Tomaru Takuya, Satoh Teturou, Yoshino Satoshi, Ishizuka Takahiro, Hashimoto Koshi, Monden Tsuyoshi, Yamada Masanobu, Mori Masatomo
Department of Medicine and Molecular Science, Gunma University Graduate School of Medicine, Maebashi 371-8511, Japan.
Endocrinology. 2006 Jan;147(1):377-88. doi: 10.1210/en.2005-0450. Epub 2005 Oct 20.
Using the DNA-binding domain (DBD) and hinge region of human peroxisome proliferator-activated receptor (PPAR)-gamma as bait in yeast two-hybrid screen, we isolated partial cDNA identical with that of the C terminal of KIAA1769. KIAA1769 encodes a 2080-amino acid protein (molecular mass, 231 kDa) that was recently identified to interact with PPARalpha and termed PPARalpha-interacting cofactor 285 (here referred to as PPARgamma-DBD-interacting protein 1 (PDIP1)-alpha). PDIP1 mRNA was expressed in 3T3-L1 adipocytes and THP-1 macrophages. We also identified the expression of the N terminal extended form of PDIP1alpha (referred to as PDIP1beta) consisting of 2649 amino acids (295 kDa) in human cultured cell lines by RT-PCR, and 5' rapid amplification of cDNA ends. Ribonuclease protection assay revealed that PDIP1beta mRNA was expressed more abundantly than PDIP1alpha mRNA. The C-terminal region of PDIP1 directly binds DBD of PPARgamma, and multiple LXXLL motifs in PDIP1 were not required for the interaction. PDIP1alpha and -beta similarly enhanced PPARgamma-mediated transactivation in transfection assays and short interfering RNA targeting PDIP1 mRNA significantly reduced transactivation by PPARgamma. No potent intrinsic activation domain was identified in either PDIP1 isoforms in mammalian one-hybrid assays, and mutation of all LXXLL motifs did not affect enhancement of PPARgamma-mediated transactivation. PDIP1alpha and -beta similarly augmented transactivation by PPARalpha, PPARdelta, thyroid hormone receptor (TR)-alpha1, TRbeta1, and retinoid X receptor-alpha. PDIP1alpha also enhanced estrogen receptoralpha- and androgen receptor-mediated transactivation, whereas PDIP1beta did not. PDIP1alpha showed receptor-specific synergism with activation function-2-interacting coactivators in PPARgamma- and TRbeta1-mediated transactivation. Together, PDIP1 might function as a transcriptional cofactor for a broad range of nuclear receptors, possibly in collaboration with specific activation function-2 interacting coactivators.
在酵母双杂交筛选中，我们以人过氧化物酶体增殖物激活受体（PPAR）-γ的DNA结合结构域（DBD）和铰链区作为诱饵，分离出了与KIAA1769 C末端相同的部分cDNA。KIAA1769编码一种2080个氨基酸的蛋白质（分子量为231 kDa），该蛋白质最近被鉴定为与PPARα相互作用，并被命名为PPARα相互作用辅因子285（此处称为PPARγ-DBD相互作用蛋白1（PDIP1）-α）。PDIP1 mRNA在3T3-L1脂肪细胞和THP-1巨噬细胞中表达。我们还通过逆转录-聚合酶链反应（RT-PCR）和5' cDNA末端快速扩增，在人培养细胞系中鉴定了由2649个氨基酸（295 kDa）组成的PDIP1α N末端延伸形式（称为PDIP1β）的表达。核糖核酸酶保护试验显示，PDIP1β mRNA的表达比PDIP1α mRNA更丰富。PDIP1的C末端区域直接结合PPARγ的DBD，并且PDIP1中的多个LXXLL基序对于这种相互作用不是必需的。在转染试验中，PDIP1α和-β同样增强了PPARγ介导的反式激活，并且靶向PDIP1 mRNA的小干扰RNA显著降低了PPARγ的反式激活。在哺乳动物单杂交试验中，在两种PDIP1亚型中均未鉴定到有效的内在激活结构域，并且所有LXXLL基序的突变均不影响PPARγ介导的反式激活的增强。PDIP1α和-β同样增强了PPARα、PPARδ、甲状腺激素受体（TR）-α1、TRβ1和视黄酸X受体-α的反式激活。PDIP1α还增强了雌激素受体α和雄激素受体介导的反式激活，而PDIP1β则没有。在PPARγ和TRβ1介导的反式激活中，PDIP1α与激活功能-2相互作用共激活因子表现出受体特异性协同作用。总之，PDIP1可能作为多种核受体的转录辅因子发挥作用，可能与特定的激活功能-2相互作用共激活因子协同作用。
