Akanksha Chaturvedi

The B Cell Receptor Governs the Subcellular Location of Toll-like Receptor 9 Leading to Hyperresponses to DNA-Containing Antigens

Synergistic engagement of the B cell receptor (BCR) and Toll-like receptor 9 (TLR9) in response to DNA-containing antigens underlies the production of many autoantibodies in systemic autoimmune diseases. However, the molecular basis of this synergistic engagement is not known. Given that these receptors are spatially segregated, with the BCR on the cell surface and TLR9 in endocytic vesicles, achieving synergy must involve unique mechanisms. We show that upon antigen binding, the BCR initiates signaling at the plasma membrane and continues to signal to activate MAP kinases as it traffics to autophagosome-like compartments. The internalized BCR signals through a phospholipase-D-dependent pathway to recruit TLR9-containing endosomes to the autophagosome via the microtubular network. The recruitment of TLR9 to the autophagosomes was necessary for hyperactivation of MAP kinases. This unique mechanism for BCR-induced TLR9 recruitment resulting in B cells hyperresponses may provide new targets for therapeutics for autoimmune diseases.

How location governs Toll like receptor signaling

Toll‐like receptors (TLRs) are a family of innate immune system receptors responsible for recognizing conserved pathogen‐associated molecular patterns (PAMPs). PAMP binding to TLRs initiates intracellular signaling pathways that lead to the upregulation of a variety of costimulatory molecules and the synthesis and secretion of various cytokines and interferons by cells of the innate immune system. TLR‐induced innate immune responses are a prerequisite for the generation of most adaptive immune responses, and in the case of B cells, TLRs directly regulate signaling from the antigen‐specific B‐cell receptor. The outcome of TLR signaling is determined, in part, by the cells in which they are expressed and by the selective use of signaling adaptors. Recent studies suggest that, in addition, both the ligand recognition by TLRs and the functional outcome of ligand binding are governed by the subcellular location of the TLRs and their signaling adaptors. In this review we describe what is known about the intracellular trafficking and compartmentalization of TLRs in innate systems dendritic cells and macrophages and in adaptive systems B cells, highlighting how location regulates TLR function.

Endocytosed BCRs sequentially regulate MAPK and Akt signaling pathways from intracellular compartments.

Binding of antigen to the B cell antigen receptor (BCR) triggers both BCR signaling and endocytosis. How endocytosis regulates BCR signaling remains unknown. Here we report that BCR signaling was not extinguished by endocytosis of BCRs; instead, BCR signaling initiated at the plasma membrane continued as the BCR trafficked intracellularly with the sequential phosphorylation of kinases. Blocking the endocytosis of BCRs resulted in the recruitment of both proximal and downstream kinases to the plasma membrane, where mitogen-activated protein kinases (MAPKs) were hyperphosphorylated and the kinase Akt and its downstream target Foxo were hypophosphorylated, which led to the dysregulation of gene transcription controlled by these pathways. Thus, the cellular location of the BCR serves to compartmentalize kinase activation to regulate the outcome of signaling.

Two-photon fluorescence properties of curcumin as a biocompatible marker for confocal imaging

Two-photon (TP) fluorescence properties of an antioxidant and anti-tumor molecule, curcumin, were investigated. The two-photon absorption (TPA) action cross-section was measured in organic solvents and found to be 6 GM in tetrahydrofuran and 2 GM in dimethyl sulfoxide. The measured TPA cross-section is comparable to that of rhodamine 6G. One-photon and TP confocal microscopy has demonstrated that curcumin is internalized in cells and can be used for imaging applications. Our investigation indicates that curcumin is a viable biocompatible TP fluorescent marker.Two-photon (TP) fluorescence properties of an antioxidant and anti-tumor molecule, curcumin, were investigated. The two-photon absorption (TPA) action cross-section was measured in organic solvents and found to be 6 GM in tetrahydrofuran and 2 GM in dimethyl sulfoxide. The measured TPA cross-section is comparable to that of rhodamine 6G. One-photon and TP confocal microscopy has demonstrated that curcumin is internalized in cells and can be used for imaging applications. Our investigation indicates that curcumin is a viable biocompatible TP fluorescent marker.

B Cells Produce Type 1 IFNs in Response to the TLR9 Agonist CpG-A Conjugated to Cationic Lipids

B cells express the innate receptor, TLR9, which signals in response to unmethylated CpG sequences in microbial DNA. Of the two major classes of CpG-containing oligonucleotides, CpG-A appears restricted to inducing type 1 IFN in innate immune cells and CpG-B to activating B cells to proliferate and produce Abs and inflammatory cytokines. Although CpGs are candidates for adjuvants to boost innate and adaptive immunity, our understanding of the effect of CpG-A and CpG-B on B cell responses is incomplete. In this study we show that both CpG-B and CpG-A activated B cells in vitro to proliferate, secrete Abs and IL-6, and that neither CpG-B nor CpG-A alone induced type 1 IFN production. However, when incorporated into the cationic lipid, DOTAP, CpG-A, but not CpG-B, induced a type 1 IFN response in B cells in vitro and in vivo. We provide evidence that differences in the function of CpG-A and CpG-B may be related to their intracellular trafficking in B cells. These findings fill an important gap in our understanding of the B cell response to CpGs, with implications for the use of CpG-A and CpG-B as immunomodulators.

Corrigendum: Endocytosed BCRs sequentially regulate MAPK and Akt signaling pathways from intracellular compartments

Corrigendum: Endocytosed BCRs sequentially regulate MAPK and Akt signaling pathways from intracellular compartments