathogenesis of the disease. The top candidate genes or loci that consistently replicate include NOD2, IL23R, ATG16L1, the IBD5 region on chromosome 5q31, and a region on 5p13.1 near the PTGER4 gene. The nature of these genes suggests that the major genetic risk factors for CD are involved in the innate immune response and destruction of intracellular bacteria.It is now clear that GWA studies provide a powerful and robust new tool for the identification of the multiple susceptibility alleles involved in complex diseases. Importantly, these types of studies have the ability to identify genes that impart only moderate increases in risk (21,22). However, most studies performed to date have identified only a few top signals, and the validation of true association among signals with lower statistical significance remains a challenge. In addition, most of the GWA studies to date have been performed by using general populations, for which very large sample sizes are required for success. They have also largely relied on single-marker analysis, with genome-wide haplotype-based association analyses receiving little attention.In early 2004, we conducted a GWA study for CD...

Many tRNA UAALeu genes from plastids contain a group I intron. An intron is also inserted in the same gene at the same position in cyanobacteria, the bacterial progenitors of plastids, suggesting an ancient bacterial origin for this intron. A group I intron has also been found in the tRNA fMet gene of some cyanobacteria but not in plastids, suggesting a more recent origin for this intron. In this study, we investigate the phylogenetic distributions of the two introns among cyanobacteria, from the earliest branching to the more derived species. The phylogenetic distribution of the tRNA UAA Leu intron follows the clustering of rRNA sequences, being either absent or present in clades of closely related species, with only one exception in the Pseudanabaena group. Our data support the notion that the tRNA UAA Leu intron was inherited by cyanobacteria and plastids through a common ancestor. Conversely, the tRNA fMet intron has a sporadic distribution, implying that many gains and losses occurred during cyanobacterial evolution. Interestingly, a phylogenetic tree inferred from intronic sequences clearly separates the different tRNA introns, suggesting that each family has its own evolutionary history.Ever since their discovery, the origin of introns has been a subject of controversy. One view, the introns-late hypothesis, proposes that introns are recent invaders and that split genes arose by late insertion of introns into originally uninterrupted genes (28). In that scenario, horizontal transfer and transposition of introns are frequent events, accounting for the scattered phylogenetic distribution of introns. Although the debate has focused on spliceosomal introns, such a scenario could apply as well to other types of introns, some of which are known to be mobile (22). In contrast, the introns-early view implies that introns are very ancient, being present in the progenote (universal ancestor) (7). The demonstration that some members of group I and group II introns are capable of in vitro autocatalytic activity (19,29,39) lends further support to the presence of these introns at an early stage of evolution, maybe as early as the putative precellular RNA world (13). In such a scenario, the observed phylogenetic distribution of introns could be explained by multiple losses in different lineages during evolution (7) and by their mobility, which is assumed to be a derived feature (2). A major obstacle for the introns-early hypothesis was the apparent absence of introns in eubacteria, although this was tentatively rationalized by pressure to streamline the genome in rapidly dividing bacteria (7). Discovery of group I introns in bacteriophages of gram-positive and gram-negative bacteria did not help to resolve the issue, due to uncertainties concerning the origin of the bacteriophages themselves (see discussion in reference 35). The recent discovery of both group I and group II introns in divergent eubacteria (4,11,12,20,31,44) was acclaimed as a breakthrough by introns-early proponents. In most cases, however, the relatio...

The mitochondrial DNA (mtDNA) of the chytridiomycete fungus Allomyces macrogynus contains 81 G+C-rich sequence elements that are 26-79 bases long and can be folded into a unique secondary structure consisting of two stem-loops. At the primary sequence level, the conservation of these double-hairpin elements (DHEs) is variable, ranging from marginal to complete identity. Forty of these DHEs are inserted in intergenic regions, 35 in introns, and 6 in variable regions of rRNA genes. Ten DHEs are inserted into other DHE elements (twins); two even form triplets. A comparison of DHE sequences shows that loop regions contain more sequence variation than helical regions and that the latter often contain compensatory base changes. This suggests a functional importance of the DHE secondary structure. We further identified nine DHEs in a 4-kb region of Allomyces arbusculus, a close relative of A. macrogynus. Eight of these DHEs are highly similar in sequence (90%-100%) to those in A. macrogynus, but only five are inserted at the same positions as in A. macrogynus. Interestingly, DHEs are also found in the mtDNAs of other chytridiomycetes, as well as certain zygomycete and ascomycete fungi. The overall distribution pattern of DHEs in fungal mtDNAs suggests that they are mobile elements.