bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2022.05.09.490905

Computational modeling of TGF-β2:TβRI:TβRII receptor complex assembly as mediated by the TGF-β co-receptor betaglycan

Ingle, M.·Madamanchi, A.·Hinck, A. P.·Umulis, D. M.
View original source ↗Export metadata ↓

Abstract

Transforming growth factor-{beta}1, -{beta}2, and -{beta}3 (TGF-{beta}1, -{beta}2, and -{beta}3) are secreted signaling ligands that play essential roles in tissue development, tissue maintenance, immune response, and wound healing. TGF-{beta} homodimers signal by assembling a heterotetrameric complex comprised of two type I receptor (T{beta}RI):type II receptor (T{beta}RII) pairs. TGF-{beta}1 and TGF-{beta}3 signal with high potency due to their high affinity for T{beta}RII, which engenders high affinity binding of T{beta}RI through a composite TGF-{beta}:T{beta}RII binding interface. However, TGF-{beta}2 binds T{beta}RII 200-500 more weakly than T{beta}RII and signals with lower potency compared to TGF-{beta}1 and -{beta}3. Remarkably, potency of TGF-{beta}2 is increased to that of TGF-{beta}1 and -{beta}3 in the presence of an additional membrane-bound co-receptor, known as betaglycan (BG), even though betaglycan does not directly participate in the signaling mechanism and is displaced as the signaling receptors, T{beta}RI and T{beta}RII, bind. To determine the role of betaglycan in the potentiation of TGF-{beta}2 signaling, we developed deterministic computational models with different modes of betaglycan binding and varying cooperativity between receptor subtypes. The models, which were developed using published kinetic rate constants for known quantities and optimization to determine unknown quantities, identified conditions for selective enhancement of TGF-{beta}2 signaling. The models provide support for additional receptor binding cooperativity that has been hypothesized, but not evaluated in the literature. The models further showed that betaglycan binding to TGF-{beta}2 ligand through two domains provides an effective mechanism for transfer to the signaling receptors that has been tuned to efficiently promote assembly of the TGF-{beta}2(T{beta}RII)2(T{beta}RI)2 signaling complex.

Source connections

Explore related subjects

biophysicsPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Ingle, M., Madamanchi, A., Hinck, A. P., Umulis, D. M.. 2022-05-09. Computational modeling of TGF-β2:TβRI:TβRII receptor complex assembly as mediated by the TGF-β co-receptor betaglycan. https://doi.org/10.1101/2022.05.09.490905

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

A Deformulating Multi-Attribute SEC Method for Simultaneous Quantification of Total RNA, Conjugated Antibody, and Free Antibody in Targeted Lipid Nanoparticles

Targeted lipid nanoparticles (tLNPs) hold promise for improving cell- and tissue-selective delivery of therapeutic payloads but introduce analytical challenges associated with characterizing both encapsulated cargo and surface-associated targeting ligands within the same formulation. Conventional nucleic acid and protein assays can be affected by nanoparticle scattering, matrix interference, or a limited ability to distinguish conjugated antibody from free antibody. Here, we report an online deformulating size-exclusion high-performance liquid chromatography (SE-HPLC) method that enables simultaneous quantification of total RNA, conjugated antibody, and free antibody from 5 L of tLNP sample. Following injection, exposure to a mobile phase containing sodium dodecyl sulfate and isopropanol disrupts the nanoparticles, enabling chromatographic separation of RNA, conjugated antibody, and free (unconjugated) antibody while minimizing sample handling prior to analysis. RNA was quantified by UV260 detection, whereas antibody was quantified by UV280 or intrinsic fluorescence detection to maximize sensitivity. The method demonstrated linear response over the evaluated concentration ranges, spike recoveries of 88-122%, and precision below 5% relative standard deviation. Application to purified tLNP formulations demonstrated simultaneous assessment of total RNA and conjugated antibody content, while confirming the absence of detectable free antibody following purification. Together, these results establish online deformulating SE-HPLC as a practical multi-attribute analytical method for characterization of tLNPs, enabling concurrent assessment of total RNA concentration, conjugated antibody concentration, and free antibody concentration using standard HPLC instrumentation.

biophysics↗
bioRxiv2026

Nanomechanical phenotyping uncovers a non-canonical smooth muscle-like hypercontractility program in lung squamous cell carcinoma myofibroblasts

Myofibroblast-like cancer-associated fibroblasts (myCAFs) drive the stiff, tumor-promoting microenvironment, yet their mechanical phenotype in non-small cell lung cancer (NSCLC) remains undefined. myCAFs from major NSCLC subtypes (lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC)) were examined combining atomic force microscopy and traction force microscopy with molecular and signaling analyses. LUAD-myCAFs displayed canonical myofibroblast features, including elevated stiffness, traction forces, ROCK-MYPT1 signaling, and -SMA-rich stress fibers. Unexpectedly, LUSC-myCAFs amplified these nanomechanical features despite reduced ROCK-MYPT1 signaling and comparable -SMA levels. Instead, they exhibited a smooth muscle cell (SMC)-like contractile program characterized by enhanced SM-MHC(MYH11) expression colocalized with -SMA, ERK1/2-RLC hyperactivation, and DNA methylation reprogramming including SMC-contractility regulators. LUSC-myCAF hypercontractility was linked with reduced proliferative capacity and smaller intratumoral vessel lumens, consistent with the low CAF density and enhanced hypoxia patterns reported in LUSC. SMC markers were enriched and associated with adverse outcomes selectively in LUSC, highlighting the relevance of SMC-like myCAFs in LUSC progression. These findings unveil subtype-specific myCAF programs in NSCLC, and identify -SMA/SM-MHC(MYH11) myCAFs as key determinants of tumor mechanics, providing a rationale for the survival benefits of antihypertensive drugs targeting SMC hypercontractility. Moreover, they demonstrate that nanomechanical phenotyping can uncover translationally relevant myCAF phenotypes invisible to molecular profiling.

biophysics↗
bioRxiv2026

Engineering Nanobody Conformational Switches

Conformational signaling is widely used in biology, but remains difficult to engineer into proteins. Here, we explore ligand-induced folding of nanobodies as a general strategy to generate conformational receptors for a broad range of target ligands. More specifically, we have generated partially unfolded nanobody variants that fold exclusively upon binding to their target ligand. To do so, we have introduced destabilizing mutations in the framework region, carefully tuning their stability while preserving molecular recognition. Given the high conservation of the framework across different nanobodies, this approach is broadly applicable to many other target ligands, which has enabled us to engineer conformational switches for three different important biomarkers and drugs: the pregnancy hormone chorionic gonadotropin, the cytokine tumor necrosis factor alpha, and the chemotherapy and immunosuppressant drug methotrexate. Another useful property of conformational signaling is that it can directly gate signal transduction, which we have leveraged to build an optical sensor for the quantitative detection of the pregnancy hormone in urine. Finally, conformational signaling introduces new mechanisms for the allosteric modulation of binding, which we have explored to control the affinity and dynamic range of the sensor. Our work thus establishes a rational strategy for engineering conformational signaling receptors with customizable properties and generalizable for many different target ligands.

biophysics↗