Loikkanen Ildikó, Toljamo Katja, Hirvikoski Pasi, Väisänen Timo, Paavonen Timo K, Vaarala Markku H
Department of Pathology, University of Oulu, Oulu University Hospital, Oulu, 90029 OYS, Finland.
Oncol Rep. 2009 Nov;22(5):991-5. doi: 10.3892/or_00000526.
Myosin VI, one of the so-called unconventional myosins, is an actin-based molecular motor involved in intracellular vesicle and organelle transport. In human prostate, myosin VI is expressed in prostate epithelium. We examined the effect of myosin VI downregulation in the LNCaP human prostate cancer cell line using an RNA interference approach. Further, the expression of myosin VI in human prostate tissue was examined using immunohistochemistry. The expression of androgen receptor (AR) and E-cadherin was examined in myosin VI knocked-down cells and control cells. We determined 3H-testosterone uptake in the myosin knocked-down LNCaP cells. Next, we cotransfected LNCaP cells with the myosin VI-specific small interfering RNA (siRNA) duplex and an androgen-responsive luciferase reporter construct and then measured luciferase activity after androgen induction. To clarify whether myosin VI and the AR are interacting proteins, we performed immunoprecipitation studies using myosin VI and AR polyclonal antibodies in androgen-induced LNCaP cells. We confirmed previous results of myosin VI overexpression in human prostate cancer tissue, as in some cases malignant epithelium was more intensively stained than benign epithelium. We found that the expression of AR decreased as a result of myosin VI knock-down. Decreased myosin VI levels did not significantly influence the testosterone uptake of the LNCaP cell line. Instead, we noted a decreased activity of the androgen-regulated mouse mammary tumor virus promoter-reporter vector construct in LNCaP cells cotransfected with myosin VI siRNA duplexes. Finally, we detected the interaction between AR and myosin VI by immunoprecipitation. We propose that myosin VI is a modulator of androgen-dependent gene transcription via interaction with the AR. Thus, myosin VI is a potential therapeutic target for prostate cancer as it could be used as a modulator of AR-dependent gene expression.
肌球蛋白VI是一种所谓的非常规肌球蛋白，是一种基于肌动蛋白的分子马达，参与细胞内囊泡和细胞器运输。在人类前列腺中，肌球蛋白VI在前列腺上皮中表达。我们使用RNA干扰方法研究了肌球蛋白VI下调对LNCaP人前列腺癌细胞系的影响。此外，使用免疫组织化学方法检测了人前列腺组织中肌球蛋白VI的表达。检测了肌球蛋白VI敲低细胞和对照细胞中雄激素受体（AR）和E-钙黏蛋白的表达。我们测定了肌球蛋白敲低的LNCaP细胞中3H-睾酮的摄取。接下来，我们用肌球蛋白VI特异性小干扰RNA（siRNA）双链体和雄激素反应性荧光素酶报告构建体共转染LNCaP细胞，然后在雄激素诱导后测量荧光素酶活性。为了阐明肌球蛋白VI和AR是否为相互作用蛋白，我们在雄激素诱导的LNCaP细胞中使用肌球蛋白VI和AR多克隆抗体进行了免疫沉淀研究。我们证实了先前关于人前列腺癌组织中肌球蛋白VI过表达的结果，因为在某些情况下，恶性上皮比良性上皮染色更强烈。我们发现，由于肌球蛋白VI敲低，AR的表达降低。肌球蛋白VI水平降低并未显著影响LNCaP细胞系的睾酮摄取。相反，我们注意到在与肌球蛋白VI siRNA双链体共转染的LNCaP细胞中，雄激素调节的小鼠乳腺肿瘤病毒启动子-报告载体构建体的活性降低。最后，我们通过免疫沉淀检测到AR和肌球蛋白VI之间的相互作用。我们提出，肌球蛋白VI通过与AR相互作用是雄激素依赖性基因转录的调节因子。因此，肌球蛋白VI可能是前列腺癌的潜在治疗靶点，因为它可作为AR依赖性基因表达的调节剂。
