Liu J L, Chiles T C, Sen R J, Rothstein T L
Department of Microbiology, Boston University Medical Center, MA 02118.
J Immunol. 1991 Mar 1;146(5):1685-91.
Constitutive expression of NF-kappa B has been associated with developmental maturity in B cells on the basis of studies using continuously growing cell lines and plasmacytomas; however, little is known about the behavior of NF-kappa B in primary, mature B cells. In the present work, the regulation of NF-kappa B expression was studied by analyzing subcellular fractions of adult murine splenic B cells with the electrophoretic mobility shift assay using a kappa B-containing oligonucleotide. Although nuclear extracts from resting B cells contained kappa B-binding activity, additional kappa B-binding activity was present in cytosolic fractions in a form that became apparent after treatment with detergent. Competition analysis indicated that the DNA binding activity detected by electrophoretic gel mobility shift assay was specific for the kappa B motif, and UV photo-cross-linking showed the molecular size of kappa B-binding protein to be similar to that of the DNA binding subunit of NF-kappa B. Nuclear expression of kappa B-binding activity was markedly induced by treatment of B cells with phorbol ester or with LPS. Most notably, kappa B-binding activity was induced after surface IgR cross-linking, and the mechanism of this induction involved PKC. Further, anti-Ig-induced activity was superinduced in the presence of cycloheximide. These results indicate that nuclear NF-kappa B is rapidly induced as a result of B cell stimulation, and further suggest that NF-kappa B may play a specific role in mature B cells after ligand binding to surface Ig distinct from its postulated developmental role as a stage-specific factor involved in kappa-enhancer function.
基于使用连续生长的细胞系和浆细胞瘤的研究，NF-κB的组成型表达已与B细胞的发育成熟相关联；然而，对于原代成熟B细胞中NF-κB的行为知之甚少。在本研究中，通过使用含κB的寡核苷酸，采用电泳迁移率变动分析来分析成年小鼠脾B细胞的亚细胞组分，从而研究NF-κB表达的调控。尽管静息B细胞核提取物含有κB结合活性，但在胞质组分中也存在额外的κB结合活性，其形式在用去污剂处理后变得明显。竞争分析表明，通过电泳凝胶迁移率变动分析检测到的DNA结合活性对κB基序具有特异性，并且紫外线光交联显示κB结合蛋白的分子大小与NF-κB的DNA结合亚基相似。用佛波酯或脂多糖处理B细胞可显著诱导κB结合活性的核表达。最值得注意的是，表面IgR交联后可诱导κB结合活性，并且这种诱导机制涉及蛋白激酶C。此外，在存在环己酰亚胺的情况下，抗Ig诱导的活性被超诱导。这些结果表明，B细胞刺激可迅速诱导核NF-κB，并且进一步表明，NF-κB在配体结合至表面Ig后在成熟B细胞中可能发挥特定作用，这与其作为参与κ增强子功能的阶段特异性因子的假定发育作用不同。
