Faculty Bibliography

The ciliate Tetrahymena thermophila is a model organism for molecular and cellular biology. Like other ciliates, this species has separate germline and soma functions that are embodied by distinct nuclei within a single cell. The germline-like micronucleus (MIC) has its genome held in reserve for sexual reproduction. The soma-like macronucleus (MAC), which possesses a genome processed from that of the MIC, is the center of gene expression and does not directly contribute DNA to sexual progeny. We report here the shotgun sequencing, assembly, and analysis of the MAC genome of T. thermophila, which is approximately 104 Mb in length and composed of approximately 225 chromosomes. Overall, the gene set is robust, with more than 27,000 predicted protein-coding genes, 15,000 of which have strong matches to genes in other organisms. The functional diversity encoded by these genes is substantial and reflects the complexity of processes required for a free-living, predatory, single-celled organism. This is highlighted by the abundance of lineage-specific duplications of genes with predicted roles in sensing and responding to environmental conditions (e.g., kinases), using diverse resources (e.g., proteases and transporters), and generating structural complexity (e.g., kinesins and dyneins). In contrast to the other lineages of alveolates (apicomplexans and dinoflagellates), no compelling evidence could be found for plastid-derived genes in the genome. UGA, the only T. thermophila stop codon, is used in some genes to encode selenocysteine, thus making this organism the first known with the potential to translate all 64 codons in nuclear genes into amino acids. We present genomic evidence supporting the hypothesis that the excision of DNA from the MIC to generate the MAC specifically targets foreign DNA as a form of genome self-defense. The combination of the genome sequence, the functional diversity encoded therein, and the presence of some pathways missing from other model organisms makes T. thermophila an ideal model for functional genomic studies to address biological, biomedical, and biotechnological questions of fundamental importance

Whole-genome comparisons are highly informative regarding genome evolution and can reveal the conservation of genome organization and gene content, gene regulatory elements, and presence of species-specific genes. Initial comparative genome analyses of the human malaria parasite Plasmodium falciparum and rodent malaria parasites (RMPs) revealed a core set of 4,500 Plasmodium orthologs located in the highly syntenic central regions of the chromosomes that sharply defined the boundaries of the variable subtelomeric regions. We used composite RMP contigs, based on partial DNA sequences of three RMPs, to generate a whole-genome synteny map of P. falciparum and the RMPs. The core regions of the 14 chromosomes of P. falciparum and the RMPs are organized in 36 synteny blocks, representing groups of genes that have been stably inherited since these malaria species diverged, but whose relative organization has altered as a result of a predicted minimum of 15 recombination events. P. falciparum-specific genes and gene families are found in the variable subtelomeric regions (575 genes), at synteny breakpoints (42 genes), and as intrasyntenic indels (126 genes). Of the 168 non-subtelomeric P. falciparum genes, including two newly discovered gene families, 68% are predicted to be exported to the surface of the blood stage parasite or infected erythrocyte. Chromosomal rearrangements are implicated in the generation and dispersal of P. falciparum-specific gene families, including one encoding receptor-associated protein kinases. The data show that both synteny breakpoints and intrasyntenic indels can be foci for species-specific genes with a predicted role in host-parasite interactions and suggest that, besides rearrangements in the subtelomeric regions, chromosomal rearrangements may also be involved in the generation of species-specific gene families. A majority of these genes are expressed in blood stages, suggesting that the vertebrate host exerts a greater selective pressure than the mosquito vector, resulting in the acquisition of diversity

We report the genome sequence of Theileria parva, an apicomplexan pathogen causing economic losses to smallholder farmers in Africa. The parasite chromosomes exhibit limited conservation of gene synteny with Plasmodium falciparum, and its plastid-like genome represents the first example where all apicoplast genes are encoded on one DNA strand. We tentatively identify proteins that facilitate parasite segregation during host cell cytokinesis and contribute to persistent infection of transformed host cells. Several biosynthetic pathways are incomplete or absent, suggesting substantial metabolic dependence on the host cell. One protein family that may generate parasite antigenic diversity is not telomere-associated

