Expression of tumour‐suppressing chemokine BRAK/CXCL14 reduces cell migration rate of HSC‐3 tongue carcinoma cells and stimulates attachment to collagen and formation of elongated focal adhesions in vitro
Expression of tumour‐suppressing chemokine BRAK/CXCL14 reduces cell migration rate of HSC‐3 tongue carcinoma cells and stimulates attachment to collagen and formation of elongated focal adhesions in vitro
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DOI：
10.1042/cbi20090108
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发表时间：
2010-05
影响因子：
3.9
通讯作者：
Kaori Sato;S. Ozawa;K. Izukuri;Y. Kato;R. Hata
中科院分区：
文献类型：
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作者：
Kaori Sato;S. Ozawa;K. Izukuri;Y. Kato;R. Hata
BRAK/CXCL 14（乳腺和肾脏表达的趋化因子/CXC趋化因子配体14）是一种在许多正常细胞和组织中表达的趋化因子，但在转化细胞和癌组织（包括HNSCC（头颈部鳞状细胞癌））中不存在或以非常低的水平表达。我们以前报道过，在HNSCC中，BRAK/CXCL 14的强制表达，与模拟载体导入相比，HSC-3 BRAK细胞降低了肿瘤形成率和肿瘤异种移植物的大小。（HSC-3模拟）细胞，即使这些细胞在体外培养条件下的生长速率相同，这表明该基因的高水平表达对于抑制体内肿瘤形成是重要的。为了研究BRAK依赖性肿瘤抑制机制的第一步，我们在体外培养条件下比较了HSC-3 BRAK和HSC-3 Mock细胞之间的特征。HSC-3 BRAK细胞的细胞迁移率低于HSC-3 Mock细胞。此外，当在I型胶原包被的培养皿上培养时，HSC-3 BRAK细胞显示出比HSC-3 Mock细胞更快的粘附，但在纤连蛋白或层粘连蛋白1包被的培养皿上则没有。这种粘附由α2β1整合素介导。对在I型胶原上培养的细胞的免疫荧光分析显示，HSC-3 BRAK细胞形成比HSC-3 Mock细胞更多的与桩蛋白和肌动蛋白应力纤维共定位的伸长的局灶性粘附。用重组BRAK处理亲本HSC-3细胞刺激了Rap 1的活化，Rap 1是ras家族的小GT3，并形成了细长的粘着斑，这表明在HSC-3 Mock和HSC-3 BRAK之间观察到的细胞特征差异不是由于选择了不同特征的克隆，而是由于细胞中BRAK的表达。HSC-3 BRAK细胞中粘着斑的特征性形态被引入HSC-3 BRAK细胞中的Ral鸟嘌呤核苷酸解离刺激剂的Rap结合结构域的表达载体（Rap 1的靶点）所扰乱，表明Rap 1调节粘着斑形态的形成。这些数据表明，BRAK的表达以自分泌或旁分泌方式刺激HSC-3细胞形成细长的局灶性粘连，其中刺激可能是细胞迁移减少的原因。
BRAK/CXCL14 (breast‐ and kidney‐expressed chemokine/CXC chemokine ligand 14) is a chemokine that is expressed in many normal cells and tissues but is absent from or expressed at very low levels in transformed cells and cancerous tissues, including HNSCC (head and neck squamous cell carcinoma). We reported previously that the forced expression of BRAK/CXCL14 in HNSCC (HSC‐3 BRAK) cells decreased the rate of tumour formation and size of tumour xenografts compared with mock‐vector‐introduced (HSC‐3 Mock) cells in athymic nude mice, even though the growth rates of these cells were the same under in vitro culture conditions, suggesting that high‐level expression of the gene is important for the suppression of tumour establishment in vivo. For the first step to study the mechanisms of BRAK‐dependent tumour suppression, we compared characteristics between HSC‐3 BRAK and HSC‐3 Mock cells under in vitro culture conditions. The cell migration rate was lower in HSC‐3 BRAK cells than in HSC‐3 Mock cells. Also, HSC‐3 BRAK cells showed more rapid adhesion than HSC‐3 Mock cells when cultured on type I collagen‐coated dishes but not on fibronectin or laminin 1‐coated ones. This adhesion was mediated by α2β1 integrin. Immunofluorescent analysis of the cells cultured on type I collagen showed that HSC‐3 BRAK cells formed much more elongated focal adhesions co‐localized with paxillin and actin stress fibres than did HSC‐3 Mock cells. Treatment of parental HSC‐3 cells with recombinant BRAK stimulated the activation of Rap1, which is a ras family small GTPase, and formation of elongated focal adhesions, indicating that the difference in cell character observed between HSC‐3 Mock and HSC‐3 BRAK was not due to selection of clones of different character but due to expression of BRAK in the cells. The characteristic morphology of focal adhesions in HSC‐3 BRAK cells was perturbed by the introduction of an expression vector of the Rap‐binding domain of the Ral guanine nucleotide dissociation stimulator, a target of Rap1, into HSC‐3 BRAK cells, suggesting that Rap1 regulated the formation of the morphology of the focal adhesions. These data indicate that the expression of BRAK stimulated the formation of elongated focal adhesions of the HSC‐3 cells in an autocrine or paracrine fashion, in which stimulation may be responsible for the reduced migration of the cells.