RI-alpha/RIAZ on Cell Growth in Breast Cancer

描述(申请人提供)：cAMP对细胞生长和增殖的影响已被深入研究，但其作用机制尚不完全清楚。CAMP的作用主要由cAMP依赖的蛋白激酶(PKA)介导，PKA由催化(C)和调节(R)两个不同的亚基组成，形成四聚体全酶R2C2。I型调节性α(RIpha)亚单位的表达与人乳腺组织的过度增殖有关，其在乳腺癌中的过度表达与恶性程度和预后不良有关。RI？表达增加？刺激生长，而C亚单位的过度表达不会产生这样的结果。最近，RIpha被证明是Carney综合征的一种肿瘤抑制基因。我们以前已经证明了RIpha的新的相互作用是独立于C亚单位激酶活性的。在这项应用中，我们通过酵母双杂交克隆实验表明，RIpha与一个新的BTB/POZ结构域的锌指转录因子相关联，称为RIpha相关的锌指蛋白(RiaZ)。我们证明RiaZ是一个cAMP反应性转录激活因子，受其与RIpha的相互作用以及与cAMP反应元件结合蛋白(CREB)的潜在合作调节。我们进一步证明RiaZ在大约15%的人类原发乳腺癌中异常表达。此外，过表达RiaZ可通过[~3H]胸腺嘧啶核苷摄取量来衡量细胞的生长抑制。CAMP对细胞生长的抑制作用增强，但不受PKA抑制剂H-89的影响。我们假设RI？与Riaz的相互作用可能是cAMP抑制生长的一种新的转录机制。在这项建议中，我们将：(1)确定RI的机制？与Riaz的相互作用和cAMP的调节；(2)研究Riaz对cAMP转录反应的机制；(3)研究RI？与里亚兹互动以回应夏令营。我们的结果将有助于揭示RIpha与Riaz在抑制生长中的这种新的相互作用。

DESCRIPTION (provided by applicant): The effects of cAMP on cell growth and proliferation have been intensely investigated, but its mechanisms of action are not completely understood. The effects of cAMP are predominantly mediated by the cAMP-dependent protein kinase (PKA), which is composed of two distinct subunits, catalytic (C) and regulatory (R), forming a tetrameric holoenzymes, R2C2. The type I regulatory alpha (RIalpha) subunit expression is associated with hyperproliferation in human breast tissue and its overexpression in human breast cancer correlates with malignancy and poor prognosis. Increased expression of RI? stimulates growth, whereas overexpression of the C subunit does not produce such consequence. Most recently, RIalpha was shown to be a tumor suppressor gene in Carney Complex Syndrome. We have demonstrated previously novel interaction of RIalpha that is independent of the C subunit kinase activity. In this application, we show by yeast two-hybrid interaction cloning experiment that RIalpha associates with a novel BTB/POZ domain zinc finger transcription factor, termed RIalpha-associated zinc finger protein (RIAZ). We demonstrate that RIAZ is a cAMP-responsive transcriptional activator regulated by its interaction with RIalpha and potential cooperation with the cAMP-response element binding protein (CREB). We show further that RIAZ is aberrantly expressed in approximately 15% of human primary breast cancer. Furthermore, overexpression of RIAZ causes growth inhibition measured by [3H]thymidine uptake. The growth inhibition is enhanced by cAMP but not affected by the PKA inhibitor H-89. We hypothesize that RI? interaction with RIAZ may be a novel transcriptional mechanism in growth inhibition transduced by cAMP. In this proposal, we will: (1) determine the mechanisms of RI? interaction with RIAZ and regulation by cAMP; (2) investigate the mechanisms of RIAZ transcriptional response to cAMP; and (3) investigate the mechanisms of growth control by RI? interaction with RIAZ in response to cAMP. Our results will shed light on this novel interaction of RIalpha with RIAZ in growth inhibition.