Fusion of a signal sequence to the interleukin-1β gene directs the protein from cytoplasmic accumulation to extracellular release

Anette Gjörloff Wingren, Olle Björkdahl, Tord Labuda, Lars Björk, Ulf Andersson, Urban Gullberg, Gunnar Hedlund, Hans Olov Sjögren, Terje Kalland, Bengt Widegren, Mikael Dohlsten

Forskningsoutput: TidskriftsbidragArtikelPeer review

34 Citeringar (Scopus)


Interleukin (IL)-1 differs from most other cytokines by the lack of a signal sequence, which results in the retention of the immature preform intracellularly (ic). Several cell types have the capacity to produce IL-1, but release has been shown to be restricted predominantly to monocytes/macrophages and associated with apoptosis of the producer cell. These features have limited the studies on IL-1 in early T cell-APC interactions. To develop a model for studying the biological effects of IL-1β release during long-lasting immune responses, we have established cells transfected with IL-1β cDNA constructs. To construct a hybrid gene for IL-1β release, the signal sequence from the related IL-1 receptor antagonist was fused to the gene encoding the 17-kDa mature form of IL-1β. A murine fibroblast cell line was transduced with retroviral technique and analyzed for the expression of human IL-1β, with or without a signal sequence (ssIL-1β and IL-1β, respectively). The fibroblasts transduced with either IL-1β or ssIL-1β expressed similar levels of human IL-1β mRNA. High levels of IL-1 bioactivity were recorded in freeze-thaw extracts from cells expressing the IL-1β protein ic, and in supernatants of ssIL-1β-transduced cells, which indicates that the initial formation of a preform of IL-1β is not required for correct folding of the protein. Treatment of ssIL-1β-transduced cells with Brefeldin A (BFA), an inhibitor of protein transport in the endoplasmatic reticulum, induced accumulation of the protein ic. BFA treatment did not affect IL-1β-transduced cells, while lipopolysaccharide-activated human monocytes increased the secretion of IL-1β. Cytoplasmic staining of single cells demonstrated that expression of the ssIL-1β gene directed the protein to a perinuclear Golgi-like compartment, whereas cells transduced with IL-1β cDNA showed a diffuse cytoplasmic distribution pattern. Secretion of IL-1β from human monocytes was under certain conditions accompanied by cell death. In contrasl, in the fibroblast cell line transduced to secrete IL-1β, no accompanying cell death could be detected. Gene targeting of IL-1 to the secretory or cytoplasmic pathway may be useful for elucidating the role of IL-1 in T cell-APC interactions, avoiding cell death of the producer cells.

Sidor (från-till)226-237
Antal sidor12
TidskriftCellular Immunology
StatusPublicerad - 1996-maj-01
Externt publiceradJa

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