Biology subjects

Mallis, R. J.

Publications and source records attributed to Mallis, R. J..

6 recordsLinked to original sources

bioRxiv2025

Cancers modulate p53 truncal neoantigen display to evade T cell detection

Summary paragraphTP53 mutations are early truncal events across cancers1,2. These are perceived to encode tumour-specific neoantigens representing prime cytotoxic T lymphocyte (CTL) targets3,4. However, studies systematically examining the physical cell surface display of p53 peptides bound to major histocompatibility complex molecules (pMHC), their relative antigenicity, and resultant immunogenicity have yet to be conducted. Here, we develop an epitope discovery platform using p53-reconstituted lung cancer cells as well as various tumour cells as pMHC sources. Combining data-independent acquisition mass spectrometry (MS), nanoscale chromatography, and peptide detection based on probabilistic measure and three-dimensional ion visualization techniques allows attomole sensitivity identification of pMHCs. This approach excluded [~]97% of algorithm-based virtual p53 immunopeptidomes, highlighting that only a few p53 pMHCs can be presented by common human MHC (human leukocyte antigen, [HLA]) alleles. Strikingly, surface expressed neoantigens are restricted to the corresponding set of such limited self-p53 peptide arrays and unaffected by enhancing p53 proteasomal turnover. Further curtailment of MS-validated, high affinity p53 neoepitopes that are structurally deviant from self-pMHC occurs in established tumours due to immune selection against the antigen presenting MHC allele or by a novel mechanism involving p53 neoepitope destruction by endoplasmic reticulum aminopeptidase 1 (ERAP1). In contrast, given the extremely weak MHC affinity and resultant short-lived cell surface pMHC expression, the common p53 neoepitope R175H/HLA-A*02:01 escapes immune selection despite CTL with high quality T-cell receptors. Rigorous tumour-protective immunoediting makes effective truncal neoepitope targeting a challenge, requiring attentive MS analysis and functional vetting to focus protective cytolytic responses.

immunology↗

bioRxiv2025

Atomistic TCR-ligand interactions instruct memory T-cell differentiation

Memory CD8 T cells provide durable protection against recurrent infection and cancer, but how distinct memory fates are specified remains unclear. Here, we define the functional landscape of 242 murine CD8 TCR{beta} clonotypes specific for influenza NP366-374/H-2D ligand (pMHC) by integrating single-cell transcriptomics and TCR sequencing with force-dependent biophysics, structural analysis, and in vivo investigation of memory development. Fate essentially tracks with TCR mechanotransduction: TCM-associated clonotypes preferentially engage pMHC through TCR{beta}, whereas TEM-associated clonotypes preferentially engage through TCR. Clonotypes with balanced, bipolar signaling generate both memory subsets, optimize expansion, and display broad heterosubtypic crossreactivity but with exhaustion susceptibility. In contrast, diverse TCM clonotypes are largely clonal singlets with considerable mutant-epitope recognition in aggregate and stemness preservation, revealing complementary strategies for immunodefense coverage. Atomistically, subtle pMHC contact differences and load transmission through the TCR holoreceptor appear to tune phosphorylation, memory differentiation, and functional durability, features applicable to adoptive T-cell immunotherapies.

immunology↗

bioRxiv2024

Load-based divergence in the dynamic allostery of two TCRs recognizing the same pMHC

Increasing evidence suggests that mechanical load on the {beta} T cell receptor (TCR) is crucial for recognizing the antigenic peptide-loaded major histocompatibility complex (pMHC) molecule. Our recent all-atom molecular dynamics (MD) simulations revealed that the inter-domain motion of the TCR is responsible for the load-induced catch bond behavior of the TCR-pMHC complex and peptide discrimination. To further examine the generality of the mechanism, we perform all-atom MD simulations of the B7 TCR under different conditions for comparison with our previous simulations of the A6 TCR. The two TCRs recognize the same pMHC and have similar interfaces with pMHC in crystal structures. We find that the B7 TCR-pMHC interface stabilizes under [~]15-pN load using a conserved dynamic allostery mechanism that involves the asymmetric motion of the TCR chassis. However, despite forming comparable contacts with pMHC as A6 in the crystal structure, B7 has fewer high-occupancy contacts with pMHC and exhibits higher mechanical compliance during the simulation. These results indicate that the dynamic allostery common to the TCR{beta} chassis can amplify slight differences in interfacial contacts into distinctive mechanical responses and nuanced biological outcomes.

