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Genomic organization and expression profile of the mucin-associated surface protein (masp) family of the human pathogen Trypanosoma cruzi.

Bartholomeu DC, Cerqueira GC, Leão AC, daRocha WD, Pais FS, Macedo C, Djikeng A, Teixeira SM, El-Sayed NM - Nucleic Acids Res. (2009)

Bottom Line: The predicted gene products, mucin-associated surface proteins (MASPs), are characterized by highly conserved N- and C-terminal domains and a strikingly variable and repetitive central region.Immunofluorescence assays using antibodies generated against a MASP peptide reveals that the expression of particular MASPs at the cell membrane is limited to subsets of the parasite population.Western blots of phosphatidylinositol-specific phospholipase C (PI-PLC)-treated parasites suggest that MASP may be GPI-anchored and shed into the medium culture, thus contributing to the large repertoire of parasite polypeptides that are exposed to the host immune system.

View Article: PubMed Central - PubMed

Affiliation: Department of Parasitology, Federal University of Minas Gerais, Belo Horizonte, MG, Brazil. daniella@icb.ufmg.br

ABSTRACT
A novel large multigene family was recently identified in the human pathogen Trypanosoma cruzi, causative agent of Chagas disease, and corresponds to approximately 6% of the parasite diploid genome. The predicted gene products, mucin-associated surface proteins (MASPs), are characterized by highly conserved N- and C-terminal domains and a strikingly variable and repetitive central region. We report here an analysis of the genomic organization and expression profile of masp genes. Masps are not randomly distributed throughout the genome but instead are clustered with genes encoding mucin and other surface protein families. Masp transcripts vary in size, are preferentially expressed during the trypomastigote stage and contain highly conserved 5' and 3' untranslated regions. A sequence analysis of a trypomastigote cDNA library reveals the expression of multiple masp variants with a bias towards a particular masp subgroup. Immunofluorescence assays using antibodies generated against a MASP peptide reveals that the expression of particular MASPs at the cell membrane is limited to subsets of the parasite population. Western blots of phosphatidylinositol-specific phospholipase C (PI-PLC)-treated parasites suggest that MASP may be GPI-anchored and shed into the medium culture, thus contributing to the large repertoire of parasite polypeptides that are exposed to the host immune system.

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MASP conserved domains and mRNA expression during the T. cruzi life cycle. (A) The coding and flanking regions of the 771 MASP members containing both the N- and C-terminal conserved domains were aligned as in Figure 4. The mRNA structure is depicted below the graph with the coding region represented by a red rectangule and the UTRs by thin lines. The polyadenylation site is indicated with an arrow. (B) Northern blot containing total RNA (10 μg) isolated from epimastigote (E), trypomastigote (T) and amastigote (A) forms was probed with the first 330 nt of the 3′ UTR. The blot was stripped and re-probed with a fragment containing part of the T. cruzi 24Sα rRNA gene.
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Figure 5: MASP conserved domains and mRNA expression during the T. cruzi life cycle. (A) The coding and flanking regions of the 771 MASP members containing both the N- and C-terminal conserved domains were aligned as in Figure 4. The mRNA structure is depicted below the graph with the coding region represented by a red rectangule and the UTRs by thin lines. The polyadenylation site is indicated with an arrow. (B) Northern blot containing total RNA (10 μg) isolated from epimastigote (E), trypomastigote (T) and amastigote (A) forms was probed with the first 330 nt of the 3′ UTR. The blot was stripped and re-probed with a fragment containing part of the T. cruzi 24Sα rRNA gene.

Mentions: An alignment of the 5′ and 3′ flanking regions from all 771 masp intact genes revealed that both regions are highly conserved (Figure 5A). Based on sequence similarity with two expressed sequence tags (ESTs) derived from an amastigote cDNA library (accession number CB923887 and S.M.R. Teixeira, unpublished data) we were able to map the polyadenylation addition site (Figure 5A) and to verify that the 3′ flanking conserved region is part of the masp 3′ UTR. This conserved region has been previously described as the repetitive TcIRE element (24), which in fact corresponds to the reverse complement of the MASP C-terminal region and the 3′ UTR conserved sequences. Consistent with our masp chromosomal location results, hybridization of PFGE blots with the TcIRE probe showed that this sequence is located in high molecular weight chromosome bands (24).Figure 5.


