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Sialic acid expression in the mosquito Aedes aegypti and its possible role in dengue virus-vector interactions.

Cime-Castillo J, Delannoy P, Mendoza-Hernández G, Monroy-Martínez V, Harduin-Lepers A, Lanz-Mendoza H, Hernández-Hernández Fde L, Zenteno E, Cabello-Gutiérrez C, Ruiz-Ordaz BH - Biomed Res Int (2015)

Bottom Line: AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface.Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome.Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology and Biotechnology Department, Biomedical Research Institute, National University of México (UNAM), 04510 México City, Mexico.

ABSTRACT
Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission.

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Related in: MedlinePlus

SDS-PAGE assay of the glycoproteins from Ae. aegypti SG protein extracts. (a) Total carbohydrates stained with Pro-Q Emerald, where the molecular weights are shown on the right. (b) Western blot assay using ConA lectin, which binds to glycoproteins that contain mannose or glucose residues.
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fig4: SDS-PAGE assay of the glycoproteins from Ae. aegypti SG protein extracts. (a) Total carbohydrates stained with Pro-Q Emerald, where the molecular weights are shown on the right. (b) Western blot assay using ConA lectin, which binds to glycoproteins that contain mannose or glucose residues.

Mentions: To confirm the presence of total sugars in the SG protein extracts from Ae. aegypti and to characterize the putative glycoprotein(s) that may recognize DENV, we separated the SG proteins by electrophoresis and stained them to detect any carbohydrates. The SG protein extracts were transferred to nitrocellulose membranes and subjected to a western blot assay. The membrane was also incubated with ConA or SNA lectins (Figures 4(b) and 5(a), lane 9). For the control assay, we used a carbohydrate staining kit (Pro-Q Emerald 300 Glycoprotein Gel Stain Kit, Molecular Probes; Figure 4(a), lane 1), and we observed a range of glycoproteins from 29 kDa to 116 kDa, with more intense bands of 29, 45, and 66 kDa. When we incubated the SG protein extracts proteins with ConA, we observed a glycoprotein of 50–60 kDa, which has not been identified previously with the carbohydrate staining kit. We also observed an increase in the intensity of the band at 97 kDa. Therefore, these proteins could have contained mannose and glucose residues (Figure 4(b)). The interaction with SNA produced several bands that ranged from 10 to 97 kDa (Figure 5(a), lane 9), so these proteins could possess Sia motifs. In agreement, we observed no significant changes when we pretreated the SG protein extracts with sialidase (Figure 5(a), lanes 2 and 3).


Sialic acid expression in the mosquito Aedes aegypti and its possible role in dengue virus-vector interactions.

Cime-Castillo J, Delannoy P, Mendoza-Hernández G, Monroy-Martínez V, Harduin-Lepers A, Lanz-Mendoza H, Hernández-Hernández Fde L, Zenteno E, Cabello-Gutiérrez C, Ruiz-Ordaz BH - Biomed Res Int (2015)

SDS-PAGE assay of the glycoproteins from Ae. aegypti SG protein extracts. (a) Total carbohydrates stained with Pro-Q Emerald, where the molecular weights are shown on the right. (b) Western blot assay using ConA lectin, which binds to glycoproteins that contain mannose or glucose residues.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385653&req=5

fig4: SDS-PAGE assay of the glycoproteins from Ae. aegypti SG protein extracts. (a) Total carbohydrates stained with Pro-Q Emerald, where the molecular weights are shown on the right. (b) Western blot assay using ConA lectin, which binds to glycoproteins that contain mannose or glucose residues.
Mentions: To confirm the presence of total sugars in the SG protein extracts from Ae. aegypti and to characterize the putative glycoprotein(s) that may recognize DENV, we separated the SG proteins by electrophoresis and stained them to detect any carbohydrates. The SG protein extracts were transferred to nitrocellulose membranes and subjected to a western blot assay. The membrane was also incubated with ConA or SNA lectins (Figures 4(b) and 5(a), lane 9). For the control assay, we used a carbohydrate staining kit (Pro-Q Emerald 300 Glycoprotein Gel Stain Kit, Molecular Probes; Figure 4(a), lane 1), and we observed a range of glycoproteins from 29 kDa to 116 kDa, with more intense bands of 29, 45, and 66 kDa. When we incubated the SG protein extracts proteins with ConA, we observed a glycoprotein of 50–60 kDa, which has not been identified previously with the carbohydrate staining kit. We also observed an increase in the intensity of the band at 97 kDa. Therefore, these proteins could have contained mannose and glucose residues (Figure 4(b)). The interaction with SNA produced several bands that ranged from 10 to 97 kDa (Figure 5(a), lane 9), so these proteins could possess Sia motifs. In agreement, we observed no significant changes when we pretreated the SG protein extracts with sialidase (Figure 5(a), lanes 2 and 3).

Bottom Line: AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface.Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome.Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues.

View Article: PubMed Central - PubMed

Affiliation: Molecular Biology and Biotechnology Department, Biomedical Research Institute, National University of México (UNAM), 04510 México City, Mexico.

ABSTRACT
Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission.

Show MeSH
Related in: MedlinePlus