Serglycin-deficient Cytotoxic T Lymphocytes Display Defective Secretory Granule Maturation and Granzyme B Storage

Mirjana Grujic,Tiago Braga,Agneta Lukinius,Maija-Leena Eloranta,Stefan D Knight,Gunnar Pejler,Magnus Åbrink

doi:10.1074/jbc.m501708200

Abstract

Cytotoxic T lymphocytes eliminate infected and tumor cells mainly by perforin/granzyme-induced apoptosis. Earlier studies suggested that serglycin-proteoglycans form macromolecular complexes with granzymes and perforin in the cytotoxic granule. Serglycin-proteoglycans may also be involved in the delivery of the cytolytic machinery into target cells. We have developed a serglycin-deficient mouse strain, and here we studied the importance of serglycin-proteoglycans for various aspects of cytotoxic T lymphocyte function. 35SO4(2-) radiolabeling of serglycin-deficient cells demonstrated a dramatic reduction of incorporated label as compared with wild type cells, indicating that serglycin is by far the dominating proteoglycan species produced by the cytotoxic T lymphocyte. Moreover, lack of serglycin resulted in impaired ability of cytotoxic T lymphocytes to produce secretory granule of high electron density, although granule of lower electron density were produced both in wild type and serglycin-deficient cells. The serglycin deficiency did not affect the mRNA expression for granzyme A, granzyme B, or perforin. However, the storage of granzyme B, but not granzyme A, Fas ligand, or perforin, was severely defective in serglycin-deficient cells. Serglycin-deficient cells did not display defects in late cytotoxicity toward target cell lines. Taken together, these results point to a key role for serglycin in the storage of granzyme B and for secretory granule maturation but argue against a major role for serglycin in the apoptosis mediated by cytotoxic T lymphocytes.

Highlights

Cytotoxic T lymphocytes (CTLs)2 and natural killer (NK) cells are key players of the immune response where they eliminate harmful cells, e.g. pathogen-infected cells and tumor cells, by initiation of apoptosis
We have developed a serglycin-deficient mouse strain, and here we studied the importance of serglycin-proteoglycans for various aspects of cytotoxic T lymphocyte function. 35SO42؊ radiolabeling of serglycin-deficient cells demonstrated a dramatic reduction of incorporated label as compared with wild type cells, indicating that serglycin is by far the dominating proteoglycan species produced by the cytotoxic T lymphocyte
In 1985 it was reported that NK cells contained a protease-resistant chondroitin sulfate PG species that was released upon incubation of the NK cells with target cells, and the possibility that these PGs are involved in the packaging of cytolytic granule compounds was discussed [16, 17]

Summary

Introduction

Cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are key players of the immune response where they eliminate harmful cells, e.g. pathogen-infected cells and tumor cells, by initiation of apoptosis (reviewed in Refs. 1 and 2). It was initially thought that perforin induces pore formation in the plasma membrane of target cells and that the granzymes are delivered through this route (reviewed in Ref. 11). This model has been challenged by the finding that granzyme B can enter target cells autonomously [12] and through the identification of granzyme receptors on the surface of target cells [13, 14]. Proteoglycans (PGs) constitute a heterogeneous group of molecules, which are all composed of a protein core to which one or more glycosaminoglycan (GAG) chains are attached It has been known for a considerable time that PGs are components of the secretory granule of cytotoxic cells. SG is not essential for storage of granzyme A, perforin, or FasL and is not necessary for cytotoxic activity of CTLs

Methods

Results

Conclusion