Genomic organization and expression profile of the mucin-associated surface protein (masp) family of the human pathogen Trypanosoma cruzi.

Bartholomeu DC, Cerqueira GC, Leão AC, daRocha WD, Pais FS, Macedo C, Djikeng A, Teixeira SM, El-Sayed NM - Nucleic Acids Res. (2009)

MASP conserved domains and mRNA expression during the T. cruzi life cycle. (A) The coding and flanking regions of the 771 MASP members containing both the N- and C-terminal conserved domains were aligned as in Figure 4. The mRNA structure is depicted below the graph with the coding region represented by a red rectangule and the UTRs by thin lines. The polyadenylation site is indicated with an arrow. (B) Northern blot containing total RNA (10 μg) isolated from epimastigote (E), trypomastigote (T) and amastigote (A) forms was probed with the first 330 nt of the 3′ UTR. The blot was stripped and re-probed with a fragment containing part of the T. cruzi 24Sα rRNA gene.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2691823&req=5

Figure 5: MASP conserved domains and mRNA expression during the T. cruzi life cycle. (A) The coding and flanking regions of the 771 MASP members containing both the N- and C-terminal conserved domains were aligned as in Figure 4. The mRNA structure is depicted below the graph with the coding region represented by a red rectangule and the UTRs by thin lines. The polyadenylation site is indicated with an arrow. (B) Northern blot containing total RNA (10 μg) isolated from epimastigote (E), trypomastigote (T) and amastigote (A) forms was probed with the first 330 nt of the 3′ UTR. The blot was stripped and re-probed with a fragment containing part of the T. cruzi 24Sα rRNA gene.
Mentions: An alignment of the 5′ and 3′ flanking regions from all 771 masp intact genes revealed that both regions are highly conserved (Figure 5A). Based on sequence similarity with two expressed sequence tags (ESTs) derived from an amastigote cDNA library (accession number CB923887 and S.M.R. Teixeira, unpublished data) we were able to map the polyadenylation addition site (Figure 5A) and to verify that the 3′ flanking conserved region is part of the masp 3′ UTR. This conserved region has been previously described as the repetitive TcIRE element (24), which in fact corresponds to the reverse complement of the MASP C-terminal region and the 3′ UTR conserved sequences. Consistent with our masp chromosomal location results, hybridization of PFGE blots with the TcIRE probe showed that this sequence is located in high molecular weight chromosome bands (24).Figure 5.

Bottom Line: The predicted gene products, mucin-associated surface proteins (MASPs), are characterized by highly conserved N- and C-terminal domains and a strikingly variable and repetitive central region.Immunofluorescence assays using antibodies generated against a MASP peptide reveals that the expression of particular MASPs at the cell membrane is limited to subsets of the parasite population.Western blots of phosphatidylinositol-specific phospholipase C (PI-PLC)-treated parasites suggest that MASP may be GPI-anchored and shed into the medium culture, thus contributing to the large repertoire of parasite polypeptides that are exposed to the host immune system.

View Article: PubMed Central - PubMed

Affiliation: Department of Parasitology, Federal University of Minas Gerais, Belo Horizonte, MG, Brazil. daniella@icb.ufmg.br

ABSTRACT
A novel large multigene family was recently identified in the human pathogen Trypanosoma cruzi, causative agent of Chagas disease, and corresponds to approximately 6% of the parasite diploid genome. The predicted gene products, mucin-associated surface proteins (MASPs), are characterized by highly conserved N- and C-terminal domains and a strikingly variable and repetitive central region. We report here an analysis of the genomic organization and expression profile of masp genes. Masps are not randomly distributed throughout the genome but instead are clustered with genes encoding mucin and other surface protein families. Masp transcripts vary in size, are preferentially expressed during the trypomastigote stage and contain highly conserved 5' and 3' untranslated regions. A sequence analysis of a trypomastigote cDNA library reveals the expression of multiple masp variants with a bias towards a particular masp subgroup. Immunofluorescence assays using antibodies generated against a MASP peptide reveals that the expression of particular MASPs at the cell membrane is limited to subsets of the parasite population. Western blots of phosphatidylinositol-specific phospholipase C (PI-PLC)-treated parasites suggest that MASP may be GPI-anchored and shed into the medium culture, thus contributing to the large repertoire of parasite polypeptides that are exposed to the host immune system.

Show MeSH
Related in: MedlinePlus