Production of soluble HLA‐class‐I molecules by IFN‐γ‐induced colon‐adenocarcinoma cells

Abstract High levels of soluble HLA‐class‐l molecules (sHLA) were found to be produced in a time‐ and dose‐dependent manner by colon‐adenocarcinoma Colo20S cells in response to IFN‐γ stimulation. Among other cytokines tested, only IL‐6, TNF, IFN‐α and IFN‐β showed weak inducibility. IFN‐γ‐induced production of sHLA was synergistically enhanced by IL‐lα, IL‐6 or TNF, and combined treatment with TNF and IL‐6 exhibited an additive to synergistic induction. Expression of sHLA is unlikely to result from IFN‐γ‐induced enhancement of overall HLA‐class‐l expression, as at low concentrations IFN‐α, IFN‐β and IFN‐γ stimulated increased expression of cell membrane HLA‐class‐l molecules in Colo205 cells with almost equal efficiency, whereas only IFN‐γ induced high level production of sHLA. Immunoblot‐ting with a monoclonal antibody recognizing, β 2 ‐microglobulin‐free HLA‐class‐l heavy chain revealed 3 major forms of sHLA heavy chain, i.e., 45/43‐, 37‐ and 33‐kDa molecules, in the culture supernatants of IFN‐γ‐induced Colo20S cells. The 45/43‐kDa proteins can be partitioned into Triton X‐114, representing intact HLA‐class‐l heavy chains shed from the cell membrane. The hydrophilic 37‐ and 33‐kDa heavy chain, which remained almost exclusively in the aqueous phase after extraction with Triton X‐l 14, could well be due to alternative RNA splicing, with deletion of exon 5 encoding the hydrophobic transmembrane region of membrane‐anchored HLA‐class‐l heavy chain.