title: Cloning and characterising an unusual perforin from chicken (Gallus gallus).
abstract: In mammals the 67 kDa 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 80 kDa 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.
authors.affiliation: Department of Biochemistry and Molecular Biology, Monash University, Wellington Road, Clayton, VIC 3800, Australia. michael.dangelo@monash.edu
authors.forename: Michael E
authors.initials: ME
authors.lastname: D'Angelo
authors.forename: Arash
authors.initials: A
authors.lastname: Arjomand
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Trapani
authors.forename: Phillip I
authors.initials: PI
authors.lastname: Bird
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D030161:Avian Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D019556:COS Cells; D002478:Cells, Cultured; D002645:Chickens / Q000235:genetics* / Q000378:metabolism; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D003208:Concanavalin A / Q000494:pharmacology; D015870:Gene Expression; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D054353:Perforin / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism
chemical_list: D030161:Avian Proteins; D003208:Concanavalin A; D054353:Perforin
keywords: CL; Chicken; ConA; Cytotoxic T lymphocyte; Granule exocytosis; MHC; Natural killer cell; Perforin; RACE; T cell receptor; TCR; concanavalin A; cytotoxic lymphocyte; major histocompatibility complex; rapid amplification of cDNA ends