DEREGULATION BY HU PROTEINS IN LUNG CANCER

Hu蛋白是一个RNA结合蛋白家族，包含三个RNA- 结合基序 HuD、HuC和Hel-N1在神经元组织中表达， 而HuR是普遍表达的。 神经元Hu蛋白也是 在所有小细胞肺癌（SCLC）中表达， 神经母细胞瘤 它们可以结合到不稳定的 原癌基因mRNA，如myc基因家族的那些。 Myc 在小细胞肺癌和神经母细胞瘤中， 通过增加Myc蛋白水平的事件。 建议的自然 Hu蛋白在RNA降解和神经元特异性 剪接会使这些蛋白质干扰正常的mRNA衰变 和癌细胞中的剪接。 该项目的具体目标是检验以下假设： 1)Hu蛋白在小细胞肺癌中的表达在转录后调控中起作用 通过改变降解和/或剪接模式的基因失调 影响细胞生长或粘附的基因编码蛋白质 特性. 2)增加细胞内Hu蛋白的水平， 没有或低水平的内源性蛋白质将导致 基因调节与观察到的一些异常相似 在SCLC。 为了验证我们的两个假设，我们提出了研究与 （a）确定以下方面的级别和特性： Hu蛋白在肺癌细胞系中表达，无论是小细胞肺癌还是肺癌， 非SCLC类型。 1b）测量表达水平和半衰期， 在缺乏和缺乏原癌基因的细胞中， 表达Hu蛋白，以确定 存在于一种或多种Hu蛋白的表达和不适当的myc mRNA降解。1c）为了确定特定的Hu蛋白是否显示出与Hu蛋白的结合。 对特定myc mRNA靶点的结合偏好，如c-，N-， 或L-myc非翻译区，并确定Hu 蛋白质在体内与myc mRNA相互作用。 1d）分析表达式 和编码细胞外基质蛋白的mRNA的剪接模式， SCLC和非SCLC类型细胞系中的生长刺激蛋白， 为了确定在细胞中是否观察到剪接变化， 表达一种或多种Hu蛋白。 2a）在细胞中表达Hu蛋白 通常缺乏它们，和2b）确定胡的后果 如上文1b和1d所概述的在这些细胞中的表达。2c）使用 在2a中建立的细胞系，以获得受 Hu蛋白表达。 Hu蛋白在小细胞肺癌中作用的研究 为我们开始理解RNA的作用提供了一个独特的机会， 结合蛋白在癌症的发展和进展中的作用。

Hu proteins are a family of RNA-binding proteins containing three RNA- binding motifs. HuD, HuC and Hel-N1 are expressed in neuronal tissue, while HuR is expressed ubiquitously. The neuronal Hu proteins are also expressed in all small cell lung cancers (SCLCs) and about half of all neuroblastomas. They can bind to the destabilizing sequences of labile proto-oncogene mRNAs, such as those of the family of myc genes. myc genes are frequently oncogenically activated in SCLC and neuroblastoma by events that increase the levels of Myc protein. The proposed natural function of Hu proteins in RNA degradation and neuronal-specific splicing would allow these proteins to interfere with proper mRNA decay and splicing in cancer cells. The specific aims of this project are to test the following hypotheses: 1) Hu protein expression in SCLC plays a role in post-transcriptional gene deregulation by altering the degradation and/or splicing pattern of mRNAs encoding proteins that affect the cell's growth or adhesion properties. 2) Increasing the levels of Hu proteins inside a cell with no or low endogenous levels of these proteins will lead to changes in gene regulation that are similar to some of the abnormalities observed in SCLC. To test our two hypotheses, we propose studies with the following specific aims: 1a) To determine the levels and identities of Hu proteins expressed in lung cancer cell lines, both of the SCLC and non-SCLC type. 1b) To measure the level of expression and half-life of mRNAs of the myc family of proto-oncogenes genes in cells lacking and expressing Hu proteins in order to determine whether a correlation exists between expression of one or more Hu proteins and improper myc mRNA degradation. 1c) To determine whether specific Hu proteins show a binding preference for particular myc mRNA targets, such as the c-, N-, or L-myc untranslated regions in vitro, and to determine whether Hu proteins interact with myc mRNA in vivo. 1d) To analyze the expression and splicing pattern of mRNAs encoding extracellular matrix proteins and growth stimulatory proteins in cell lines of the SCLC and non-SCLC type, in order to determine whether splicing changes are observed in cells expressing one or more Hu proteins. 2a) to express Hu proteins in cells that normally lack them, and 2b) to determine the consequences of Hu expression in these cells as outlined under 1b and 1d above. 2c) To use the cell lines established in 2a to obtain genes that are affected by Hu protein expression. The study of the role of Hu proteins in SCLC offers a unique opportunity to begin to understand the role of RNA- binding proteins in the development and progression of cancer.