Arup K. Chakraborty

T cells can respond to even a single molecular binding event of antigen to a TCR. A key step in the TCR signaling pathway that definitively exhibits this single-molecule response is the initiation of calcium influx by activation of PLCγ1 in the LAT protein condensate. Here, we describe detailed kinetic measurements examining how protein condensation of LAT regulates activation of PLCγ1 using a reconstituted membrane system. The results reveal that membrane recruitment of PLCγ1 is tightly controlled by the LAT phosphorylation state, with no measurable independent recruitment to PIP ₂ or PIP ₃ lipids via the PLCγ1 PH domains. We further observe PLCγ1 is rapidly activated by membrane-associated kinase upon recruitment, irrespective of the LAT condensation state. These studies also reveal a crosstalk mechanism in which the TEC family kinases responsible for PLCγ1 activation also phosphorylate LAT. This interaction establishes a positive feedback loop in LAT phosphorylation, mediated through LAT condensation, which drives a bimodal LAT phosphorylation response to TCR activation. Kinetic modeling reveals how this LAT phosphorylation response can cooperatively gate PLCγ1 activation from a single TCR. These results suggest the LAT condensate facilitates both signal amplification and noise suppression in PLCγ1 activation through a bimodal switch affecting LAT phosphorylation.

Despite decades of research, an effective HIV vaccine or cure has not yet been developed. An important reason is the virus’s mutability, which allows it to rapidly evade human immune responses. Often, the virus also takes advantage of epistatic pathways to evolve escape mutations through compensatory effects that allow maintaining its fitness. We used a fitness landscape of HIV, which explicitly considers interactions between mutations, to design an immunogen that contains parts of the proteome that are “multidimensionally” conserved. Based on inferred fitness landscapes, these regions are predicted to be under mutational constraints. Our immunogen also incorporates predicted immunogenic regions and offers broad coverage for the Caucasian population. We further investigate the relative contributions of various proteins in our design and compare properties with other immunogens. We hope that our proposed immunogen will serve as a candidate for experimental evaluation in pre-clinical studies in T cell-based vaccine or immunotherapy contexts.

The formation of condensates by the Linker for the Activation of T cells (LAT) is a key signal gating and amplification step in the T-cell receptor signaling pathway. LAT condensation is challenging to study in vivo and is therefore often investigated using reconstitution experiments. While these experiments recapitulate key aspects of LAT condensation, they also exhibit some puzzling features. Here, we describe the mechanisms underlying these observations using two complementary models. First, we employ a Smoluchowski aggregation model to show that the delay time before condensation is observed arises from a low effective binding probability between LAT monomers. Second, we propose a field-theoretic model that reproduces all condensate morphologies observed in experiments, showing that they can arise from common underlying dynamics, modulated by variations in experimental conditions. This result unifies different experimental observations reported previously. While this article addresses open questions regarding the formation of LAT condensates, our results also provide a common framework for understanding the condensation of other multivalent membrane proteins such as EGFR, FGFR2, and nephrin.

The adaptive immune system can mount pathogen-specific responses against a vast world of evolving pathogens (antigens). The humoral immune system is a part of adaptive immunity. It has the remarkable ability to learn about an antigen after exposure to it through a mutation-selection evolutionary process, resulting in more potent responses than those that emerge initially. Recent studies have revealed that upon reexposure to antigens, feedback regulation mediated by past responses steers the evolutionary learning algorithm to generate new responses. This feature enables the immune system to learn to generalize responses to target unseen variants of the original pathogen upon “training” with a limited number of samples. We summarize these new findings and suggest new questions that emerge from them. This is the right time to address these questions by bringing together statistical physics, machine learning, immunology, and evolutionary biology.

The ongoing diversification of influenza virus necessitates annual vaccine updating. The vaccine antigen, the viral spike protein hemagglutinin (HA), tends to elicit strain-specific neutralizing activity, predicting that sequential immunization with the same HA strain will boost antibodies with narrow coverage. However, repeated vaccination with homologous SARS-CoV-2 vaccine eventually elicits neutralizing activity against highly unmatched variants, questioning this immunological premise. We evaluated a longitudinal influenza vaccine cohort, where each year the subjects received the same, novel H1N1 2009 pandemic vaccine strain. Repeated vaccination gradually enhanced receptor-blocking antibodies (HAI) to highly unmatched H1N1 strains within individuals with no initial memory recall against these historical viruses. An in silico model of affinity maturation in germinal centers (GCs) integrated with a model of differentiation and expansion of memory cells outside GCs during a recall response provides insight into the potential mechanisms underlying these results and shows how repeated exposure to the same immunogen can broaden the antibody response against diversified targets.

The formation of condensates by the Linker for the Activation of T-cells (LAT) is a key signal gating and amplification step in the T-cell receptor signaling pathway. LAT condensation is challenging to study in-vivo and is therefore often investigated using reconstitution experiments. While these experiments recapitulate key aspects of LAT condensation, they also exhibit some puzzling features. Here, we describe the mechanisms underlying these observations using two complementary models. First, we employ a Smoluchowski aggregation model to show that the delay time before condensation is observed arises from a low effective binding probability between LAT monomers. Second, we propose a field-theoretic model that reproduces all condensate morphologies observed in experiments, showing that they can arise from common underlying dynamics modulated by variations in experimental conditions. This result unifies different experimental observations reported previously. While this article addresses open questions regarding the formation of LAT condensates, our results also provide a common framework for understanding condensation of other multivalent membrane proteins such as EGFR, FGFR2, and nephrin.

SUMMARY: Antiretroviral therapy (ART) inhibits Human Immunodeficiency Virus (HIV) replication to maintain undetectable viral loads in people living with HIV, but does not result in a cure. Due to the significant challenges of lifelong ART for many, there is strong interest in therapeutic strategies that result in cure. Recent clinical trials have shown that administration of broadly neutralizing antibodies (bnAbs) when there is some viremia can lead to ART-free viral control in some people; however, the underlying mechanisms are unclear. Our computational modeling shows that bnAbs administered in the presence of some viremia promote the evolution of autologous antibodies (aAbs) that target diverse epitopes of HIV spike proteins. This “net” of polyclonal aAbs could confer control since evasion of this response would require developing mutations in multiple epitopes. Our results provide a common mechanistic framework underlying recent clinical observations upon bnAb/ART therapy, and they should also motivate and inform new trials.