Pathni, A. | Search bioRxiv

bioRxiv2025

Nuclear morphology and chromatin organization modulate T cell cytoskeletal remodeling and immune synapse formation

T cell activation is characterized by rapid reorganization of the actin cytoskeleton and cell spreading on the antigen presenting cell. The T cell nucleus occupies a large fraction of the cell volume, and its mechanical properties are likely to act as a key determinant of activation. However, the contribution of nuclear mechanics to T cell spreading and activation is not well understood. Mechanical rigidity of lymphocyte nuclei is conferred by chromatin compaction and dense packing of heterochromatin. We find that nuclear deformation and increased chromatin compaction accompany T cell spreading, in T cells. Reducing chromatin compaction leads to increased cell spread area and nuclear deformation, while diminishing accumulation and peripheral enrichment of F-actin at the immune synapse. In contrast, enhanced chromatin compaction reduced spread area and nuclear deformation, which was accompanied by increased peripheral F-actin organization at the immune synapse. These findings suggest a reciprocal interaction between chromatin compaction and actin cytoskeletal organization. We identified SUN proteins and myosin as critical elements through which chromatin compaction orchestrates actin morphology and cell shape, facilitating T cell adaptation to antigen-presenting surfaces of varying stiffness. These results emphasize the crucial role of chromatin compaction in T cell activation, underlining the mechanical relationship between the nucleus and the cytoskeleton during immune responses, and suggest new avenues for understanding T cell mechano-responsiveness.

biophysics↗

bioRxiv2024

Preventing trogocytosis by cathepsin B inhibition augments CAR T cell function

Chimeric antigen receptor (CAR) T cell therapy has shown remarkable efficacy in cancer treatment. Still, most patients receiving CAR T cells relapse within 5 years of treatment. CAR-mediated trogocytosis (CMT) is a potential tumor escape mechanism in which cell surface proteins transfer from tumor cells to CAR T cells. CMT results in the emergence of antigen-negative tumor cells, which can evade future CAR detection, and antigen-positive CAR T cells, which has been suggested to cause CAR T cell fratricide and exhaustion. Whether CMT indeed causes CAR T cell dysfunction and the molecular mechanisms conferring CMT remain unknown. Using a selective degrader of trogocytosed antigen in CAR T cells, we show that the presence of trogocytosed antigen on the CAR T cell surface directly causes CAR T cell fratricide and exhaustion. By performing a small molecule screening using a custom high throughput CMT-screening assay, we found that the cysteine protease cathepsin B is essential for CMT and that inhibition of cathepsin B is sufficient to prevent CAR T cell fratricide and exhaustion, leading to improved long-term CAR T cell persistence and anti-tumor activity. Our data demonstrate that it is feasible to separate CMT from cytotoxic activity, that CAR T cell persistence, a key factor associated with clinical CAR T cell efficacy, is directly linked to cathepsin B activity in CAR T cells, and that it is possible to improve CAR T cell function through selective inhibition of CMT. One sentence summary: CAR-mediated trogocytosis is mediated by the cysteine protease cathepsin B and directly causes CAR T cell exhaustion and fratricide. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=116 SRC="FIGDIR/small/598379v3_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@5c058dorg.highwire.dtl.DTLVardef@1707fa7org.highwire.dtl.DTLVardef@ccdff1org.highwire.dtl.DTLVardef@82800b_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