Loss of LRRC33-dependent TGFβ1 activation enhances antitumor immunity and checkpoint blockade therapy
Jiang, A., Qin, Y. & Springer, T.A. Loss of LRRC33-dependent TGFβ1 activation enhances antitumor immunity and checkpoint blockade therapy. Cancer Immunol Res 10, 4, 453-467 (2022).Abstract
TGFβ has multiple roles and gene products (TGFβ1, -β2, and -β3), which make global targeting of TGFβ undesirable. Expression of TGFβ requires association with milieu molecules, which localize TGFβ to the surface of specific cells or extracellular matrices. Here, we found that LRRC33 was specifically associated with TGFβ1, not TGFβ2 and TGFβ3, and was required for surface display and activation of TGFβ1 on tumor-infiltrating myeloid cells. Loss of LRRC33-dependent TGFβ1 activation slowed tumor growth and metastasis by enhancing innate and adaptive antitumor immunity in multiple mouse syngeneic tumor models. LRRC33 loss resulted in a more immunogenic microenvironment, with decreased myeloid-derived suppressor cells, more active CD8+ T and NK cells, and more skewing toward tumor-suppressive M1 macrophages. LRRC33 loss and PD-1 blockade synergized in controlling B16.F10 tumor growth. Our results demonstrate the importance of LRRC33 in tumor biology and highlight the therapeutic potential of dual blockade of the LRRC33/TGFβ1 axis and PD-1/PD-L1 in cancer immunotherapy.
Protection of the Prodomain α1-Helix Correlates with Latency in the Transforming Growth Factor-β Family
Le, V.Q., et al. Protection of the Prodomain α1-Helix Correlates with Latency in the Transforming Growth Factor-β Family. J Mol Biol 434, 5, 167439 (2022).Abstract
The 33 members of the transforming growth factor beta (TGF-β) family are fundamentally important for organismal development and homeostasis. Family members are synthesized and secreted as pro-complexes of non-covalently associated prodomains and growth factors (GF). Pro-complexes from a subset of family members are latent and require activation steps to release the GF for signaling. Why some members are latent while others are non-latent is incompletely understood, particularly because of large family diversity. Here, we have examined representative family members in negative stain electron microscopy (nsEM) and hydrogen deuterium exchange (HDX) to identify features that differentiate latent from non-latent members. nsEM showed three overall pro-complex conformations that differed in prodomain arm domain orientation relative to the bound growth factor. Two cross-armed members, TGF-β1 and TGF-β2, were each latent. However, among V-armed members, GDF8 was latent whereas ActA was not. All open-armed members, BMP7, BMP9, and BMP10, were non-latent. Family members exhibited remarkably varying HDX patterns, consistent with large prodomain sequence divergence. A strong correlation emerged between latency and protection of the prodomain α1-helix from exchange. Furthermore, latency and protection from exchange correlated structurally with increased α1-helix buried surface area, hydrogen bonds, and cation-pi bonds. Moreover, a specific pattern of conserved basic and hydrophobic residues in the α1-helix and aromatic residues in the interacting fastener were found only in latent members. Thus, this first comparative survey of TGF-β family members reveals not only diversity in conformation and dynamics but also unique features that distinguish latent members.
hao headshot

Yuxin Hao, PhD

Postdoctoral Research Fellow
Boston Children's Hospital, Harvard Medical School
Aiping Jiang

Aiping Jiang

Postdoctoral Research Fellow
Boston Children's Hospital, Harvard Medical School

Bo Zhao, Ph.D.

Assistant Professor, Department of Immunology, Molecular Cancer Research Center, School of Medicine, Sun Yat-sen University, Shenzhen, China
L., L., et al. Carbon nanotube-assisted optical activation of TGF-β signalling by near-infrared light. Nat Nanotechnol. 10, 5, 465-71 (2015).Abstract

Receptor-mediated signal transduction modulates complex cellular behaviours such as cell growth, migration and differentiation. Although photoactivatable proteins have emerged as a powerful tool for controlling molecular interactions and signalling cascades at precise times and spaces using light, many of these light-sensitive proteins are activated by ultraviolent or visible light, which has limited tissue penetration. Here, we report a single-walled carbon nanotube (SWCNT)-assisted approach that enables near-infraredlight-triggered activation of transforming growth factor β (TGF-β) signal transduction, an important signalling pathway in embryonic development and cancer progression. The protein complex of TGF-β and its latency-associated peptide is conjugated onto SWCNTs, where TGF-β is inactive. Upon near-infrared irradiation, TGF-β is released through the photothermal effect of SWCNTs and becomes active. The released TGF-β activates downstream signal transduction in live cells and modulates cellular behaviours. Furthermore, preliminary studies show that the method can be used to mediate TGF-β signalling in living mice.

Dong, X., Hudson, N.E., Lu, C. & Springer, T.A. Structural determinants of integrin β-subunit specificity for latent TGF-β. Nat Struct Mol Biol. 21, 12, 1091-6 (2014).Abstract

Eight integrin α-β heterodimers recognize ligands with an Arg-Gly-Asp (RGD) motif. However, the structural mechanism by which integrins differentiate among extracellular proteins with RGD motifs is not understood. Here, crystal structures, mutations and peptide-affinity measurements show that αVβ6 binds with high affinity to a RGDLXXL/I motif within the prodomains of TGF-β1 and TGF-β3. The LXXL/I motif forms an amphipathic α-helix that binds in a hydrophobic pocket in the β6 subunit. Elucidation of the basis for ligand binding specificity by the integrin β subunit reveals contributions by three different βI-domain loops, which we designatespecificity-determining loops (SDLs) 1, 2 and 3. Variation in a pair of single key residues in SDL1 and SDL3 correlates with the variation of the entire β subunit in integrin evolution, thus suggesting a paradigmatic role in overall β-subunit function.

Viet Le, PhD

Viet Le, PhD

Instructor in Pediatrics, Harvard Medical School
Research Associate, Boston Children's Hospital
Structure of bone morphogenetic protein 9 procomplex
Mi, L.-Z., et al. Structure of bone morphogenetic protein 9 procomplex. Proc Natl Acad Sci USA 112, 3710-5 (2015).Abstract

Bone morphogenetic proteins (BMPs) belong to the TGF-β family, whose 33 members regulate multiple aspects of morphogenesis. TGF-β family members are secreted as procomplexes containing a small growth factor dimer associated with two larger prodomains. As isolated procomplexes, some members are latent, whereas most are active; what determines these differences is unknown. Here, studies on pro-BMP structures and binding to receptors lead to insights into mechanisms that regulate latency in the TGF-β family and into the functions of their highly divergent prodomains. The observed open-armed, nonlatent conformation of pro-BMP9 and pro-BMP7 contrasts with the cross-armed, latent conformation of pro-TGF-β1. Despite markedly different arm orientations in pro-BMP and pro-TGF-β, the arm domain of the prodomain can similarly associate with the growth factor, whereas prodomain elements N- and C-terminal to the arm associate differently with the growth factor and may compete with one another to regulate latency and stepwise displacement by type I and II receptors. Sequence conservation suggests that pro-BMP9 can adopt both cross-armed and open-armed conformations. We propose that interactors in the matrix stabilize a cross-armed pro-BMP conformation and regulate transition between cross-armed, latent and open-armed, nonlatent pro-BMP conformations.