Regulation of Gene Expression by DNA Binding Ligands

描述（由申请人提供）：可渗透人细胞的序列特异性DNA结合小分子可能调节特定基因的转录。在人类基因组已被绘制和测序的时代，生物学和人类医学领域已经揭示了许多人类疾病与异常基因表达有关，DNA结合配体的基础研究可能会导致功能基因组学试剂和可能的一类新的人类治疗方法。我们的目标是阐明设计小分子的化学原理，这些小分子以转录因子（TF）的亲和力和特异性结合预定的DNA序列。聚酰胺可以抑制TF在启动子中的结合，在无细胞系统中抑制（和激活）特异性基因表达，并结合浓缩的DNA、染色质。合成聚酰胺已被证明可以抑制原代人T细胞中的病毒基因表达，并调节复杂生物体（果蝇）中表型的获得（和丧失）。关于具体目标：（1）将探索一类新的与转运蛋白肽缀合的聚酰胺，用于增强核摄取，以拓宽细胞类型的库（人、酵母、果蝇），其允许核摄取，（2）将在细胞培养物中研究由连接至肽活化结构域的聚酰胺组成的人工转录活化剂，（3）将使用缀合至HDAC抑制剂的聚酰胺来调节转录，（4）一类新型聚酰胺嵌入剂将用于阐明MEF 2转录因子在间期染色质基质相关区域中的作用，（5）抑制逆转录病毒整合酶的聚酰胺将在细胞培养中进行测试，（6）聚酰胺将用于研究调节静息CD 4+淋巴细胞中HIV潜伏期的宿主细胞分子机制，（7）聚酰胺-苯丁酸氮芥缀合物将用于NB 4人细胞中以靶向PML启动子，和（8）聚酰胺将用于控制活斑马鱼胚胎中的基因表达和发育。

DESCRIPTION (provided by applicant): Sequence specific DNA binding small molecules that can permeate human cells could potentially regulate transcription of specific genes. In an era when the human genome has been mapped and sequenced and the fields of biology and human medicine have revealed that many human diseases are linked to aberrant gene expression, fundamental research in DNA binding ligands could lead to reagents for functional genomics and possibly a new class of human therapeutics. Our objective has been to elucidate chemical principles for the design of small molecules which bind predetermined DNA sequences with the affinity and specificity of transcription factors (TF). Polyamides can inhibit binding of TF in promoters, inhibit (and activate) specific gene expression in cell-free systems and bind condensed DNA, chromatin. Synthetic polyamides have been shown to inhibit viral gene expression in primary human T-cells, and to modulate gain (and loss) of phenotype in a complex organism (Drosophila). With regard to specific aims: (1) a new class of polyamides conjugated to transporter peptides for enhanced nuclear uptake will be explored to broaden the repertoire of cell types (human, yeast, Drosophila) which allow nuclear uptake, (2) artificial transcriptional activators comprised of polyamide linked to peptide activation domains will be studied in cell culture, (3) polyamides conjugated to HDAC inhibitors will be used to modulate transcription, (4) a new class of polyamide-intercalators will be used to elucidate the role of MEF2 transcription factor in the matrix associated region in interphase chromatin, (5) polyamides which inhibit retroviral integrase will be tested in cell culture, (6) polyamides will be used to study the host cell molecular mechanism that regulates HIV latency in resting CD4+ lymphocytes, (7) polyamides-chlorambucil conjugates will be used in NB4 human cells to target the PML promoter, and (8) polyamides will be used to control gene expression and development in living zebrafish embryos.