Exploiting the RUSH System to Study Lytic Granule Biogenesis in Cytotoxic T Lymphocytes

Nagaja Capitani; Chiara Cassioli; Keerthana Ravichandran; Cosima T Baldari

doi:10.1007/978-1-0716-3135-5_27

Exploiting the RUSH System to Study Lytic Granule Biogenesis in Cytotoxic T Lymphocytes

Methods Mol Biol. 2023:2654:421-436. doi: 10.1007/978-1-0716-3135-5_27.

Authors

Nagaja Capitani¹, Chiara Cassioli², Keerthana Ravichandran³, Cosima T Baldari²

Affiliations

¹ Department of Life Sciences, University of Siena, Siena, Italy. [email protected].
² Department of Life Sciences, University of Siena, Siena, Italy.
³ Cellular Neurophysiology, Center for Integrative Physiology and Molecular Medicine (CIPMM), Saarland University, Homburg, Germany.

Abstract

The Retention Using Selective Hooks (RUSH) system allows for the synchronized release of one or more proteins of interest from a donor endomembrane compartment, usually the endoplasmic reticulum, and the subsequent monitoring of their traffic toward acceptor compartments. Here we describe the RUSH system applied to cytotoxic T cells to characterize the biogenesis of lytic granules, using as a proof-of-concept granzyme B trafficking to this specialized compartment.

Keywords: Confocal microscopy; Cytotoxic granules; Granzyme B; Intracellular trafficking; Lytic granule biogenesis; RUSH system; Spinning disk microscopy.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cytoplasmic Granules / metabolism
Proteins* / metabolism
T-Lymphocytes, Cytotoxic* / metabolism

Substances

Proteins

Grants and funding

951329/ERC_/European Research Council/International