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Tumors Exploit Dedicated Intracellular Vesicles to Program T cell Responses

Edward W. Roberts, Megan K. Ruhland, En Cai, Adriana M. Mujal, Kyle Marchuk, Casey Beppler, David Nam, Nina K. Serwas, Mikhail Binnewies, Matthew F. Krummel

Preprint posted on July 04, 2019 https://www.biorxiv.org/content/10.1101/691873v1.abstract

Dendritic cells bring enough to share: cell-cell transfer of tumor antigens in lymph nodes.

Selected by Tim Fessenden

Categories: cancer biology, immunology

Context

Efficient immune responses to diseased tissues require that the sentinels of the adaptive immune system constantly sample their target tissues. These sentinels – most importantly dendritic cells – transport material they encounter to lymph nodes, where they can activate T cells against specific pathogens or tumor cells. Immunologists have long had the tools to analyze the final state of this process – called cross-presentation – by assessing T cell activation. Similarly, our knowledge of dendritic cell subsets and their corresponding behaviors has sharpened over the past few years. What remains somewhat murky is the set of steps between these two: how do dendritic cells ensure that proteins from the diseased tissue are transported and efficiently cross-presented to T cells? A diverse population of dendritic cells manages this, but the precise mechanisms connecting each step in this complex cascade are unclear. Here, Max Krummel’s lab set out to examine how dendritic cells traffic antigens from tumors to lymph nodes.

 

Data

In order to follow cell debris derived from tumor cells as it is ingested and transported by dendritic cells, the authors make use of a fluorescent protein, ZsGreen, which remains stable at low pH. From mice that bear ZsGreen-expressing tumor cells, the authors use flow cytometry to analyze all immune cells that retain fluorescence from ZsGreen. With this assay they find that all phagocytic cells ingest at least some tumor cell debris. As dendritic cells specialize in transporting cell debris from tumors to lymph nodes, the authors analyze lymph nodes and indeed find cells containing intracellular puncta of ZsGreen. However, when they compare dendritic cells from the tumor vs from the lymph node, they note that lymph nodes exhibit relatively more ZsGreen+ cells that are however of lower fluorescence intensity. This observation motivates the foundational hypothesis of this work: that a few dendritic cells ingest a lot of cell debris in the tumor, which they then distribute among many dendritic cells within the lymph node. Their operating metaphor is the game of mancala in which game pieces are distributed equally among a set of holes, recalled dimly by this reader from his childhood.

To test this hypothesis, the authors isolate ZsGreen+ dendritic cells from a tumor-bearing mouse and coculture these ex vivo with dendritic cells from a naïve mouse. This assay clearly demonstrates that tumor-derived material is spread to the naïve dendritic cells, in a process that requires direct contact between cells. Live imaging of dendritic cells confirms that intracellular vesicles containing ZsGreen are transferred between apposed dendritic cells in vivo and ex vivo. Most interestingly, they find this sharing is most efficient from stimulatory dendritic cells previously associated with a robust anti-tumor immune response. This observation associates antigen sharing with a dendritic cell subset whose activities are required for a productive anti-tumor immune response.

What about the kinetics of this transfer of tumor cell material? The authors use a tumor cell line in which they can turn on ZsGreen expression using tamoxifen. This enables a pulse/chase experimental setup, and demonstrates that a migratory subset of dendritic cells takes up the tumor cell material first, and then transfers this material to nonmigratory dendritic cells residing in the lymph node. Finally the authors correlate this cell-cell transfer with cross-presentation and activation of T cells in the lymph node or ex vivo.

 

Implications

The authors report observations that redefine the capabilities of dendritic cells as transporters and presenters of antigens derived from tumors. Some hints at a unique promiscuity of dendritic cells exist already, for example from evidence that they can exchange membrane with nonimmune cells and thereby take up exogenous MHC-I receptors already bearing antigen[1]. However, the present work marks an important and provocative conceptual advance for the logic of antigen cross-presentation, and opens new avenues to investigate dendritic cells as not only capturing and transporting but also disseminating antigens from target tissues.

This work does not furnish details on the nature of the cell-cell contacts through which antigen-bearing vesicles might pass, nor on the trafficking of vesicles themselves. Readers must await future studies to examine these contacts and the frequency of such antigen sharing. Overall the present data suggest that as they survey and integrate cell debris to present to T cells, dendritic cells do not respect boundaries between cells but share their contents liberally to ensure an optimal immune response.

1. Wakim, L.M. and M.J. Bevan, Cross-dressed dendritic cells drive memory CD8+ T-cell activation after viral infection. Nature, 2011. 471(7340): p. 629-632.

Tags: b16, dendritic cell, immunotherapy, microscopy

Posted on: 7th October 2019

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  • Author's response

    Megan K. Ruhland shared

    Do you think that antigen sharing via this or other mechanisms would also mediate immune responses to intracellular viruses? Or is there a particular feature of tumors that renders them especially suited to immune activation via this mechanism?

    It is entirely possible that viral antigen could be shunted into the antigen trafficking pathway we describe. We have ongoing studies that look further into other sources of antigen including steady-state self-antigen and commensal and pathogenic bacteria-derived antigens. In these cases we find interesting similarities and important differences when comparing with tumor-derived antigen. These findings likely will be informative beyond anti-tumor immune responses and include how the immune system senses and transmits information about various types of antigen in order to mount the appropriate response.

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