Cloning and characterising an unusual perforin from chicken (Gallus gallus)

Abstract

In mammals the 67kDa pore-forming protein perforin is essential to the granule exocytosis pathway used by cytotoxic lymphocytes to eliminate virally infected and malignant cells. There is indirect evidence that this pathway exists in lower vertebrates such as teleost fish and birds, although in genome databases for the chicken and other bird species the perforin gene is incomplete and no full length expressed sequence tag has been reported. We present here the full gene and transcript sequence of chicken perforin. The inferred protein product contains an extended C-terminus that is at least 90 amino acids longer than any mammalian perforin, which is also evident in partial genomic sequences from other birds. To determine whether this extension is present in the translated protein, we raised two polyclonal antisera. The antisera identified a protein of just less than 80kDa in both transfected COS-1 cells and concanavalin A stimulated chicken splenocytes, indicating that the extended C-terminus is present in the mature protein. Our findings confirm that perforin exists in birds, and show that it is considerably longer than perforin of non-avian vertebrates.