Evolutionary trace-based peptides identify a novel asymmetric interaction that mediates oligomerization in nuclear receptors
Evolutionary trace-based peptides identify a novel asymmetric interaction that mediates oligomerization in nuclear receptors
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DOI：
10.1074/jbc.m501924200
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发表时间：
2005-09-09
影响因子：
4.8
通讯作者：
Cooney, AJ
中科院分区：
文献类型：
--
作者：
Gu, PL;Morgan, DH;Cooney, AJ
生殖细胞核因子 (GCNF) 是一种孤儿核受体，通过与 DR0 元件结合来抑制 Oct4、GDF9 和 BMP15 等必需基因的表达，在发育和生殖中发挥重要作用。令人惊讶的是，虽然重组 GCNF 作为同二聚体与 DR0 序列结合，但内源 GCNF 并不作为同二聚体存在，而是作为称为瞬时类视黄醇诱导因子 (TRIF) 的大型复合物的一部分存在。在这里，我们使用进化追踪（ET）分析来设计突变和肽，以探究这种不寻常复合物形成的分子基础。我们发现 GCNF 同二聚化和 TRIF 复合物形成是 DNA 依赖性的，ET 表明二聚化涉及螺旋 3 和螺旋 11 上的关键功能位点，这些位点位于配体结合域的相对表面。 GCNF 任一螺旋中的靶向突变都会破坏同源二聚体和内源 TRIF 复合物的形成。此外，这两个 ET 确定位点的肽模拟物抑制二聚化和 TRIF 复合物形成。这表明一种新的螺旋 3-螺旋 11 异型相互作用介导 GCNF 相互作用并促进寡聚化。事实上，已确定内源TRIF复合物由GCNF寡聚物组成。这些发现揭示了一种进化选择机制，揭示了 GCNF 不寻常的 DNA 结合、二聚化和寡聚化特性。
Germ cell nuclear factor ( GCNF) is an orphan nuclear receptor that plays important roles in development and reproduction, by repressing the expression of essential genes such as Oct4, GDF9, and BMP15, through binding to DR0 elements. Surprisingly, whereas recombinant GCNF binds to DR0 sequences as a homodimer, endogenous GCNF does not exist as a homodimer but rather as part of a large complex termed the transiently retinoid-induced factor ( TRIF). Here, we use evolutionary trace ( ET) analysis to design mutations and peptides that probe the molecular basis for the formation of this unusual complex. We find that GCNF homodimerization and TRIF complex formation are DNA-dependent, and ET suggests that dimerization involves key functional sites on both helix 3 and helix 11, which are located on opposing surfaces of the ligand binding domain. Targeted mutations in either helix of GCNF disrupt the formation of both the homodimer and the endogenous TRIF complex. Moreover, peptide mimetics of both of these ET-determined sites inhibit dimerization and TRIF complex formation. This suggests that a novel helix 3-helix 11 heterotypic interaction mediates GCNF interaction and would facilitate oligomerization. Indeed, it was determined that the endogenous TRIF complex is composed of a GCNF oligomer. These findings shed light on an evolutionarily selected mechanism that reveals the unusual DNA-binding, dimerization, and oligomerization properties of GCNF.