Expression, processing and transcriptional regulation of granulysin in short-term activated human lymphocytes.
BACKGROUND
Granulysin, a cytotoxic protein expressed in human natural killer cells and activated T lymphocytes, exhibits cytolytic activity against a variety of intracellular microbes. Expression and transcription have been partially characterised in vitro and four transcripts (NKG5, 519, 520, and 522) were identified. However, only a single protein product of 15 kDa was found, which is subsequently processed to an active 9 kDa protein.


RESULTS
In this study we investigated generation of granulysin in lymphokine activated killer (LAK) cells and antigen (Listeria) specific T-cells. Semiquantitative RT-PCR revealed NKG5 to be the most prominent transcript. It was found to be up-regulated in a time-dependent manner in LAK cells and antigen specific T-cells and their subsets. Two isoforms of 519 mRNA were up-regulated under IL-2 and antigen stimulation. Moreover, two novel transcripts, without any known function, comprising solely parts of the 5 prime region of the primary transcript, were detected. A significant increase of granulysin expressing LAK cells as well as antigen specific T-cells was shown by fluorescence microscopy. On the subset level, increase in CD4+ granulysin expressing cells was found only under antigen stimulation. Immunoblotting showed the 15 kDa form of granulysin to be present in the first week of stimulation either with IL-2 or with bacterial antigen. Substantial processing to the 9 kDa form was detected during the first week in LAK cells and in the second week in antigen specific T-cells.


CONCLUSIONS
This first comprehensive study of granulysin gene regulation in primary cultured human lymphocytes shows that the regulation of granulysin synthesis in response to IL-2 or bacterial antigen stimulation occurs at several levels: RNA expression, extensive alternative splicing and posttranslational processing.
Latinovic-Golic
Sonja
S
Division of Cell Biology, Institute of Anatomy, Zürich, Switzerland. sonja.latinovic@anatom.unizh.ch <sonja.latinovic@anatom.unizh.ch>
Walch
Michael
M
Sundstrom
Hanna
H
Dumrese
Claudia
C
Groscurth
Peter
P
Ziegler
Urs
U
D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics* / Q000378:metabolism*; D000939:Epitopes / Q000276:immunology; D005786:Gene Expression Regulation; D006801:Humans; D015979:Killer Cells, Lymphokine-Activated / Q000166:cytology / Q000276:immunology*; D007700:Kinetics; D008087:Listeria / Q000276:immunology; D008213:Lymphocyte Activation; D018613:Microscopy, Confocal; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology*; D013997:Time Factors; D014158:Transcription, Genetic
D000945:Antigens, Differentiation, T-Lymphocyte; D000939:Epitopes; C075005:GNLY protein, human; D012333:RNA, Messenger