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Expression of 6-Cys Gene Superfamily Defines Babesia bovis Sexual Stage Development within Rhipicephalus microplus

View Article: PubMed Central - PubMed

ABSTRACT

Babesia bovis, an intra-erythrocytic tick-borne apicomplexan protozoan, is one of the causative agents of bovine babesiosis. Its life cycle includes sexual reproduction within cattle fever ticks, Rhipicephalus spp. Six B. bovis 6-Cys gene superfamily members were previously identified (A, B, C, D, E, F) where their orthologues in Plasmodium parasite have been shown to encode for proteins required for the development of sexual stages. The current study identified four additional 6-Cys genes (G, H, I, J) in the B. bovis genome. These four genes are described in the context of the complete ten 6-Cys gene superfamily. The proteins expressed by this gene family are predicted to be secreted or surface membrane directed. Genetic analysis comparing the 6-Cys superfamily among five distinct B. bovis strains shows limited sequence variation. Additionally, A, B, E, H, I and J genes were transcribed in B. bovis infected tick midgut while genes A, B and E were also transcribed in the subsequent B. bovis kinete stage. Transcription of gene C was found exclusively in the kinete. In contrast, transcription of genes D, F and G in either B. bovis infected midguts or kinetes was not detected. None of the 6-Cys transcripts were detected in B. bovis blood stages. Subsequent protein analysis of 6-Cys A and B is concordant with their transcript profile. The collective data indicate as in Plasmodium parasite, certain B. bovis 6-Cys family members are uniquely expressed during sexual stages and therefore, they are likely required for parasite reproduction. Within B. bovis specifically, proteins encoded by 6-Cys genes A and B are markers for sexual stages and candidate antigens for developing novel vaccines able to interfere with the development of B. bovis within the tick vector.

No MeSH data available.


Characterization of Bbo 6-Cys proteins and 6-Cys domain.A. Schematic representation of 6-Cys family proteins. The number and relative localization of the 6-Cys domains, represented as blue boxes, the predicted signal peptides (SP) and transmembrane domains (Tm), and the number of amino acids are indicated. B. Schematic representation of the arrangement and number of the 6-Cys domains in each of the B. bovis 6-Cys proteins. The number of residues found in each domain is represented inside each box. The lengths of the inter-domain regions (IDR) in the proteins containing two 6-Cys domains are relatively similar at 163,161,159,162, 164, in the A, B, C, D and E proteins, whereas it contains 227 and 521 aa in H and G’s 6-Cys domain, respectively. Since protein F has three 6-Cys domains so it has two IDRs with lengths 164 aa and 214 aa for IDR1 and IDR2 respectively. C. Schematic representation of the predicted disulfide bonds in the B. bovis 6-Cys proteins using the Prosite program (top). Description of the three conserved domains identified in the 6-Cys domains among the ten 6-Cys B. bovis proteins using MEME analysis. Amino acids are represented using letter symbols. The highest letters represent strictly conserved residues. Font sizes are proportional to the relative frequency of each residue for each position.
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pone.0163791.g002: Characterization of Bbo 6-Cys proteins and 6-Cys domain.A. Schematic representation of 6-Cys family proteins. The number and relative localization of the 6-Cys domains, represented as blue boxes, the predicted signal peptides (SP) and transmembrane domains (Tm), and the number of amino acids are indicated. B. Schematic representation of the arrangement and number of the 6-Cys domains in each of the B. bovis 6-Cys proteins. The number of residues found in each domain is represented inside each box. The lengths of the inter-domain regions (IDR) in the proteins containing two 6-Cys domains are relatively similar at 163,161,159,162, 164, in the A, B, C, D and E proteins, whereas it contains 227 and 521 aa in H and G’s 6-Cys domain, respectively. Since protein F has three 6-Cys domains so it has two IDRs with lengths 164 aa and 214 aa for IDR1 and IDR2 respectively. C. Schematic representation of the predicted disulfide bonds in the B. bovis 6-Cys proteins using the Prosite program (top). Description of the three conserved domains identified in the 6-Cys domains among the ten 6-Cys B. bovis proteins using MEME analysis. Amino acids are represented using letter symbols. The highest letters represent strictly conserved residues. Font sizes are proportional to the relative frequency of each residue for each position.

Mentions: The predicted protein size of the ten Bbo 6-Cys ranges from 568 aa [66.73 KDa] to 1,689 aa [187.27 KDa] (Fig 2A, Table 1). All, except Bbo 6-Cys G, have signal peptide (SP), suggesting that they may be surface exposed or secreted proteins. Earlier gene modeling analysis did not predict a signal peptide for Bbo 6-Cys protein F. Additional visual analysis revealed the presence of a 23 aa hydrophobic SP prior to the three 6-Cys domains (PF07422PF07422-PTZ00360) at the N-terminus of the protein (Fig 2A and 2C, S1 Fig). Bbo 6-Cys protein C is the only member with a predicted transmembrane (Tm) domain in its N-terminus region and none of the Bbo 6-Cys proteins contain glycosyl-phosphatidylinositol anchors (GPI) (Fig 2A). This feature is in contrast to the orthologous Pf48/45 protein which is predicted to contain a GPI anchor and is located on the gamete surface although another orthologous soluble protein, Pfs230, also lacks a GPI anchor [38, 39]


