DPP8

DPP8 is an intracellular, non-glycosylated serine dipeptidyl aminopeptidase that cleaves N-terminal Xaa-Pro dipeptides from cytosolic substrates at neutral pH, distinguishing it from lysosomal and membrane-anchored post-proline peptidases (PMID:11012666, PMID:15039077). It is enzymatically active with kinetics comparable to DPP-IV but imposes strict selectivity for hydrophobic or basic residues at the P2 site; its activity depends on conserved C-terminal loop residues and on residues lining the S1/S2 pockets that also contribute to dimerization, though dimerization alone is insufficient for catalysis (PMID:17040910, PMID:20536396). A defining cellular role is as an N-terminal processing enzyme that exposes IAP-binding motifs on substrates such as adenylate kinase 2 (AK2) and EIF2A, licensing IAP E3 ligase-mediated proteasomal degradation, a fate counteracted by NatA-mediated N-terminal acetylation (PMID:40312560). Functionally, DPP8 (with DPP9) acts as an intracellular checkpoint restraining inflammasome activation: its pharmacological inhibition triggers gasdermin D-dependent pyroptosis through pro-caspase-1, sensed by Nlrp1b in mouse myeloid cells and by CARD8 in human myeloid cells and T lymphocytes in a cell-state-dependent manner (PMID:27820798, PMID:29396289, PMID:29967349, PMID:32796818). DPP8 additionally promotes TGF-β1/Smad-dependent epithelial-to-mesenchymal transition and fibrotic extracellular-matrix deposition (PMID:33932609, PMID:38458339), and carries catalysis-independent functions in cell migration, adhesion, and apoptosis (PMID:16704418).

1. 2000 High

   Establishing that DPP8 was a genuine post-proline-cleaving peptidase distinct in localization from the known DPP-IV answered whether a cytoplasmic enzyme could perform Xaa-Pro dipeptidase chemistry outside the lysosomal/membrane compartment.

   Evidence Recombinant expression in COS-7 cells with confocal localization and chromogenic substrate assays

   PMID:11012666

   Open questions at the time

   • No physiological substrate identified at this stage
   • Quaternary structure and active-site architecture undefined
2. 2004 High

   Purification to homogeneity and kinetic characterization established that DPP8 is intrinsically catalytically competent with DPP-IV-like turnover, settling whether activity required cellular cofactors.

   Evidence Baculovirus expression in Sf9 cells, purification, kinetic assays with multiple chromogenic substrates

   PMID:15039077

   Open questions at the time

   • Natural substrates remained unknown
   • Did not address oligomeric state requirements
3. 2006 High

   Structural and mutational dissection defined DPP8 as a dimer with a distinct P2 substrate preference and showed that specific C-terminal loop and pocket residues govern catalysis independently of dimer assembly, refining how the enzyme achieves selectivity.

   Evidence Analytical ultracentrifugation, native gels, site-directed mutagenesis, and enzyme kinetics; homology modeling

   PMID:17040910 PMID:20536396

   Open questions at the time

   • No experimental crystal structure
   • Substrate selectivity inferred from synthetic peptides, not endogenous targets
4. 2006 Medium

   Overexpression with catalytic-serine mutants revealed that DPP8 modulates cell migration, adhesion, wound healing, and apoptosis through a mechanism independent of its peptidase activity, introducing extraenzymatic roles.

   Evidence GFP-fusion overexpression with transwell/wound-healing and apoptosis assays plus catalytic mutant controls

   PMID:16704418

   Open questions at the time

   • No endogenous loss-of-function validation
   • Molecular partners mediating the non-catalytic effects unidentified
   • Single lab
5. 2016 High

   Demonstrating that DPP8/9 inhibition triggers ASC-independent, caspase-1- and gasdermin D-dependent pyroptosis defined these enzymes as suppressors of an inflammatory cell-death pathway.

   Evidence Selective inhibitors, caspase-1 knockout mice, gasdermin D cleavage immunoblots, cell-death assays

   PMID:27820798

   Open questions at the time

   • The upstream sensor was not yet identified
   • Endogenous substrate linking DPP8/9 activity to the checkpoint unknown
6. 2018 High

   Genetic epistasis identified the sensors downstream of DPP8/9 inhibition, with Nlrp1b operating in mouse myeloid cells and CARD8 in human myeloid cells, establishing species-specific inflammasome wiring of the checkpoint.

