The helminth TGF-β mimic TGM4 is a modular ligand that binds CD44, CD49d and TGF-β receptors to preferentially target myeloid cells

Abstract

The murine helminth parasiteHeligmosomoides polygyrusexpresses a family of modular proteins which, replicating the functional activity of the immunomodulatory cytokine TGF-β, have been named TGM (TGF-β Μimic). Multiple domains bind to different receptors, including TGF-β receptors TβRI (ALK5) and TβRII through domains 1-3, and prototypic family member TGM1 binds the cell surface co-receptor CD44 through domains 4-5. This allows TGM1 to induce T lymphocyte Foxp3 expression, characteristic of regulatory (Treg) cells, and to activate a range of TGF-β-responsive cell types. In contrast, a related protein, TGM4, targets a much more restricted cell repertoire, primarily acting on myeloid cells, with less potent effects on T cells and lacking activity on other TGF-β-responsive cell types. TGM4 binds avidly to myeloid cells by flow cytometry, and can outcompete TGM1 for cell binding. Analysis of receptor binding in comparison to TGM1 reveals a 10-fold higher affinity than TGM1 for TGFβR-I (TβRI), but a 100-fold lower affinity for TβRII through Domain 3. Consequently, TGM4 is more dependent on co-receptor binding; in addition to CD44, TGM4 also engages CD49d (Itga4) through Domains 1-3, as well as CD206 and Neuropilin-1 through Domains 4 and 5. TGM4 was found to effectively modulate macrophage populations, inhibiting lipopolysaccharide-driven inflammatory cytokine production and boosting interleukin (IL)-4-stimulated responses such as Arginase-1in vitroandin vivo. These results reveal that the modular nature of TGMs has allowed the fine tuning of the binding affinities of the TβR- and co-receptor binding domains to establish cell specificity for TGF-β signalling in a manner that cannot be attained by the mammalian cytokine.

TGM6, a helminth secretory product, mimics TGF-β binding to TβRII to antagonize TGF-β signaling in fibroblasts

“…S12 ) [29]. TGM4 which binds TβRII more weakly than TGM1 [32] and does not signal in MFB-F11 fibroblasts [29], also lacks a basic residue homologous to TGM6 Arg 82 , but also absent is the Pro 94 -Arg 95 dipeptide of TGM6 or the Lys 254 -Asn 255 dipeptide of TGM1. TGM7, which has not been found to have any TGF-β signaling activity in MFB-F11 fibroblasts [29], or TGM8, which has not functionally characterized, retain an arginine homologous to TGM6 Arg 82 , but have either a His-Asn or Leu-Asn dipeptide in place of the KN dipeptide of TGM1 or the PR dipeptide of TGM6.…”

“…TGM1, for example, was shown to bind the co-receptor CD44 through domains 4 and 5 [31]. This increases the signaling potency and biases TGM1 activity to cells which have abundant CD44, such as T lymphocytes and myeloid cells [31, 32]. TGM4, which shares the same five-domain structure as TGMs 1, 2, and 3, was recently shown to bind TβRI 10-fold more strongly than TGM1, but TβRII 100-fold less avidly than TGM1 [32].…”

“…This increases the signaling potency and biases TGM1 activity to cells which have abundant CD44, such as T lymphocytes and myeloid cells [31, 32]. TGM4, which shares the same five-domain structure as TGMs 1, 2, and 3, was recently shown to bind TβRI 10-fold more strongly than TGM1, but TβRII 100-fold less avidly than TGM1 [32]. This combination of receptor binding affinities, together with binding of CD44 and several other co-receptors, including CD206, Itga4, and neuropilin-1, biases TGM4 activity toward different subclasses of myeloid cells that express these co-receptors.…”

“…The targeting of the TGF-β pathway by the helminth H. polygyrus therefore has potentially adverse consequences for the host. To overcome potentially deleterious off-target effects, and thus limit damage to the host, the parasite has adapted the multidomain CCP scaffold proteins, not only to bind TβRI and TβRII to activate the TGF-β pathway, but also to bind co-receptors that precisely target them to relevant immune cell subsets [6, 31, 32].…”

“…1A ) [6]. While TGM6 D3, D4, and D5 share 66%, 42%, and 32% identity with TGM1’s D3, D4, and D5, it does not signal in TGF-β functional assays, unlike TGM1, TGM2, and TGM3 which signal in both fibroblasts and T cells, or TGM4 which signals in T cells and myeloid cells [29, 32]. In this work, we show that TGM6 binds TβRII and that it does so through D3, but it does not bind TβRI or other type I and type II receptors of the TGF-β family.…”