Expression of 6-Cys Gene Superfamily Defines Babesia bovis Sexual Stage Development within Rhipicephalus microplus
Characterization of Bbo 6-Cys proteins and 6-Cys domain.A. Schematic representation of 6-Cys family proteins. The number and relative localization of the 6-Cys domains, represented as blue boxes, the predicted signal peptides (SP) and transmembrane domains (Tm), and the number of amino acids are indicated. B. Schematic representation of the arrangement and number of the 6-Cys domains in each of the B. bovis 6-Cys proteins. The number of residues found in each domain is represented inside each box. The lengths of the inter-domain regions (IDR) in the proteins containing two 6-Cys domains are relatively similar at 163,161,159,162, 164, in the A, B, C, D and E proteins, whereas it contains 227 and 521 aa in H and G’s 6-Cys domain, respectively. Since protein F has three 6-Cys domains so it has two IDRs with lengths 164 aa and 214 aa for IDR1 and IDR2 respectively. C. Schematic representation of the predicted disulfide bonds in the B. bovis 6-Cys proteins using the Prosite program (top). Description of the three conserved domains identified in the 6-Cys domains among the ten 6-Cys B. bovis proteins using MEME analysis. Amino acids are represented using letter symbols. The highest letters represent strictly conserved residues. Font sizes are proportional to the relative frequency of each residue for each position.
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pone.0163791.g002: Characterization of Bbo 6-Cys proteins and 6-Cys domain.A. Schematic representation of 6-Cys family proteins. The number and relative localization of the 6-Cys domains, represented as blue boxes, the predicted signal peptides (SP) and transmembrane domains (Tm), and the number of amino acids are indicated. B. Schematic representation of the arrangement and number of the 6-Cys domains in each of the B. bovis 6-Cys proteins. The number of residues found in each domain is represented inside each box. The lengths of the inter-domain regions (IDR) in the proteins containing two 6-Cys domains are relatively similar at 163,161,159,162, 164, in the A, B, C, D and E proteins, whereas it contains 227 and 521 aa in H and G’s 6-Cys domain, respectively. Since protein F has three 6-Cys domains so it has two IDRs with lengths 164 aa and 214 aa for IDR1 and IDR2 respectively. C. Schematic representation of the predicted disulfide bonds in the B. bovis 6-Cys proteins using the Prosite program (top). Description of the three conserved domains identified in the 6-Cys domains among the ten 6-Cys B. bovis proteins using MEME analysis. Amino acids are represented using letter symbols. The highest letters represent strictly conserved residues. Font sizes are proportional to the relative frequency of each residue for each position.
Mentions: The predicted protein size of the ten Bbo 6-Cys ranges from 568 aa [66.73 KDa] to 1,689 aa [187.27 KDa] (Fig 2A, Table 1). All, except Bbo 6-Cys G, have signal peptide (SP), suggesting that they may be surface exposed or secreted proteins. Earlier gene modeling analysis did not predict a signal peptide for Bbo 6-Cys protein F. Additional visual analysis revealed the presence of a 23 aa hydrophobic SP prior to the three 6-Cys domains (PF07422PF07422-PTZ00360) at the N-terminus of the protein (Fig 2A and 2C, S1 Fig). Bbo 6-Cys protein C is the only member with a predicted transmembrane (Tm) domain in its N-terminus region and none of the Bbo 6-Cys proteins contain glycosyl-phosphatidylinositol anchors (GPI) (Fig 2A). This feature is in contrast to the orthologous Pf48/45 protein which is predicted to contain a GPI anchor and is located on the gamete surface although another orthologous soluble protein, Pfs230, also lacks a GPI anchor [38, 39]

View Article: PubMed Central - PubMed

ABSTRACT

Babesia bovis, an intra-erythrocytic tick-borne apicomplexan protozoan, is one of the causative agents of bovine babesiosis. Its life cycle includes sexual reproduction within cattle fever ticks, Rhipicephalus spp. Six B. bovis 6-Cys gene superfamily members were previously identified (A, B, C, D, E, F) where their orthologues in Plasmodium parasite have been shown to encode for proteins required for the development of sexual stages. The current study identified four additional 6-Cys genes (G, H, I, J) in the B. bovis genome. These four genes are described in the context of the complete ten 6-Cys gene superfamily. The proteins expressed by this gene family are predicted to be secreted or surface membrane directed. Genetic analysis comparing the 6-Cys superfamily among five distinct B. bovis strains shows limited sequence variation. Additionally, A, B, E, H, I and J genes were transcribed in B. bovis infected tick midgut while genes A, B and E were also transcribed in the subsequent B. bovis kinete stage. Transcription of gene C was found exclusively in the kinete. In contrast, transcription of genes D, F and G in either B. bovis infected midguts or kinetes was not detected. None of the 6-Cys transcripts were detected in B. bovis blood stages. Subsequent protein analysis of 6-Cys A and B is concordant with their transcript profile. The collective data indicate as in Plasmodium parasite, certain B. bovis 6-Cys family members are uniquely expressed during sexual stages and therefore, they are likely required for parasite reproduction. Within B. bovis specifically, proteins encoded by 6-Cys genes A and B are markers for sexual stages and candidate antigens for developing novel vaccines able to interfere with the development of B. bovis within the tick vector.

No MeSH data available.