Abstract
Cytotoxic lymphocytes contain granules that have the ability to induce apoptosis in susceptible target cells. The granule contents include perforin, a pore-forming molecule, and several granzymes, including A and B, which are the most abundant serine proteases in these granules. Granzyme B-deficient cytotoxic T lymphocytes (CTL) have a severe defect in their ability to rapidly induce apoptosis in their targets, but have an intact late cytotoxicity pathway that is in part perforin-dependent. In this report, we have created mice that are deficient for granzyme A and characterized their phenotype. These mice have normal growth and development and normal lymphocyte development, activation, and proliferation. Granzyme A-deficient CTL have a small but reproducible defect in their ability to induce 51Cr and 125I-UdR release from susceptible allogeneic target cells. Since other granzyme A-like tryptases could potentially account for the residual cytotoxicity in granzyme A-deficient CTL, we cloned the murine granzyme K gene, which is linked to granzyme A in humans, and proved that it is also tightly linked with murine granzyme A. The murine granzyme K gene (which encodes a tryptase similar to granzyme A) is expressed at much lower levels than granzyme A in CTL and LAK cells, but its expression is unaltered in granzyme A-/- mice. The minimal cytotoxic defect in granzyme A-/- CTL could be due to the existence of an intact, functional early killing pathway (granzyme B dependent), or to the persistent expression of additional granzyme tryptases like granzyme K.
Highlights
The cytolytic granules of cytotoxic T lymphocytes (CTL)1 and natural killer (NK) cells contain several different neutral serine proteases called granzymes, the pore-forming protein perforin, and other proteins of unknown function
Our laboratory has demonstrated that the insertion of PGK-Neo cassette in the murine granzyme B gene disrupts the expression of granzymes C, D, F, and G in LAK/NK cells, potentially confounding interpretations of some of the results obtained from these mice [38]
Cytotoxic lymphocytes derived from these mice have normal programs of activation and proliferation, and express normal levels of the tightly linked granzyme K gene
Summary
The cytolytic granules of cytotoxic T lymphocytes (CTL)1 and natural killer (NK) cells contain several different neutral serine proteases called granzymes, the pore-forming protein perforin, and other proteins of unknown function. Genomic Organization and Linkage of the Granzyme K and A Genes in Mice—We screened a mouse BAC library derived from 129/SvJ mice using a 500-bp genomic fragment containing the alternatively spliced leader exons of mouse granzyme A, and obtained four clones (BAC 1254 –1257) that hybridized with this probe.
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