NOVEL TRANSLOCATION PRODUCT IN APL

粒细胞成熟停滞的分子机制 急性早幼粒细胞白血病(APL)的特征仍不清楚。多数 APL患者表现为t(15；17)染色体易位， 结果表达了PML-RAR融合蛋白。竞争车型 提示PML-RAR是PML的显性否定，PML是显性的 野生型维甲酸受体α(RARA)阴性，或流氓 转录激活剂。为了更深入地了解 APL的潜在机制我们研究了一例带有t(5；17)的APL患者 染色体易位。我们发现这种易位将5‘ 核仁蛋白核磷脂(NPM)的成分到核仁蛋白的3‘端成分 拉拉。在我们的初步数据中，我们鉴定了嵌合cDNA并对其进行了测序， 并证明其编码52kd的蛋白质。我们假设这是 融合通过一种新的方式影响保留的RAR结构域的功能 改变RAR结构域所在核区的机制 隔离。我们假设NPM基序将RAR结构域重定向到一个 异常的核隔室，融合在那里结合并隔离RAR 异二聚化伙伴。通过这种新的机制重定向其 融合到核的一个不习惯的建筑区域， NPM将链接的RAR结构域转换为对RARA的显性否定，并且 从而阻止髓系成熟。为了检验这一假设，我们将 完成NPM-RAR的全开放阅读框架的测序，以及 确定融合蛋白确实在t(5；17)中表达 白血病细胞。我们将确定NPM-RAR是否复制了APL 通过研究NPM-RAR强制表达对细胞表型的影响 HL-60细胞的分化潜能。我们将确定是否 NPM-RAR的表达调节野生型RARA功能，以及NPM-RAR是否 与RARA异二聚体的络合物。我们将通过以下方式确定 免疫荧光检测NPM-RAR在细胞内的分布，以及 它的表达影响了拉拉所在的建筑隔间 二聚化合作伙伴本地化。最后，我们将确定一个 将RAR引导到一个不习惯的核的人造结构 构造域类似地阻止髓系分化。这些 调查将检验破坏维甲酸的假说 信号转导是APL发育成熟的基础。此外，他们还将 作为未来绘制核功能图工作的开创性研究 车厢。

The molecular mechanism underlying the granulocytic maturation arrest that characterizes Acute Promyelocytic Leukemia (APL) remains obscure. Most patients with APL manifest the t(15;17) chromosomal translocation, which results in expression of the PML-RAR fusion protein. Competing models suggest that PML-RAR acts as a dominant negative for PML, a dominant negative for wild type retinoic acid receptor alpha (RARa), or a rogue transcriptional activator. In an effort to gain greater insight into the mechanism underlying APL we have studies an APL patient with a t(5;17) chromosomal translocation. We show that this translocation links 5' elements of the nucleolar protein nucleophosmin (NPM) to 3' elements of RARa. In our preliminary data we identify and sequence the chimeric cDNA, and demonstrate that it encodes a 52 kd protein. We hypothesize that this fusion affects the function of the retained RAR domains through a novel mechanism of changing the nuclear compartment in which the RAR domains segregate. We hypothesize that NPM motifs redirect RAR domains into an aberrant nuclear compartment, where the fusion binds to and sequesters RAR heterodimerization partners. By this novel mechanism of redirecting its fusion partner into an unaccustomed architectural region of the nucleus, NPM converts the linked RAR domain into a dominant negative for RARa, and thereby blocks myeloid maturation. To test this hypothesis we will complete the sequencing of the full open reading frame for NPM-RAR, and establish that the fusion protein is indeed expressed in the t(5;17) leukemic cells. We will determine whether NPM-RAR reproduces the APL phenotype by investigating the effects of forced expression of NPM-RAR on the differentiation potential of HL-60 cells. We will determine whether NPM-RAR expression modulates wild type RARa function, and whether NPM-RAR complexes with RARa heterodimerization partners. We will determine by immunofluorescence the distribution of NPM-RAR in the cell, and whether its expression affects the architectural compartment in which RARa dimerization partners localize. Finally, we will determine whether an artificial construct that directs RAR to an unaccustomed nuclear architectural domain similarly blocks myeloid differentiation. These investigations will test the hypothesis that disrupted retinoic acid signaling underlies the maturational block in APL. In addition, they will serve as seminal studies for future work mapping nuclear functional compartments.