The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform

Pavithra Aravamudhan - One of the best experts on this subject based on the ideXlab platform.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein σ1 binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements for different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, CMP N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels similar to those of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface and mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    bioRxiv, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    ABSTRACT Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are unknown. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes identified as required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains differing in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Infectivity of T3SA+ was diminished to levels of T3SA-in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection. IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

Kelly Urbanek - One of the best experts on this subject based on the ideXlab platform.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein σ1 binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements for different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, CMP N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels similar to those of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface and mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    bioRxiv, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    ABSTRACT Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are unknown. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes identified as required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains differing in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Infectivity of T3SA+ was diminished to levels of T3SA-in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection. IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

Danica M Sutherland - One of the best experts on this subject based on the ideXlab platform.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein σ1 binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements for different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, CMP N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels similar to those of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface and mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    bioRxiv, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    ABSTRACT Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are unknown. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes identified as required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains differing in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Infectivity of T3SA+ was diminished to levels of T3SA-in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection. IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

Jonathan J Knowlton - One of the best experts on this subject based on the ideXlab platform.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    Journal of Virology, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    Engagement of cell surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein σ1 binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements for different cell types are not entirely known. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, CMP N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains that differ in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for reovirus infection. Following CRISPR-Cas9 disruption of either gene, cell surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Furthermore, infectivity of T3SA+ was diminished to levels similar to those of T3SA- in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection.IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface and mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

  • cytidine monophosphate n acetylneuraminic acid synthetase and Solute Carrier Family 35 member a1 are required for reovirus binding and infection
    bioRxiv, 2020
    Co-Authors: Kelly Urbanek, Danica M Sutherland, Robert C Orchard, Craig B Wilen, Jonathan J Knowlton, Pavithra Aravamudhan
    Abstract:

    ABSTRACT Engagement of cell-surface receptors by viruses is a critical determinant of viral tropism and disease. The reovirus attachment protein, σ1, binds sialylated glycans and proteinaceous receptors to mediate infection, but the specific requirements on different cell types are unknown. To identify host factors required for reovirus-induced cell death, we conducted a CRISPR-knockout screen targeting over 20,000 genes in murine microglial BV2 cells. Candidate genes identified as required for reovirus to cause cell death were highly enriched for sialic acid synthesis and transport. Two of the top candidates identified, cytidine monophosphate N-acetylneuraminic acid synthetase (Cmas) and Solute Carrier Family 35 member A1 (Slc35a1), promote sialic acid expression on the cell surface. Two reovirus strains differing in the capacity to bind sialic acid, T3SA+ and T3SA-, were used to evaluate Cmas and Slc35a1 as potential host genes required for infection. Following CRISPR-Cas9 disruption of either gene, cell-surface expression of sialic acid was diminished. These results correlated with decreased binding of strain T3SA+, which is capable of engaging sialic acid. Disruption of either gene did not alter the low-level binding of T3SA-, which does not engage sialic acid. Infectivity of T3SA+ was diminished to levels of T3SA-in cells lacking Cmas and Slc35a1 by CRISPR ablation. However, exogenous expression of Cmas and Slc35a1 into the respective null cells restored sialic acid expression and T3SA+ binding and infectivity. These results demonstrate that Cmas and Slc35a1, which mediate cell-surface expression of sialic acid, are required in murine microglial cells for efficient reovirus binding and infection. IMPORTANCE Attachment factors and receptors are important determinants of dissemination and tropism during reovirus-induced disease. In a CRISPR cell-survival screen, we discovered two genes, Cmas and Slc35a1, which encode proteins required for sialic acid expression on the cell surface, that mediate reovirus infection of microglial cells. This work elucidates host genes that render microglial cells susceptible to reovirus infection and expands current understanding of the receptors on microglial cells that are engaged by reovirus. Such knowledge may lead to new strategies to selectively target microglial cells for oncolytic applications.

Angelica Linden Hirschberg - One of the best experts on this subject based on the ideXlab platform.

  • insulin regulation of Solute Carrier Family 2 member 1 glucose transporter 1 expression and glucose uptake in decidualizing human endometrial stromal cells an in vitro study
    Reproductive Biology and Endocrinology, 2020
    Co-Authors: Ivika Jakson, Dorina Ujvari, Sebastian Gidlof, Angelica Linden Hirschberg
    Abstract:

    Solute Carrier Family 2 member 1 (SLC2A1; previously known as glucose transporter 1), is the most abundant glucose transporter in human endometrium and is up-regulated during decidualization, whereas high insulin may have a negative impact on this process. The present study aimed to investigate the effect of insulin on the expression of SLC2A1 and glucose uptake in decidualizing human endometrial stromal cells. We induced in vitro decidualization of endometrial stromal cells obtained from regularly menstruating healthy non-obese women. The cells were treated with increasing concentrations of insulin, and the involvement of the transcription factor forkhead box O1 (FOXO1) was evaluated using a FOXO1 inhibitor. SLC2A1 mRNA levels were measured by Real-Time PCR and protein levels were evaluated by immunocytochemistry. Glucose uptake was estimated by an assay quantifying the cellular uptake of radioactive glucose. One-way ANOVA, Dunnett’s multiple comparisons test and paired t-test were used to determine the statistical significance of the results. We found that insulin dose-dependently decreased SLC2A1 mRNA levels and decreased protein levels of SLC2A1 in decidualizing human endometrial stromal cells. Transcriptional inactivation of FOXO1 seems to explain at least partly the down-regulation of SLC2A1 by insulin. Glucose uptake increased upon decidualization, whereas insulin treatment resulted in a slight inhibition of the glucose uptake, although not significant for all insulin concentrations. These results indicate an impairment of decidualization by high concentrations of insulin. Future studies will determine the clinical significance of our results for endometrial function and decidualization in women with insulin resistance and hyperinsulinemia.