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Salmonella type III effector SopB modulates host cell exocytosis.

Perrett CA, Zhou D - Emerg Microbes Infect (2013)

Bottom Line: Type III effectors interact with the host cell endocytic pathway to aid replication.We investigated whether Salmonella effector proteins may also interact with the host's exocytic pathway.The 4-phosphatase activity of SopB was crucial to its effect on exocytosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Purdue University , West Lafayette, IN 47907, USA.

ABSTRACT
Salmonella enterica pathogenesis is dependent on its ability to enter and replicate inside host cells. Replication occurs inside the Salmonella-containing vacuole (SCV), a vacuolar compartment that is modified by bacterial effectors secreted through the two type III secretion systems (T3SS-1 and T3SS-2). Type III effectors interact with the host cell endocytic pathway to aid replication. We investigated whether Salmonella effector proteins may also interact with the host's exocytic pathway. A secreted alkaline phosphatase (SEAP) assay indicated three Salmonella effectors inhibited the secretory pathway, although only Salmonella outer protein B (SopB) was confirmed to block exocytosis using a vesicular stomatitis virus glycoprotein-green fluorescent protein (VSVG-GFP) transport assay. The 4-phosphatase activity of SopB was crucial to its effect on exocytosis. The interaction with the secretory pathway could potentially be important for providing replicating Salmonella with nutrients, contributing membrane material necessary for SCV biogenesis, altering antibacterial peptide/protein secretion or manipulating cell surface proteins important in the host response to infection.

No MeSH data available.


Related in: MedlinePlus

The 4-phosphatase activity is necessary for SopB inhibition of exocytosis. SEAP assays were performed on 293T cells transfected with pEGFP-N2 plasmids encoding the indicated forms of SopB. sopB mutation abbreviations: C460S (C/S), R468A (R/A), and K530A (K/A). The protein secretion index is displayed. Values are the mean±SD for three independent experiments. Asterisks indicate a statistical significant difference compared to vector.
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fig2: The 4-phosphatase activity is necessary for SopB inhibition of exocytosis. SEAP assays were performed on 293T cells transfected with pEGFP-N2 plasmids encoding the indicated forms of SopB. sopB mutation abbreviations: C460S (C/S), R468A (R/A), and K530A (K/A). The protein secretion index is displayed. Values are the mean±SD for three independent experiments. Asterisks indicate a statistical significant difference compared to vector.

Mentions: Among the three Salmonella type III effectors tested, SopB appeared to be the most potent in inhibiting SEAP secretion. We decided to test whether the phosphatidylinositol phosphatase activity of SopB was required for this effect. Point mutations were made in catalytic residues known to be essential for the phosphatase activity of SopB. Mutation of the 420 cysteine (C420S; C/S) or mutation of the 468 arginine (R468A; R/A), both of which are conserved between E. coli and Shigella IpgD proteins and human inositol phosphatase sequences, have been shown to be essential for 4-phosphatase activity.6,16 The lysine residue at position 530 lies within the putative synaptojanin-homology domain that is essential for the 5-phosphatase activity of SopB.17 When examined in the SEAP secretion assay, mutation of either C420 or R468 restores SEAP secretion to levels comparable to the empty vector indicating the 4-phosphatase region but not the 5-phosphatase domain is necessary for SopB inhibition of exocytosis (Figure 2).


Salmonella type III effector SopB modulates host cell exocytosis.

Perrett CA, Zhou D - Emerg Microbes Infect (2013)

The 4-phosphatase activity is necessary for SopB inhibition of exocytosis. SEAP assays were performed on 293T cells transfected with pEGFP-N2 plasmids encoding the indicated forms of SopB. sopB mutation abbreviations: C460S (C/S), R468A (R/A), and K530A (K/A). The protein secretion index is displayed. Values are the mean±SD for three independent experiments. Asterisks indicate a statistical significant difference compared to vector.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3675406&req=5

fig2: The 4-phosphatase activity is necessary for SopB inhibition of exocytosis. SEAP assays were performed on 293T cells transfected with pEGFP-N2 plasmids encoding the indicated forms of SopB. sopB mutation abbreviations: C460S (C/S), R468A (R/A), and K530A (K/A). The protein secretion index is displayed. Values are the mean±SD for three independent experiments. Asterisks indicate a statistical significant difference compared to vector.
Mentions: Among the three Salmonella type III effectors tested, SopB appeared to be the most potent in inhibiting SEAP secretion. We decided to test whether the phosphatidylinositol phosphatase activity of SopB was required for this effect. Point mutations were made in catalytic residues known to be essential for the phosphatase activity of SopB. Mutation of the 420 cysteine (C420S; C/S) or mutation of the 468 arginine (R468A; R/A), both of which are conserved between E. coli and Shigella IpgD proteins and human inositol phosphatase sequences, have been shown to be essential for 4-phosphatase activity.6,16 The lysine residue at position 530 lies within the putative synaptojanin-homology domain that is essential for the 5-phosphatase activity of SopB.17 When examined in the SEAP secretion assay, mutation of either C420 or R468 restores SEAP secretion to levels comparable to the empty vector indicating the 4-phosphatase region but not the 5-phosphatase domain is necessary for SopB inhibition of exocytosis (Figure 2).

Bottom Line: Type III effectors interact with the host cell endocytic pathway to aid replication.We investigated whether Salmonella effector proteins may also interact with the host's exocytic pathway.The 4-phosphatase activity of SopB was crucial to its effect on exocytosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Purdue University , West Lafayette, IN 47907, USA.

ABSTRACT
Salmonella enterica pathogenesis is dependent on its ability to enter and replicate inside host cells. Replication occurs inside the Salmonella-containing vacuole (SCV), a vacuolar compartment that is modified by bacterial effectors secreted through the two type III secretion systems (T3SS-1 and T3SS-2). Type III effectors interact with the host cell endocytic pathway to aid replication. We investigated whether Salmonella effector proteins may also interact with the host's exocytic pathway. A secreted alkaline phosphatase (SEAP) assay indicated three Salmonella effectors inhibited the secretory pathway, although only Salmonella outer protein B (SopB) was confirmed to block exocytosis using a vesicular stomatitis virus glycoprotein-green fluorescent protein (VSVG-GFP) transport assay. The 4-phosphatase activity of SopB was crucial to its effect on exocytosis. The interaction with the secretory pathway could potentially be important for providing replicating Salmonella with nutrients, contributing membrane material necessary for SCV biogenesis, altering antibacterial peptide/protein secretion or manipulating cell surface proteins important in the host response to infection.

No MeSH data available.


Related in: MedlinePlus