Fhit protein inhibits cell growth by attenuating the signaling mediated by nuclear factor-κB in colon cancer cell lines
Fhit protein inhibits cell growth by attenuating the signaling mediated by nuclear factor-κB in colon cancer cell lines
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DOI：
10.1016/j.yexcr.2006.04.004
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发表时间：
2006-08-01
影响因子：
3.7
通讯作者：
Akao, Yukihiro
中科院分区：
文献类型：
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作者：
Nakagawa, Yoshihito;Akao, Yukihiro
脆性组氨酸三联体（FHIT）基因参与了多种肿瘤中3p14.2区域的缺失。我们研究了Fhit蛋白在细胞生长中的作用，通过检查受Fhit影响的信号通路。本研究使用了3种人结肠癌细胞系SW 480、DLD-1和COLO 201。用pIRES 1 neo载体（SW/IRES细胞）、野生型FHIT载体（SW/FHIT细胞）或mt-FHIT（密码子96，His变为Asn）载体（SW/mt-FHIT细胞）转染完全不表达Fhit的SW 480细胞。与亲本或SWIRES细胞相比，SW/FHIT或SW/mt-FHIT细胞的生长受到抑制。特别是SW/FHIT细胞的生长受到明显抑制。另一方面，在携带Fhit的SW/FHIT或DLD-1细胞中通过siRNA沉默FHIT，表明与载体对照或非特异性siRNA对照相比，FHIT siRNA处理的细胞的生长显著增强。因此，我们发现Fhit对结肠癌细胞系中的细胞生长有负面贡献。此外，SW/FHIT细胞表现出更高的敏感性，线粒体电子传递或蛋白酶体的抑制剂诱发的氧化应激相比，任何控制转染。SW/FHIT细胞中磷酸化-I κ B-α（p-I κ B-α）的基线量与其他转染子中的相比减少。相反，FHIT siRNA处理的SW/FHIT和DLD-1细胞在FHIT的RNAi实验中表现出升高的p-I κ B-α水平。SW 480细胞的野生型Fhit表达体倾向于对柳氮磺胺吡啶或小白菊糖敏感，这是NF-κ B抑制剂，强烈提示了核因子（NF）-κ B信号传导的扰动。用TNF-α处理后，I κ B激酶（IKK）复合物（IKK α/β、磷酸-IKK α/β和IKK γ）水平的时间进程在转染子之间相似。尽管SW/FHIT细胞中的p-I κ B-α和磷酸化NF-κ B p65（p-NF-κ B）与其他转染子中的那些一样对TNF-α有反应，但在处理5分钟后，SW/FHIT细胞中的p-I κ B-α和p-NF-κ B水平的增加低于其他转染子。这些结果共同表明Fhit通过抑制I κ B-α的磷酸化从而阻断NF-κ B信号传导而作为抗癌蛋白发挥作用。(c)2006年爱思唯尔公司All rights reserved.
Fragile histidine triad (FHIT) gene is involved in the deletions at the 3p14.2 region in various cancers. We investigated the role of Fhit protein in cell growth by examining the signaling pathway affected by Fhit. We used 3 human colon cancer cell lines, SW480, DLD-1 and COLO201, in the study. SW480 cells, in which the expression of Fhit is completely absent, were transfected with pIRES1neo vector (SW/IRES cells), wild-type FHIT vector (SW/FHIT cells) or mt-FHIT (codon 96, His changed to Asn) vector (SW/mt-FHIT cells). The growth of SW/FHIT or SW/mt-FHIT cells was suppressed in comparison with that of parent or SWIRES cells. Especially, the growth of SW/FHIT cells was considerably suppressed. on the other hand, the silencing of FHIT by an siRNA for it in SW/FHIT or DLD-1 cells harboring Fhit demonstrated that the growth of FHIT siRNA-treated cells was significantly enhanced in comparison with that of the vector control or nonspecific siRNA control. Thus, we found that Fhit negatively contributed to cell growth in the colon cancer cell lines. Moreover, SW/FHIT cells exhibited a higher sensitivity to oxidative stress evoked by inhibitors of mitochondrial electron transport or proteasomes compared with any of the control transfectants. The base line amount of phospho-I kappa B-alpha (p-I kappa B-alpha) was reduced in SW/FHIT cells compared with that in the other transfectants. On the contrary, the FHIT siRNA-treated SW/FHIT and DLD-1 cells exhibited an elevated p-I kappa B-alpha level in an RNAi experiment on FHIT. Perturbation of nuclear factor (NF)-kappa B signaling was strongly suggested by the fact that the wild-type Fhit expressants of SW480 cells tended to be sensitive to sulfasarazine or parthenolide, which are inhibitors of NF-kappa B. The time course of the level Of I kappa B kinase (IKK) complex (IKK alpha/beta, phospho-IKK alpha/beta and IKK gamma) after the treatment with TNF-alpha was similar between the transfectants. Although p-I kappa B-alpha and phospho-NF-kappa B p65 (p-NF-kappa B) in SW/FHIT cells responded to TNF-alpha as those in other transfectants, the increase in the levels Of P-I kappa B-alpha and p-NF-kappa B after a 5-min treatment was less in SW/FHIT cells than in the other transfectants. These results altogether suggest that Fhit functions as an anti-oncoprotein by inhibiting the phosphorylation Of I kappa B-alpha and thereby blocking NF-kappa B signaling. (c) 2006 Elsevier Inc. All rights reserved.