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Analysis of consensus sequence patterns in Giardia cytoskeleton gene promoters.

ABSTRACT: Protein-coding genes in the ancient eukaryote Giardia lamblia lack typical promoter consensus elements. We have analysed the immediate 5' flanking sequences of seven genes of related function (structural cytoskeleton proteins) to identify shared DNA motifs that might have a role in transcription initiation. Transcription start sites for five genes have been determined previously. Genomic mapping and mRNA primer extension experiments demonstrate additionally that the genes for beta-giardin and median body protein are (i) present as single copies in the genome, (ii) transcribed with very short 5' leader sequences. Two search algorithms designed to extract conserved motifs from either aligned or non-aligned sequences independently discovered three sites constituting a common pattern in all seven promoters. Sites were optimally aligned using weight matrix building trials to achieve the maximum 'information content'. Profiling the information content of best alignments defines the extent of the homologies as: a 9 bp box (initiator) at the start site and upstream 18 and 6 bp boxes. The initiator is the most highly conserved element and contains a universal Py-A-Pu motif at which transcription starts. We show that the best matrices can be combined in a search pattern that correctly locates transcription start sites in genomic DNA sequences.

SUBMITTER: Holberton DV

PROVIDER: S-EPMC307134 | biostudies-other | 1995 Aug

REPOSITORIES: biostudies-other

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Analysis of consensus sequence patterns in Giardia cytoskeleton gene promoters.

Holberton D V DV Marshall J J

Nucleic acids research 19950801 15

Protein-coding genes in the ancient eukaryote Giardia lamblia lack typical promoter consensus elements. We have analysed the immediate 5' flanking sequences of seven genes of related function (structural cytoskeleton proteins) to identify shared DNA motifs that might have a role in transcription initiation. Transcription start sites for five genes have been determined previously. Genomic mapping and mRNA primer extension experiments demonstrate additionally that the genes for beta-giardin and me ...[more]

PMID: 7659516

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Project description:BACKGROUND: Genetic heterogeneity has become a major inconvenience in the genotyping and molecular epidemiology of the intestinal protozoan parasite Giardia intestinalis, in particular for the major human infecting genotype, assemblage B. Sequence-based genotyping of assemblage B Giardia from patient fecal samples, where one or several of the commonly used genotyping loci (beta-giardin, triosephosphate isomerase and glutamate dehydrogenase) are implemented, is often hampered due to the presence of sequence heterogeneity in the sequencing chromatograms. This can be due to allelic sequence heterozygosity (ASH) and /or co-infections with parasites of different assemblage B sub-genotypes. Thus, two important questions have arisen; i) does ASH occur at the single cell level, and/or ii) do multiple sub-genotype infections commonly occur in patients infected with assemblage B, G. intestinalis isolates? RESULTS: We used micromanipulation in order to isolate single Giardia intestinalis, assemblage B trophozoites (GS isolate) and cysts from human patients. Molecular analysis at the tpi loci of trophozoites from the GS lineage indicated that ASH is present at the single cell level. Analyses of assemblage B Giardia cysts from clinical samples at the bg and tpi loci also indicated ASH at the single cell level. Additionally, alignment of sequence data from several different cysts that originated from the same patient yielded different sequence patterns, thus suggesting the presence of multiple sub-assemblage infections in congruence with ASH within the same patient. CONCLUSIONS: Our results conclusively show that ASH does occur at the single cell level in assemblage B Giardia. Furthermore, sequence heterogeneity generated during sequence-based genotyping of assemblage B isolates may possess the complexity of single cell ASH in concurrence with co-infections of different assemblage B sub-genotypes. These findings explain the high abundance of sequence heterogeneity commonly found when performing sequence based genotyping of assemblage B Giardia, and illuminates the necessity of developing new G. intestinalis genotyping tools.

Dataset Information

Analysis of consensus sequence patterns in Giardia cytoskeleton gene promoters.

Publications

Analysis of consensus sequence patterns in Giardia cytoskeleton gene promoters.

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