HkRP3 Is a Microtubule-Binding Protein Regulating Lytic Granule Clustering and NK Cell Killing

Secretion of misfolded tau, a microtubule-binding protein enriched in nerve cells, is linked to the progression of tau pathology. However, the molecular mechanisms underlying tau secretion are poorly understood. Recent work by Lee et al. [Biochemical J. (2021) 478: 1471–1484] demonstrated that the transmembrane domains of syntaxin6 and syntaxin8 could be exploited for tau release, setting a stage for testing a novel hypothesis that has profound implications in tauopathies (e.g. Alzheimer's disease, FTDP-17, and CBD/PSP) and other related neurodegenerative diseases. The present commentary highlights the importance and limitations of the study, and discusses opportunities and directions for future investigations.

Download Full-text

IMMU-42. OPTIMIZING IMMUNOTHERAPY FOR GLIOMA USING CYTOKINE-ACTIVATED NATURAL KILLER CELLS

Neuro-Oncology ◽

10.1093/neuonc/noab196.401 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi101-vi102

Author(s):

Amber Kerstetter-Fogle ◽

Folashade Otegbeye ◽

David Soler ◽

Peggy Harris ◽

Alankrita Raghavan ◽

...

Keyword(s):

Nk Cells ◽

Natural Killer ◽

Glioma Cell ◽

Cell Function ◽

Nk Cell ◽

Cell Killing ◽

Cellular Immunotherapy ◽

Stereotactic Radiation ◽

Tumor Bed

Abstract INTRODUCTION Glioblastoma multiforme (GBM) is the most common primary central nervous system malignancy associated with a 12-15 month survival after surgery and radio-chemotherapy. Utilizing adoptive cellular immunotherapy using natural killer (NK) cells has developed over the past two decades for a variety of hematologic malignancies. This approach in solid malignancies is limited by questions of cell dose versus tumor burden, insufficient tumor infiltration, and a tumor microenvironment that suppresses NK cell function. METHODS We isolated NK cells from healthy volunteers and activated them using IL-2, -15, -12, -18, then perform cytotoxic assays in the presence of glioma stem cells. We also tested the efficacy of the NK cells with intracranial delivery in a pre-clinical murine model of glioma. We tested various concentrations of IL-2 and IL-15 on the cytokine culture platform. RESULTS In this study, we demonstrate human NK cells, activated using a cytokine cocktail of interleukins-2, -15, -12, and -18, exert strong cytotoxic events against glioma cell lines. To further examine the efficacy of activated NK cells in vitro, we utilized intracranially xenografted glioma lines and demonstrated a survival benefit with tumor bed injections of these cytokine-activated NK cells (p=0.0089). We were able to confirm that NK cells cultured with low doses (200u IL2; 50ng/ml IL15) of both cytokines are just as effective as higher doses. This is important, as in vivoexhaustion of NK cells stimulated with high doses of either cytokine has been well validated. We also found that low-dose irradiation (4Gy) of glioma cells prior to co-culture with cytokine-activated NK cells promoted increased targeted glioma cell killing within 4 hours(32% cell killing). CONCLUSIONS These findings suggest that in a clinical study, injection of cytokine-activated NK cells into the glioblastoma tumor bed could be used as adjuvant treatment following either stereotactic radiation or surgical resection.

Download Full-text

Inscuteable-dependent apical localization of the microtubule-binding protein Cornetto suggests a role in asymmetric cell division

Journal of Cell Science ◽

10.1242/jcs.114.20.3655 ◽

2001 ◽

Vol 114 (20) ◽

pp. 3655-3662 ◽

Cited By ~ 1

Author(s):

Silvia Bulgheresi ◽

Elke Kleiner ◽

Juergen A. Knoblich

Keyword(s):

Cell Fate ◽

Mitotic Spindle ◽

Binding Protein ◽

Protein Localization ◽

Apical Cell ◽

Coiled Coil ◽

Cell Cortex ◽

Dependent Manner ◽

Genetic Redundancy ◽

Microtubule Binding

Drosophila neuroblasts divide asymmetrically along the apical-basal axis. The Inscuteable protein localizes to the apical cell cortex in neuroblasts from interphase to metaphase, but disappears in anaphase. Inscuteable is required for correct spindle orientation and for asymmetric localization of cell fate determinants to the opposite (basal) cell cortex. Here, we show that Inscuteable also directs asymmetric protein localization to the apical cell cortex during later stages of mitosis. In a two-hybrid screen for Inscuteable-binding proteins, we have identified the coiled-coil protein Cornetto, which shows a highly unusual subcellular distribution in neuroblasts. Although the protein is uniformly distributed in the cytoplasm during metaphase, it concentrates apically in anaphase and forms an apical crescent during telophase in an inscuteable-dependent manner. Upon overexpression, Cornetto localizes to astral microtubules and microtubule spin-down experiments demonstrate that Cornetto is a microtubule-binding protein. After disruption of the actin cytoskeleton, Cornetto localizes with microtubules throughout the cell cycle and decorates the mitotic spindle during metaphase. Our results reveal a novel pattern of asymmetric protein localization in Drosophila neuroblasts and are consistent with a function of Cornetto in anchoring the mitotic spindle during late phases of mitosis, even though our cornetto mutant analysis suggests that this function might be obscured by genetic redundancy.

Download Full-text

Enhancing network activation in natural killer cells: predictions from in silico modeling

Integrative Biology ◽

10.1093/intbio/zyaa008 ◽

2020 ◽

Vol 12 (5) ◽

pp. 109-121

Author(s):

Sahak Z Makaryan ◽

Stacey D Finley

Keyword(s):

Nk Cells ◽

Natural Killer ◽

In Silico ◽

Intracellular Signaling ◽

Cell Activation ◽

Nk Cell ◽

Mechanistic Model ◽

Cell Killing ◽

Published Data ◽

Network Activation

Abstract Natural killer (NK) cells are part of the innate immune system and are capable of killing diseased cells. As a result, NK cells are being used for adoptive cell therapies for cancer patients. The activation of NK cell stimulatory receptors leads to a cascade of intracellular phosphorylation reactions, which activates key signaling species that facilitate the secretion of cytolytic molecules required for cell killing. Strategies that maximize the activation of such intracellular species can increase the likelihood of NK cell killing upon contact with a cancer cell and thereby improve efficacy of NK cell-based therapies. However, due to the complexity of intracellular signaling, it is difficult to deduce a priori which strategies can enhance species activation. Therefore, we constructed a mechanistic model of the CD16, 2B4 and NKG2D signaling pathways in NK cells to simulate strategies that enhance signaling. The model predictions were fit to published data and validated with a separate dataset. Model simulations demonstrate strong network activation when the CD16 pathway is stimulated. The magnitude of species activation is most sensitive to the receptor’s initial concentration and the rate at which the receptor is activated. Co-stimulation of CD16 and NKG2D in silico required fewer ligands to achieve half-maximal activation than other combinations, suggesting co-stimulating these pathways is most effective in activating the species. We applied the model to predict the effects of perturbing the signaling network and found two strategies that can potently enhance network activation. When the availability of ligands is low, it is more influential to engineer NK cell receptors that are resistant to proteolytic cleavage. In contrast, for high ligand concentrations, inhibiting phosphatase activity leads to sustained species activation. The work presented here establishes a framework for understanding the complex, nonlinear aspects of NK cell signaling and provides detailed strategies for enhancing NK cell activation.

Download Full-text