Marucha P T, Zeff R A, Kreutzer D L
Department of Periodontology, Ohio State University College of Dentistry, Columbus 43210.
J Immunol. 1991 Oct 15;147(8):2603-8.
We have previously demonstrated that Il-1 and TNF could rapidly, but transiently, induce gene expression of Il-1 beta in human polymorphonuclear leukocytes (PMN) at both the protein and mRNA level. Additionally, we demonstrated a cooperative effect of Il-1 and TNF on the kinetics of induction of Il-1 beta mRNA and protein. In order to better understand the molecular basis of Il-1 beta induction, we have further investigated the regulation of Il-1 and TNF-induced gene expression in the PMN. Using nuclear run-on transcription analysis, we found that within 1 h Il-1, TNF, and TNF plus Il-1 induced the transcription of the Il-1 beta gene by 33-, 61-, and 99-fold, respectively. By 2 h, the levels of transcription had been reduced to approximately 50% of peak levels for TNF- and TNF plus Il-1-treated PMN, and to near noninduced levels in Il-1-treated PMN. We also found that these cytokines induced stable mRNA, i.e., Il-1 beta mRNA t1/2 for Il-1-, TNF-, and TNF plus Il-1-induced PMN were 57, 94, and 86 min, respectively. By 2 h, when steady state levels of Il-1 beta mRNA were found to decrease, Il-1 beta mRNA t1/2 had fallen to approximately 18 min for all cytokine treatments. To determine if protein synthesis was required for induction of Il-1 beta gene expression, we treated PMN simultaneously with cytokines and cycloheximide, and found that cycloheximide enhanced the accumulation of Il-1-induced Il-1 beta mRNA, but abrogated the accumulation of Il-1 beta mRNA, by TNF- or TNF plus Il-1-treated PMN. This abrogation of Il-1 beta mRNA accumulation was not caused by inhibition of induction of Il-1 beta transcription because TNF induction of transcription of Il-1 beta was not affected by simultaneous treatment with cycloheximide. Thus, we report that Il-1 and TNF regulate IL-1 beta gene expression via both transcriptional and post-transcriptional mechanisms in vitro.
我们先前已证明，白细胞介素-1（Il-1）和肿瘤坏死因子（TNF）可迅速但短暂地在人多形核白细胞（PMN）中诱导Il-1β基因在蛋白质和mRNA水平的表达。此外，我们还证明了Il-1和TNF对Il-1β mRNA和蛋白质诱导动力学具有协同作用。为了更好地理解Il-1β诱导的分子基础，我们进一步研究了PMN中Il-1和TNF诱导的基因表达调控。通过核转录分析，我们发现，在1小时内，Il-1、TNF以及TNF加Il-1分别使Il-1β基因的转录增加了33倍、61倍和99倍。到2小时时，TNF和TNF加Il-1处理的PMN中，转录水平已降至峰值水平的约50%，而Il-1处理的PMN中则降至接近未诱导水平。我们还发现，这些细胞因子诱导了稳定的mRNA，即Il-1、TNF以及TNF加Il-1诱导的PMN中Il-1β mRNA的半衰期分别为57分钟、94分钟和86分钟。到2小时时，当发现Il-1β mRNA的稳态水平下降时，所有细胞因子处理下Il-1β mRNA的半衰期已降至约18分钟。为了确定Il-1β基因表达的诱导是否需要蛋白质合成，我们同时用细胞因子和环己酰亚胺处理PMN，发现环己酰亚胺增强了Il-1诱导的Il-1β mRNA的积累，但消除了TNF或TNF加Il-1处理的PMN中Il-1β mRNA的积累。Il-1β mRNA积累的这种消除不是由抑制Il-1β转录的诱导引起的，因为TNF对Il-1β转录的诱导不受同时用环己酰亚胺处理的影响。因此，我们报告，在体外，Il-1和TNF通过转录和转录后机制调节IL-1β基因表达。
