Evidence against a Human Cell-Specific Role for LRP6 in Anthrax Toxin Entry

Patricia L Ryan,John A T Young,Debbie Fox

doi:10.1371/journal.pone.0001817

Patricia L Ryan, John A T Young + Show 1 more

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https://doi.org/10.1371/journal.pone.0001817

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Abstract

The role of the cellular protein LRP6 in anthrax toxin entry is controversial. Previous studies showed that LRP6 was important for efficient intoxication of human M2182 prostate carcinoma cells but other studies performed with cells from gene-knockout mice demonstrated no role for either LRP6 or the related LRP5 protein in anthrax toxin entry. One possible explanation for this discrepancy is that LRP6 may be important for anthrax toxin entry into human, but not mouse, cells. To test this idea we have investigated the effect of knocking down LRP6 or LRP5 expression with siRNAs in human HeLa cells. We show here that efficient knockdown of either LRP6, LRP5, or both proteins has no influence on the kinetics of anthrax lethal toxin entry or MEK1 substrate cleavage in these cells. These data argue against a human-specific role for LRP6 in anthrax toxin entry and suggest instead that involvement of this protein may be restricted to certain cell types independently of their species of origin.

Highlights

Bacillus anthracis, the etiological agent of anthrax, secretes a tripartite toxin, which is one of two major virulence factors
Anthrax toxin is composed of the receptor binding moiety, protective antigen (PA), and two catalytic moieties: Lethal factor (LF), a zinc –dependent metalloprotease that cleaves MAP kinase kinases (MAPKKs), [1,2,3] and edema factor (EF), a calcium- and calmodulin-dependent adenylate cyclase that raises cAMP levels [4]
In this report we show that siRNA-mediated knockdown of LRP6 and/or LRP5 levels has no impact on the kinetics of anthrax toxin entry into human HeLa cells

Summary

Introduction

The etiological agent of anthrax, secretes a tripartite toxin, which is one of two major virulence factors. SiRNA-mediated knockdown of LRP6 levels in these cells reduced their toxin sensitivity by several orders of magnitude, an effect that was partially overcome by expression of an siRNA-resistant form of LRP6. LRP6 deficiency in M2182 cells was associated with reduced levels of PA binding and pore formation, and expression of a dominant-negative form of LRP6, lacking its cytoplasmic domain, rendered these cells resistant to intoxication.

Results

Conclusion