Genomic variation of Trypanosoma cruzi: involvement of multicopy genes

Abstract

By using improved pulsed field gel conditions, the karyotypes of several strains of the protozoan parasite Trypanosoma cruzi were analyzed and compared with those of Leishmania major and two other members of the genus Trypanosoma. There was no difference in chromosome migration patterns between different life cycle stages of the T. cruzi strains analyzed. However, the sizes and numbers of chromosomal bands varied considerably among T. cruzi strains. This karyotype variation among T. cruzi strains was analyzed further at the chromosomal level by using multicopy genes as probes in Southern hybridizations. The chromosomal location of the genes encoding a-and 13-tubulin, ubiquitin, rRNA, spliced leader RNA, and an 85-kilodalton protein remained stable during developmental conversion of the parasite. The sizes and numbers of chromosomes containing these sequences varied among the different strains analyzed, implying multiple rearrangements of these genes during evolution of the parasites. During continuous in vitro cultivation of T. cruzi Y, the chromosomal location of the spliced leader gene shifted spontaneously. The spliced leader gene encodes a 35-nucleotide RNA that is spliced in trans from a 105-nucleotide donor RNA onto all mRNAs in T. cruzi. The spliced leader sequences changed in their physical location in both the cloned and uncloned Y strains. Associated with the complex changes was an increase in the infectivity of the rearranged variant for tissue culture cells. Our results indicate that the spliced leader gene clusters in T. cruzi undergo high-frequency genomic rearrangements.

Chromosomal size conservation through the cell cycle supports karyotype stability in Trypanosoma cruzi

“…In this work, we show that T. cruzi karyotype is stable at the level of individual chromosomal size during the exponential and stationary phases of growth, in the cell cycle and during differentiation from epimastigotes to trypomastigotes. These observations are consistent with previous reports showing that the molecular karyotype is constant after successive and prolonged passages of 'vitro' cultures [20].…”

Identification of a novel large extrachromosomal DNA (LED) in the Trypanosomatidae

“…2B). In previous karyotypic studies of these parasites, no DNA smaller than the 270 kbp L. ma/or chromosome could be detected under pulsed-field conditions which separated linear DNA (Wagner and So, 1990). When we modified the parameters for pulsed-field separation we observed EtBrstainable material in each of the Trypanosomatidae migrating in front of and away from the compression zone where the linear chromosomes remained.…”

“…We extended our previous pulsed-field studies of the chromosomes of T. cruzi and related Kinetoptastida (Fig. 1;Wagner and So, 1990). To determine whether low-molecular-weight DNA structures exist in these parasites, we used an electrophoresis method initially developed to separate covalently closed circular DNA from the linear chromosomes of virus-infected cells (Carroll et ai, 1987).…”

“…1989); RNA editing Shaw, 1989: Stuart, 1989); the existence of the kinetoplast DNA (kDNA), a complicated network of maxi-and minicircular DNAs (Stuart, 1989;Simpson, 1986) and the astonishing plasticity of the karyotype among members of the same species (Aymerich and Goldenberg, 1989;Engman et al. 1987;Gibson and Miles, 1986;lovannisci and Beverley, 1989;Stuart etal.. 1989;Van der Ploeg etai, 1984;Wagner and So, 1990).…”

“…As in many trypanosomatid species, the size and number of chromosomes vary among different T. cruz; strains. These chromosomes range in size from 500 to 1600 kbp (Aymerich and Goldenberg, 1989;Gibson and Miles, 1986;Engman et al 1987;Wagner and So, 1990). Minichromosomes between 50 and 100 kbp have been observed in Trypanosoma brucei (Van der Ptoeg ef a/., 1984).…”