Duffy antigen is the receptor used by Plasmodium vivax to invade erythrocytes. Consequently, individuals lacking Duffy antigen [Fy(-)] do not develop blood-stage infections. We hypothesized that naturally exposed Fy(-) humans may develop immune responses mainly to pre-erythrocytic stages and could be used to study acquired immunity to P. vivax and to identify liver-stage antigens. We report here that antibody and IFN-gamma responses to known sporozoite antigens were significantly induced by natural exposure in Fy(-) humans, whereas responses to blood-stage antigens were significantly induced in Fy(+) humans. IFN-gamma responses to sporozoite antigens were lower in Fy(+) than in Fy(-) humans, indicating that in Fy(+) humans blood-stage infections may have suppressed T cell responses to pre-erythrocytic stages. We evaluated the immune responses to 18 novel P. vivax homologs of P. falciparum sporozoite proteins identified from the P. vivax genome sequence. Eight proteins recalled IFN-gamma responses in P. vivax-exposed but not in unexposed individuals. Of these, 3 antigens elicited IFN-gamma responses in Fy(-) but not in Fy(+) individuals. These results suggest that differential immune responses observed in naturally exposed Fy(-) and Fy(+) individuals can be exploited to identify P. vivax stage-specific antigens

Rab proteins are pivotal components of the membrane trafficking machinery in all eukaryotes. Distinct Rab proteins locate to specific endomembrane compartments and genomic studies suggest that Rab gene diversity correlates with endomembrane system complexity; for example unicellular organisms generally possess 5-20 Rab family members and the size of the repertoire increases to 25-60 in multicellular systems. Here we report 65 open reading frames from the unicellular protozoan Trichomonas vaginalis that encode distinct Rab proteins (TvRabs), indicating a family with complexity that rivals Homo sapiens in number. The detection of gene transcripts for the majority of these genes and conservation of functional motifs strongly suggests that TvRabs retain functionality and likely roles in membrane trafficking. The T. vaginalis Rab family includes orthologues of the conserved subfamilies, Rab1, Rab5, Rab6, Rab7 and Rab11, but the majority of TvRabs are not represented by orthologues in other systems and includes six novel T. vaginalis specific Rab subfamilies (A-F). The extreme size of the T. vaginalis Rab family, the presence of novel subfamilies plus the divergent nature of many TvRab sequences suggest both the presence of a highly complex endomembrane system within Trichomonas and potentially novel Rab functionality. A family of more than 65 Rab genes in a unicellular genome is unexpected, but may be a requirement for progression though an amoeboid life-cycle phase as both Dictyostelium discoideum and Entamoeba histolytica share with T. vaginalis both an amoeboid life cycle stage and very large Rab gene families

In the past few years, the area of comparative genomics of malaria parasites has begun to come of age, with the completion of genome sequencing projects of four Plasmodium species, and several functional genomics studies. A picture is emerging of a parasite genome that is highly adapted to its mammalian and vector hosts, and which uses post-transcriptional gene-silencing as one method for the control of gene expression. The genome is compartmentalized into a core of conserved housekeeping genes, sandwiched between subtelomerically located genes encoding surface antigens. Species-specific gene families shape the preference of the parasite for host cells, in addition to determining interactions with the host immune-system. Recent research has led to the description of a motif that is conserved across Plasmodium species and which plays a central role in protein export into the host cell

Despite the significance of Plasmodium vivax as the most widespread human malaria parasite and a major public health problem, gene expression in this parasite is poorly understood. To accelerate gene discovery and facilitate the annotation phase of the P. vivax genome project, we have undertaken a transcriptome approach to study gene expression in the mixed blood stages of a P. vivax field isolate. Using a cDNA library constructed from purified blood stages, we have obtained single-pass sequences for approximately 21,500 expressed sequence tags (ESTs), the largest number of transcript tags obtained so far for this species. Cluster analysis revealed that the library is highly redundant, resulting in 5407 clusters. Clustered ESTs were searched against public protein databases for functional annotation, and more than one-third showed a significant match, the majority of these to Plasmodium falciparum proteins. The most abundant clusters were to genes encoding ribosomal proteins and proteins involved in metabolism, consistent with the predominance of trophozoites in the field isolate sample. In spite of the scarcity of other parasite stages in the field isolate, we could identify genes that are expressed in rings, schizonts and gametocytes. This study should facilitate our understanding of the gene expression in P. vivax asexual stages and provide valuable data for gene prediction and annotation of the P. vivax genome sequence

A comparison of gene content and genome architecture of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major, three related pathogens with different life cycles and disease pathology, revealed a conserved core proteome of about 6200 genes in large syntenic polycistronic gene clusters. Many species-specific genes, especially large surface antigen families, occur at nonsyntenic chromosome-internal and subtelomeric regions. Retroelements, structural RNAs, and gene family expansion are often associated with syntenic discontinuities that-along with gene divergence, acquisition and loss, and rearrangement within the syntenic regions-have shaped the genomes of each parasite. Contrary to recent reports, our analyses reveal no evidence that these species are descended from an ancestor that contained a photosynthetic endosymbiont