Functional mapping of a trypanosome centromere by chromosome fragmentation identifies a 16-kb GC-rich transcriptional “strand-switch” domain as a major feature

Abstract

Trypanosomatids are an ancient family that diverged from the main eukaryotic lineage early in evolution, which display several unique features of gene organization and expression. Although genome sequencing is now complete, the nature of centromeres in these and other parasitic protozoa has not been resolved. Here, we report the functional mapping of a centromere in the American trypanosome, Trypanosoma cruzi, a parasite with an unusual mechanism of genetic exchange that involves the generation of aneuploidy by nuclear hybridization. Using a telomere-associated chromosome fragmentation approach, we show that the region required for the mitotic stability of chromosome 3 encompasses a transcriptional "strand-switch" domain constituted by a 16-kb GC-rich island. The domain contains several degenerate retrotransposon-like insertions, but atypically, lacks the arrays of satellite repeats normally associated with centromeric regions. This unusual type of organization may represent a paradigm for centromeres in T. cruzi and other primitive eukaryotes.

Chromosome-Wide Analysis of Gene Function by RNA Interference in the African Trypanosome

“…However, two of the smaller polycistrons fall within a region located between kbp 730 and 810; all eight ORFs sampled in this region gave no phenotype. This region is syntenic with a putative centromere recently described for T. cruzi (29), suggesting that the centromeric region of chromosome I of T. brucei resides here. As the analysis is limited to chromosome I, any potential functional clustering within the other megabase chromosomes of T. brucei is not addressed by the present study and must await further work.…”

“…Hence, phenotype-associated ORFs are not strongly clustered on chromosome I. Two of the smaller polycistrons located between kbp 730 and 810 fall within a region displaying synteny with a potential centromeric region identified in T. cruzi (29). None of the eight ORFs sampled from these polycistrons had a phenotype, and microarray analysis did not reveal detectable transcription in this region (S. Melville et al, personal communication), suggesting that this region of chromosome I is unlikely to encode protein products.…”

“…Both the great evolutionary divergence and novel functions likely contribute to this aspect; the molecular systems mediating antigenic variation, kinetoplast/basal body assembly, chromosomal segregation, and intracellular signaling may all require the activity of novel protein factors that are not conserved across the eukaryota. Major noncoding features of the trypanosome genome also remain cryptic, including centromeres (29) and RNA maturation and turnover motifs (23). Experimental confirmation of in silico predicted gene models is thus highly important for defining the functional potential within the trypanosome genome.…”

Histone Acetylation and Methylation at Sites Initiating Divergent Polycistronic Transcription in Trypanosoma cruzi

“…The importance of the SSRs is reflected by the rarity of sequence polymorphisms within and around SSRs on chromosome I in T. brucei (41). Also centromere function has been assigned to a SSR on T. cruzi chromosome 3 (42). However, genome analysis shows that although gene clusters are syntenic between the trypanosomatid parasites, a surprisingly high number of breaks of synteny occur at or close to SSRs (1).…”

The Genome of the Kinetoplastid Parasite, Leishmania major

“…The H2B, H3, and H4 variants appear to be novel, but may have roles in gene silencing, gene expression, DNA repair, and centromere function. Centromeres have been reported in T. cruzi (17), but no homologs were found in the Tritryps for CenH3, which is required for kinetochore assembly during mitosis. The Tritryp genomes encode a number of enzymes involved in histone modification (table S4) that may influence transcription, replication, repair, and recombination.…”