   Evidence Nlrp1b and CARD8 knockout/knockdown with selective inhibitors and caspase-1 activation assays in mouse and human cells including primary AML

   PMID:29396289 PMID:29967349

   Open questions at the time

   • Biochemical signal generated by DPP8/9 inhibition that activates the sensors not defined
   • Did not resolve why sensitivity differs across cell types
7. 2019 Medium

   Showing that all functional rodent NLRP1 alleles respond to DPP8/9 inhibition, mirroring Toxoplasma gondii sensitivity, indicated a universal NLRP1-sensed signal and a possible pathogen-mimicking mechanism.

   Evidence Overexpression of diverse NLRP1 alleles with inhibitors and comparison to T. gondii infection

   PMID:31383852

   Open questions at the time

   • Molecular identity of the shared signal unresolved
   • Single lab
8. 2020 Medium

   Extending the checkpoint to lymphocytes revealed that DPP8/9 inhibition drives CARD8-dependent pyroptosis in resting but not activated T cells, establishing a cell-state-dependent control of the pathway.

   Evidence CARD8 knockdown and selective inhibitors in resting versus activated human and rodent T cells

   PMID:32796818

   Open questions at the time

   • Mechanism conferring resistance in activated T cells unknown
   • Single lab
9. 2021 Medium

   Systematic peptide-degradation profiling demonstrated that DPP8/9 do not process most proline-containing proteasomal peptides, narrowing the enzymes' substrate scope from earlier reporter-based assumptions.

   Evidence Broad peptide degradation assays in cell lysates with inhibitors and knockout

   PMID:37982048

   Open questions at the time

   • Negative result; did not pinpoint the bona fide physiological substrates
   • Lysate context may not reflect intact-cell processing
10. 2025 High

    Identifying AK2 and EIF2A as DPP8/9 substrates whose N-terminal processing unmasks IAP-binding motifs for proteasomal degradation provided the first defined endogenous substrate mechanism, linking peptidase activity to regulated protein turnover and a degron logic counteracted by NatA acetylation.

    Evidence Co-IP, N-terminal processing and IBM mutagenesis, NatA knockdown, genome-wide in silico screen with EIF2A validation; live-cell DiPAK sensor linking DPP8/9 activity to B-cell activation

    PMID:40312560 PMID:40355159

    Open questions at the time

    • Only two of 129 candidate substrates experimentally validated
    • Link between substrate processing and the NLRP1/CARD8 checkpoint not directly established
11. 2021 Medium

    Loss-of-function with rescue placed DPP8 as a positive regulator of TGF-β1/Smad and Akt signaling driving EMT and fibrotic ECM deposition, broadening its physiological role beyond immunity.

    Evidence siRNA knockdown with lentiviral rescue, DPP8/9 inhibitor, UUO mouse model, and Smad/Akt immunoblots in HK-2 and mesangial cells

    PMID:33932609 PMID:38458339

    Open questions at the time

    • Whether the fibrotic effect requires catalytic activity not resolved
    • Direct DPP8 substrate in the TGF-β1 pathway unknown
12. 2026 Low

    Positioning DPP8 downstream of MST1 in microglial NLRP1/caspase-1/GSDMD pyroptosis extended the checkpoint to a neuroinflammatory disease context.

    Evidence MST1 knockdown in 5xFAD mice with downstream immunoblots and behavioral assays

    PMID:41689046

    Open questions at the time

    • DPP8 placement is indirect with no direct DPP8 manipulation
    • How MST1 controls DPP8 expression undefined
    • Single study

• It remains unknown which endogenous substrate(s) processed by DPP8/9 generate the signal sensed by NLRP1/CARD8, and how catalytic versus extraenzymatic activities are partitioned across immunity, fibrosis, and migration.
• No direct biochemical link between identified substrates (AK2/EIF2A) and inflammasome activation
• Extraenzymatic partners undefined
• No experimental high-resolution structure of human DPP8