Bacterial esterases reverse lipopolysaccharide ubiquitylation to block host immunity.

Aspects of how Burkholderia escape the host's intrinsic immune response to replicate in the cell cytosol remain enigmatic. Here, we show that Burkholderia has evolved two mechanisms to block the activity of Ring finger protein 213 (RNF213)-mediated non-canonical ubiquitylation of bacterial lipopolysaccharide (LPS), thereby preventing the initiation of antibacterial autophagy. First, Burkholderia's polysaccharide capsule blocks RNF213 association with bacteria and second, the Burkholderia deubiquitylase (DUB), TssM, directly reverses the activity of RNF213 through a previously unrecognized esterase activity. Structural analysis provides insight into the molecular basis of TssM esterase activity, allowing it to be uncoupled from its isopeptidase function. Furthermore, a putative TssM homolog also displays esterase activity and removes ubiquitin from LPS, establishing this as a virulence mechanism. Of note, we also find that additional immune-evasion mechanisms exist, revealing that overcoming this arm of the host's immune response is critical to the pathogen.

Dedicated bacterial esterases reverse lipopolysaccharide ubiquitylation to block immune sensing.

Pathogenic bacteria have evolved diverse mechanisms to counteract cell-autonomous immunity, which otherwise guards both immune and non-immune cells from the onset of an infection1,2. The versatile immunity protein Ring finger protein 213 (RNF213)3-6 mediates the non-canonical ester-linked ubiquitylation of lipopolysaccharide (LPS), marking bacteria that sporadically enter the cytosol for destruction by antibacterial autophagy4. However, whether cytosol-adapted pathogens are ubiquitylated on their LPS and whether they escape RNF213-mediated immunity, remains unknown. Here we show that Burkholderia deubiquitylase (DUB), TssM7-9, is a potent esterase that directly reverses the ubiquitylation of LPS. Without TssM, cytosolic Burkholderia became coated in polyubiquitin and autophagy receptors in an RNF213-dependent fashion. Whereas the expression of TssM was sufficient to enable the replication of the non-cytosol adapted pathogen Salmonella, we demonstrate that Burkholderia has evolved a multi-layered defence system to proliferate in the host cell cytosol, including a block in antibacterial autophagy10-12. Structural analysis provided insight into the molecular basis of TssM esterase activity, allowing it to be uncoupled from isopeptidase function. TssM homologs conserved in another Gram-negative pathogen also reversed non-canonical LPS ubiquitylation, establishing esterase activity as a bacterial virulence mechanism to subvert host cell-autonomous immunity.