biophysics↗

bioRxiv2023

Parsing digital or analogue TCR performance through piconewton forces

{beta} T-cell receptors (TCRs) recognize aberrant peptides bound to major histocompatibility complex molecules (pMHCs) on unhealthy cells, amplifying specificity and sensitivity through physical load placed on the TCR-pMHC bond during immunosurveillance. To understand this mechanobiology, TCRs stimulated by abundantly and sparsely arrayed epitopes (NP366-374 /Db and PA224-233/Db, respectively) following in vivo influenza A virus infection were studied with optical tweezers. While certain NP repertoire CD8 T lymphocytes require many ligands for activation, others are digital, needing just few. Conversely, all PA TCRs perform digitally, exhibiting pronounced bond lifetime increases through sustained, energizing volleys of structural transitioning. Optimal digital performance is superior in vivo, correlating with ERK phosphorylation, CD3 loss, and activation marker upregulation in vitro. Given neoantigen array paucity, digital TCRs are likely critical for immunotherapies. One Sentence SummaryQuality of ligand recognition in a T-cell repertoire is revealed through application of physical load on clonal T-cell receptor (TCR)-pMHC bonds

immunology↗

bioRxiv2023

Asymmetric framework motion of TCR alpha beta controls load-dependent peptide discrimination

Mechanical force is critical for the interaction between an {beta}T cell receptor (TCR) and a peptide-bound major histocompatibility complex (pMHC) molecule to initiate productive T-cell activation. However, the underlying mechanism remains unclear. We use all-atom molecular dynamics simulations to examine the A6 TCR bound to HLA-A*02:01 presenting agonist or antagonist peptides under different extensions to simulate the effects of applied load on the complex, elucidating their divergent biological responses. We found that TCR and {beta} chains move asymmetrically, which impacts the interface with pMHC, in particular the peptide-sensing CDR3 loops. For the wild-type agonist, the complex stabilizes in a load-dependent manner while antagonists destabilize it. Simulations of the C{beta} FG-loop deletion, which reduces the catch bond response, and simulations with in silico mutant peptides further support the observed behaviors. The present results highlight the combined role of interdomain motion, fluctuating forces, and interfacial contacts in determining the mechanical response and fine peptide discrimination by a TCR, thereby resolving the conundrum of nearly identical crystal structures of TCR{beta}-pMHC agonist and antagonist complexes.

biophysics↗

bioRxiv2022

Pre-T cell receptor Self-MHC Sampling Restricts Thymocyte Dedifferentiation

Summary paragraphProgramming T lymphocytes to distinguish self from non-self is a vital, multi-step process arising in the thymus1-4. Signalling through the pre-T cell receptor (preTCR), a CD3-associated heterodimer comprising an invariant pT chain and a clone-specific {beta} chain, constitutes a critical early checkpoint in thymocyte development within the {beta} T-cell lineage5, 6. Recent work demonstrates that preTCRs arrayed on double negative (DN) thymocytes, like {beta} TCRs appearing on double positive (DP) thymocytes, ligate peptides bound to MHC molecules (pMHC) on thymic stroma but via a different molecular docking strategy7-10. Here we show the consequences of those distinctive interactions for thymocyte progression, using synchronized fetal thymic progenitor cultures differing in the presence or absence of pMHC on support stroma, determining single cell transcriptomes at key thymocyte developmental transitions. Although MHC negative stroma fosters {beta} T lymphocyte differentiation, the absence of pMHC-preTCR interplay leads to deviant thymocyte transcriptional programming associated with de-differentiation. Highly proliferative DN and DP subsets with antecedent characteristics of T cell lymphoblastic and myeloid malignancies emerge. Thus, at least in vitro, beyond fostering {beta} chain repertoire broadening for subsequent {beta} TCR utilization, preTCR-pMHC interaction limits cellular plasticity to facilitate normal thymocyte differentiation and proliferation that, if absent, introduces significant developmental vulnerabilities.

immunology↗