The Type III Secretion Effector NleF of Enteropathogenic Escherichia coli Activates NF-κB Early during Infection

Abstract

bThe enteric pathogens enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli employ a type 3 secretion system (T3SS) to manipulate the host inflammatory response during infection. Previously, it has been reported that EPEC, in a T3SS-dependent manner, induces an early proinflammatory response through activation of NF-B via extracellular signal-regulated kinases 1 and 2 (ERK1/2) and protein kinase C (PKC). However, the activation of NF-B during infection has not yet been attributed to an effector. At later time points postinfection, NF-B signaling is inhibited through the translocation of multiple effectors, including NleE and NleC. Here we report that the highly conserved non-LEE (locus of enterocyte effacement)-encoded effector F (NleF) shows both diffuse and mitochondrial localization during ectopic expression. Moreover, NleF induces the nuclear translocation of NF-B p65 and the expression of interleukin 8 (IL-8) following ectopic expression and during EPEC infection. Furthermore, the proinflammatory activity and localization of NleF were dependent on the C-terminal amino acids LQCG. While the C-terminal domain of NleF has previously been shown to be essential for interaction with caspase-4, caspase-8, and caspase-9, the proinflammatory activity of NleF was independent of interaction with caspase-4, -8, or -9. In conclusion, EPEC, through the T3SS-dependent translocation of NleF, induces a proinflammatory response in an NF-B-dependent manner in the early stages of infection.

Tir Triggers Expression of CXCL1 in Enterocytes and Neutrophil Recruitment during Citrobacter rodentium Infection

“…rodentium translocates multiple effectors that contribute to coordinated cytoskeleton remodeling (including Map, which activates Cdc42 and Rac1 [53], EspM, which activates RhoA [54], EspT, which activates Rac1 [55] and EspJ, which inhibits Src kinases [56] that play a role in Tir tyrosine phosphorylation) and subversion of innate immune responses, including NF-B (e.g., NleC, NleD, NleE, NleB, and NleF [13,[57][58][59][60][61][62]). Importantly, the difference in neutrophil recruitment seen at day 14 between wild-type C. rodentium and C. rodentium expressing Tir_Y451A/Y471A is at the time when both infections are close to being cleared.…”

Modulation of the host innate immune and inflammatory response by translocated bacterial proteins

“…IpaH9.8 NEMO (component of the IKK complex) E3 ubiquitin ligase, ubiquitylates NEMO and targets it for degradation Ashida et al, 2010 IpaH4.5 p65 subunit of the NF-κB complex Ubiquitylates the p65 subunit of NF-κB to limit NF-κB transcription Wang et al, 2013 OspG Ubiquitin pathway Serine/threonine kinase that interacts with E2 enzymes, uses ubiquitin as a cofactor to promote kinase activity Kim et al, 2005Pruneda et al, 2014 IpaH7.8 NLRC4 inflammasome Ubiquitylates GLMN and targets it for degradation to promote inflammasome activation Suzuki et al, 2014 EHEC and EPEC NleF NF-κB and caspases 4, 8 and 9 Promotes nuclear translocation of NF-κB p65, promotes caspase-dependent cell death pathways Pallett et al, 2014Blasche et al, 2013 Human ribosomal protein S3 (RPS3) Prevents phosphorylation and activation of RPS3 Wan et al, 2011 NleC NF-κB Zinc metalloprotease that cleaves p65 to inhibit NF-κB function Yen et al, 2010Baruch et al, 2011Pearson et al, 2011Mühlen et al, 2011Sham et al, 2011 NleE TAB2 and TAB3 (NF-κB pathway) Methyltransferase that modifies TAB2 and TAB3 to prevent downstream degradation of IκB Zhang et al, Newton et al, 2010Nadler et al, 2010 NleB FADD, TRADD, RIPK1 N-acetylglucosamine (GlcNAc) transferase that disrupts death domains to prevent protein-protein interactions Li et al, 2013aPearson et al, 2013 Salmonella enterica serovar Typhimurium Helicobacter pylori CagA NF-κB, apoptosis Activates NF-κB, degrades p53 and prevents apoptosis…”

“…Interestingly, enteropathogenic and enterohemorrhagic Escherichia coli translocate T3SS substrates that can both promote and inhibit NF-κB activation. NleF is translocated into tissue culture cells early during infection, and can promote nuclear localization of NF-κB p65 (Pallett et al, 2014). NleF can also bind to caspase 8 to inhibit apoptosis (Blasche et al, 2013).…”

“…Although these activities seem distinct, there is some evidence that caspase 8 can activate NF-κB signalling (Chaudhary et al, 2000), indicating that NleF binding to caspase 8 could promote interactions with additional host signalling proteins. Following NleF translocation, NleH1, NleE and NleC are injected into host cells (Mills et al, 2008;Pallett et al, 2014), all of which inhibit NF-κB signalling. NleH1 prevents phosphorylation and activation of ribosomal protein S3 (RPS3), which can function as a non-Rel NF-κB subunit (Wan et al, 2007;.…”

Distinct Roles of the Antiapoptotic Effectors NleB and NleF from Enteropathogenic Escherichia coli

“…In addition, ectopic expression of NleF led to increased transcription of an NF-B-dependent reporter, as well as an increase in IL-8 mRNA expression. While this activity was dependent on the LQCG motif of NleF, it occurred independently of caspase involvement (25). Although both NleF and NleB1 can block FasL/FAS signaling, their distinct roles in vivo may be due to their ability to target other innate immune signaling pathways either directly or indirectly.…”