Alpha-kinase 1 is a cytosolic innate immune receptor for bacterial ADP-heptose

Abstract

Immune recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors often activates proinflammatory NF-κB signalling. Recent studies indicate that the bacterial metabolite D-glycero-β-D-manno-heptose 1,7-bisphosphate (HBP) can activate NF-κB signalling in host cytosol, but it is unclear whether HBP is a genuine PAMP and the cognate pattern recognition receptor has not been identified. Here we combined a transposon screen in Yersinia pseudotuberculosis with biochemical analyses and identified ADP-β-D-manno-heptose (ADP-Hep), which mediates type III secretion system-dependent NF-κB activation and cytokine expression. ADP-Hep, but not other heptose metabolites, could enter host cytosol to activate NF-κB. A CRISPR-Cas9 screen showed that activation of NF-κB by ADP-Hep involves an ALPK1 (alpha-kinase 1)-TIFA (TRAF-interacting protein with forkhead-associated domain) axis. ADP-Hep directly binds the N-terminal domain of ALPK1, stimulating its kinase domain to phosphorylate and activate TIFA. The crystal structure of the N-terminal domain of ALPK1 and ADP-Hep in complex revealed the atomic mechanism of this ligand-receptor recognition process. HBP was transformed by host adenylyltransferases into ADP-heptose 7-P, which could activate ALPK1 to a lesser extent than ADP-Hep. ADP-Hep (but not HBP) alone or during bacterial infection induced Alpk1-dependent inflammation in mice. Our findings identify ALPK1 and ADP-Hep as a pattern recognition receptor and an effective immunomodulator, respectively.

ADP heptose, a novel pathogen‐associated molecular pattern identified in Helicobacter pylori

“…Recognition is specific, as evidenced by the fact that the α-configurated forms elicited no response. By directly demonstrating that ADP heptose is present in gram-negative bacteria at sufficient concentrations, these results complement the recent demonstration by Zhou et al (12) that ADP heptose is the ligand for ALPK1.…”

“…After screening fractionated H. pylori lysates for their capacity to stimulate NF-κB, we identified ADP heptose by MS as a novel, potent NF-κB–activating PAMP acting via the ALPK1-TIFA signaling axis [see also Pfannkuch et al (11)]. In parallel to our studies, Zhou et al (12) recently demonstrated that ADP heptose is indeed the ligand for ALPK1. Because ADP heptose is present in most LPS-containing bacteria, this molecule likely is of relevance for numerous infections with gram-negative bacteria.…”

“…A dynamin-dependent uptake has been proposed for HBP stimulation of Jurkat cells (8), and a similar mechanism could apply in the case of ADP heptose. The study by Zhou et al (12) also suggested that HBP is converted inside the host cell to heptose-7-phosphate, which is also able to activate ALPK1. In support of this theory, we too observed NF-κB activation in cells treated with ADP heptose-7-phosphate (unpublished results).…”

Review: Helicobacter: Inflammation, immunology, and vaccines

“…4 Initially, it was reported that TIFA is responsible for H pylori T4SS-dependent proinflammatory responses via its recognition of the LPS intermediate metabolite, heptose-1,7-bisphosphate (HBP). Elegant biochemical studies showed that the N-terminal domain of ALPK1 directly binds ADP-heptose, which leads to the phosphorylation and activation of the adaptor molecule, tumor necrosis factor (TNF), a receptor-associated factor-interacting protein with a forkhead-associated domain (TIFA).…”

ADP‐heptose is a newly identified pathogen‐associated molecular pattern of Shigella flexneri

“…In conclusion, our data rule out a major contribution of βHBP in S. flexneri infection and identify ADP‐heptose as a new potent bacterial PAMP that can be detected down to a concentration of 10 −10 M. The delay in inflammatory signaling observed in response to βHBP suggested that this bacterial metabolite is not directly recognized by host defense mechanisms. At the time of finalizing this manuscript, Zhou et al confirmed this hypothesis by showing that βHBP must be converted into ADP‐heptose 7‐phosphate by host adenylyltransferase enzymes of the NMNAT family to induce inflammatory signaling. Although βHBP is a bacteria‐derived factor triggering an immune response, its classification as a genuine PAMP is challenged by the fact that it is not directly sensed via the interaction with a cognate pathogen recognition receptor.…”