Identification of an endogenous ligand bound to a native orphan nuclear receptor.
Identification of an endogenous ligand bound to a native orphan nuclear receptor.
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DOI：
10.1371/journal.pone.0005609
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发表时间：
2009
期刊：
影响因子：
3.7
通讯作者：
Sladek FM
中科院分区：
文献类型：
--
作者：
Yuan X;Ta TC;Lin M;Evans JR;Dong Y;Bolotin E;Sherman MA;Forman BM;Sladek FM
孤儿核受体在识别新的信号通路和治疗靶点方面发挥了重要作用。然而，识别这些受体的配体通常是基于随机化合物筛选或其他有偏见的方法。因此，在许多情况下，尚不清楚所报道的配体是否是真正的内源性配体，即在体内未受干扰的情况下与受体结合的配体。技术上的限制限制了我们根据这一严格定义识别配体的能力。孤儿受体肝细胞核因子4α(HNF4α)是许多代谢途径的关键调节因子，与糖尿病、动脉粥样硬化、血友病和癌症等多种疾病有关。在这里，我们利用亲和分离/质谱仪(AIMS)的方法证明，在哺乳动物细胞和喂养的小鼠的肝脏中，亚油酸(LA，C18：2，α6)选择性地占据了HNF4的ω。在禁食状态和携带突变的受体中，受体占有率显著降低，该突变源自成熟期起病的青年糖尿病患者(MODY1)。然而，有趣的是，配体占有率似乎对HNF4HNF4DNA转录活性没有显著影响，这一点从人类结肠来源细胞的全基因组表达谱中得到了证明。我们还使用AIMS来表明LA结合在完整细胞中是可逆的，表明HNF4α可能是一个可行的药物靶点。这项研究建立了一种通用的方法来鉴定核受体(和其他脂结合蛋白)的真正内源性配体，而不依赖于转录功能，并追踪生理相关条件下体内受体的占据情况。
Orphan nuclear receptors have been instrumental in identifying novel signaling pathways and therapeutic targets. However, identification of ligands for these receptors has often been based on random compound screens or other biased approaches. As a result, it remains unclear in many cases if the reported ligands are the true endogenous ligands, – i.e., the ligand that is bound to the receptor in an unperturbed in vivo setting. Technical limitations have limited our ability to identify ligands based on this rigorous definition. The orphan receptor hepatocyte nuclear factor 4 α (HNF4α) is a key regulator of many metabolic pathways and linked to several diseases including diabetes, atherosclerosis, hemophilia and cancer. Here we utilize an affinity isolation/mass-spectrometry (AIMS) approach to demonstrate that HNF4α is selectively occupied by linoleic acid (LA, C18:2ω6) in mammalian cells and in the liver of fed mice. Receptor occupancy is dramatically reduced in the fasted state and in a receptor carrying a mutation derived from patients with Maturity Onset Diabetes of the Young 1 (MODY1). Interestingly, however, ligand occupancy does not appear to have a significant effect on HNF4α transcriptional activity, as evidenced by genome-wide expression profiling in cells derived from human colon. We also use AIMS to show that LA binding is reversible in intact cells, indicating that HNF4α could be a viable drug target. This study establishes a general method to identify true endogenous ligands for nuclear receptors (and other lipid binding proteins), independent of transcriptional function, and to track in vivo receptor occupancy under physiologically relevant conditions.
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影响因子：
4.8
作者：
Dhe-Paganon, S;Duda, K;Shoelson, SE
通讯作者：
Shoelson, SE
影响因子：
5.3
作者：
Hayhurst, GP;Lee, YH;Gonzalez, FJ
通讯作者：
Gonzalez, FJ
影响因子：
10.5
作者：
Gupta, Rana K.;Gao, Nan;Kaestner, Klaus H.
通讯作者：
Kaestner, Klaus H.
影响因子：
--
作者：
DuSell, Carolyn D.;Umetani, Michihisa;McDonnell, Donald P.
通讯作者：
McDonnell, Donald P.
DOI：
10.1073/pnas.94.13.6803
发表时间：
1997-06-24
影响因子：
11.1
作者：
Escriva, H;Safi, R;Laudet, V
通讯作者：
Laudet, V