Abstract
The expression, cellular localization, and activation of the NF-kappaB/Rel transcription factors are altered during neuronal differentiation, but the significance is unclear. Here we investigate the requirement for NF-kappaB/Rel proteins in neuronal differentiation. SH-SY5Y neuroblastoma cells were induced to differentiate with retinoic acid (RA) or 12-O-tetradecanoylphorbol 13-acetate (TPA), and differentiation was demonstrated by morphological criteria and the enhanced expression of Bcl-2. NF-kappaB was transiently activated after the addition of the differentiation inducers before the morphological signs of differentiation and the enhanced Bcl-2 synthesis. The onset of NF-kappaB activation coincided with a significant reduction in the amount of only one of four NF-kappaB-inhibitory proteins examined (I-kappaBbeta). In contrast, NF-kappaB activation and the reduction in I-kappaBbeta failed to occur in SH-SY5Y cells transformed with I-kappaBalphaM, a dominant-negative inhibitor of NF-kappaB/Rel proteins. These I-kappaBalphaM-expressing cells failed to differentiate into neuronal cell types when treated with RA or TPA, and the increased Bcl-2 synthesis was blocked. Therefore, NF-kappaB/Rel proteins are required for neuronal differentiation of SH-SY5Y neuroblastoma cells.
Highlights
The human SH-SY5Y neuroblastoma cell line is a well established system for studying neuronal differentiation [1, 2]
In one of several I-B␣Mexpressing SH-SY5Y clones, similar amounts of endogenous I-B␣ and the faster migrating I-B␣M protein were initially synthesized, but only I-B␣M steadily accumulated in the 24 h following retinoic acid (RA) or tetradecanoylphorbol 13-acetate (TPA) treatments (Fig. 1A, lower panels)
Up-regulation of Bcl-2 Is Blocked in I-B␣M-expressing SHSY5Y Cell Lines—Bcl-2 levels are progressively elevated during neuronal differentiation of some neuroblastoma cells induced by RA [17], TPA [18], or nerve growth factor [19]; and Bcl-2 up-regulation can be considered as a biochemical marker of neuronal differentiation [17,18,19,20,21,22]
Summary
The human SH-SY5Y neuroblastoma cell line is a well established system for studying neuronal differentiation [1, 2]. Studies on B cells from p50/NF-B knockout mice and on B cells and thymocytes transformed with I-B␣M, a strong dominant-negative inhibitor of the activation of different NFB/Rel complexes that bind I-B proteins, have indicated that NF-B is required for or participates in immune cell differentiation and development [8, 9].
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