Morath, K. | Search bioRxiv

bioRxiv2025

HIV-1 Reprograms CD4 T Cell Responses by Impairing Antigen-specific Communication with Dendritic Cells

HIV-1 infection causes general dysfunction of adaptive immune cells that persists even under therapy but the underlying mechanisms remain elusive. Antigen-specific interactions of the main target cells of HIV, CD4 T cells, with dendritic cells (DCs) orchestrate global T cell responses and convey help to CD8 T cells. Here we report that HIV-1, by virtue of its pathogenesis factor Nef, impairs activation and transcriptionally reprograms CD4 T cells to dampen Th1 differentiation in response to antigen-specific stimulation by DCs. These alterations also disrupt functional communication to DCs to reduce DC activation and limit Th1 helper cytokine production. Mechanistically, Nef achieves this modulation of antigen-specific CD4 T cell function by reducing T cell surface levels of CD4. These results define modulation of CD4 T cell-DC communication as pathogenic principle by which HIV-1 disrupts adaptive immunity and emphasize the direct role of CD4 in immune cell communication. One Sentence SummaryHIV-1 suppresses Th1 polarization by disrupting the communication between CD4 T cells and Dendritic cells through cell surface CD4 downregulation by the viral pathogenesis factor Nef.

immunology↗

bioRxiv2023

ediTONSIL: Activation-Neutral Gene Editing of CD4 T cells in Human ex Vivo Tonsil Cultures

MotivationCD4 T cells are central players of adaptive immunity that rapidly change activation states in response to exogenous T cell receptor (TCR) stimulation. While molecular processes in activated human CD4 T cells from peripheral blood are well studied, resting CD4 T cells are refractory to gene-editing transduction and transfection methods without prior activation. Knowledge on the molecular biology of truly resting CD4 T cells is therefore lacking. We present here the culture and editing workflow ediTONSIL that allows for gene editing of tissue-resident CD4 T cells without compromising their activation state or immunological function. This will enable molecular mechanistic analyses of this cell type in their native/physiological tissue context. SummaryThe molecular and immunological properties of resting CD4 T cell biology are understudied due to the lack of suitable gene editing methods. Here we describe the ex vivo culture and gene editing methodology ediTONSIL for tissue-resident CD4 T cells from human tonsil tissue. CRISPR/Cas9 RNP nucleofection under optimized culture conditions and cytokine concentrations results in knock out efficacies of over 90%, which are e.g. sufficient to prevent HIV-1 infection when targeting the viral entry co-receptor CXCR4. EdiTONSIL does not impair tonsil CD4 T cell viability, activation state or immunocompetence and does not require exogenous activation. Editing can be performed on multiple cell types in bulk cultures or of CD4 T cells previously isolated from tonsils that can be added back to the non-CD4 T cell fraction post gene editing to reassemble into immunocompetent organotypic lymphoid aggregate structures. This highly efficient and versatile workflow for gene editing of tonsillar CD4 T cells enables the dissection of molecular mechanisms in ex vivo cultures of human lymphoid tissue and can be adapted to other tonsil-resident cell types. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/554089v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@1ece503org.highwire.dtl.DTLVardef@1af2d0corg.highwire.dtl.DTLVardef@1b67f14org.highwire.dtl.DTLVardef@510d63_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