Interferon-alpha but not -beta genes require de novo protein synthesis for efficient expression in human monocytes.
Monocytes produce interferon-alpha (IFN)-alpha) and -beta when human peripheral blood mononuclear cells (PBMCs) are stimulated in vitro by Sendai virus (SV). We found that about 70% of the IFN-producing cells (IPCs) expressed both IFN-alpha and -beta mRNA; the rest expressed only IFN-beta mRNA. In the presence of the protein synthesis inhibitor cycloheximide (CHX), the frequency of IFN-alpha mRNA-containing cells, measured after 6h, was decreased by 85-90%. Results of nuclear run-on transcription assays showed that CHX inhibited IFN-alpha gene expression. The frequency of IFN-beta mRNA-containing cells was not reduced by CHX. Actually, a threefold increase was observed at the lower CHX concentrations. Studies on the kinetics of IFN-alpha/beta mRNA induction showed that CHX accelerated the appearance of IFN-beta mRNA-containing cells, increased IFN-beta mRNA levels, and delayed the normally occurring post-inductional decrease of IFN-beta mRNA. Unexpectedly, an initially normal or even accelerated IFN-alpha mRNA response was seen in the presence of CHX during the first 3-4 h after SV stimulation. This occurred in a small proportion of the potential IPCs. However, CHX prevented the subsequent marked increase of IFN-alpha mRNA levels. Preincubation of PBMCs for 6 h in conditioned medium (CM) containing IFN and other cytokines prevented the CHX-mediated inhibition of IFN-alpha mRNA. Without preincubation this was not seen. The preincubation in CM caused an accelerated appearance of IFN-alpha mRNA, resembling that of IFN-beta mRNA. The results suggest that IFN-alpha and -beta genes are differentially regulated in the same monocytes, the former requiring de novo synthesis of intracellular protein(s) for efficient expression.
Gobl
A E
AE
Interferon Laboratory, Uppsala University, Sweden.
Cederblad
B
B
Sandberg
K
K
Alm
G V
GV
D015152:Blotting, Northern; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D015342:DNA Probes; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D016898:Interferon-alpha / Q000096:biosynthesis / Q000235:genetics*; D016899:Interferon-beta / Q000096:biosynthesis / Q000235:genetics*; D009000:Monocytes / Q000378:metabolism*; D010222:Parainfluenza Virus 1, Human / Q000276:immunology; D012333:RNA, Messenger / Q000187:drug effects
D015342:DNA Probes; D016898:Interferon-alpha; D012333:RNA, Messenger; D016899:Interferon-beta; D003513:Cycloheximide