ZNF326通过DBC1促进乳腺癌恶性表型的机制

Previous studies had demonstrated that ZNF326 could interact with DBC1,however, the expression pattern of ZNF326 as well as its biological role were still uncertain. We found that ZNF326 was highly expressed in triple-negative breast cancer and significantly correlated with advanced TNM stage and positive lymph node metastasis. Overexpressing ZNF326 may upregualte Snail and CyclinB1, and downregulate E-cadherin, thus enhanced the malignant phonotype of breast carcinoma. We speculated that ZNF326 may bind with the S1RNA domain in the N terminal of DBC1 through its C2H2 zinc finger domain. The interaction between ZNF326 and DBC1 may activate ERK/AKT signaling, thus promoted the process of tumor progression. We will up- or down-regulated the expression of ZNF326 and DBC-1 and construct mutant that ZNF326 could not bind with DBC-1 as well as the mutant that ZNF326 and DBC1 could not translocate into the nucleus, using co-IP, EMSA , western blot and Real-time PCR, to find out the binding site which ZNF326 bond with DBC1, to clarify the effect on the activity of ERK and/or AKT signaling and their downstream factor CyclinB1,Snail and E-cadherin.We also tried to explore, both in vivo and in vitro, the mechanism on how ZNF326 promoted proliferation, invasion and metastasis of breast carcinoma via binding with DBC1, and evaluated the clinical therapy value of ZNF326 in triple-negative breast cancer.

已知ZNF326可以与DBC1结合，但其在肿瘤中表达意义及作用机制不清。我们发现ZNF326在三阴性乳腺癌中明显高表达且与高TNM分期和淋巴结转移相关，转染ZNF326可通过上调CyclinB1和Snail，下调E-cadherin促进乳腺癌细胞恶性表型。我们推测ZNF326通过其锌指结构与DBC1的N端S1RNA结构域结合后，通过激活ERK和/或AKT信号活性，发挥其促癌作用。本研究拟双向表达ZNF326和DBC1，并转染二者各种突变质粒,使用免疫共沉淀DNA电泳迁移率、免疫印迹及实时定量PCR等方法，证实二者的结合位点，明确二者在不同的结合状态下对ERK和/或AKT活性及下游蛋白表达的影响。结合体外细胞功能试验、裸鼠移植试验和大样本临床标本检测和分析等，探索ZNF326通过DBC1促进乳腺癌细胞恶型表型的分子机制，评估检测ZNF326在三阴性乳腺癌中的临床价值。

锌指蛋白-326（Zinc-finger protein-326）在恶性肿瘤中的作用和机制尚没有报道。我们发现ZNF326在乳腺癌中存在着核内高表达（60/111，54.1%）,并与乳腺癌患者的高pTNM分期（p=0.002），淋巴结转移（p=0.004）和不良预后（p=0.0097）呈正相关，并且与ER/PR/HER-2状态有关。转染ZNF326可上调MMP7，EMT-相关蛋白（Snail和Slug）和细胞周期相关蛋白 （cyclinA2和cyclinB1），下调E-cadherin的表达，在体内外发挥促进乳腺癌细胞的增殖和侵袭能力。免疫共沉淀和免疫荧光实验证实ZNF326与DBC1在乳腺癌细胞中存在着相互作用和共定位；敲除DBC1后可以降低ZNF326上调MMP7、EMT-和细胞周期相关蛋白的表达、促进乳腺癌细胞的增殖和侵袭能力的作用。综上，ZNF326在乳腺癌中高表达并与不良预后相关，ZNF326通过与DBC1相互作用发挥促进乳腺癌细胞恶性表型的生物学功能。