Belak Zachery R, Ovsenek Nick
Department of Anatomy and Cell Biology, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
J Biol Chem. 2007 Dec 28;282(52):37913-20. doi: 10.1074/jbc.M708057200. Epub 2007 Nov 1.
The early stages of vertebrate development depend heavily on control of maternally transcribed mRNAs that are stored for long periods in complexes termed messenger ribonucleoprotein particles (mRNPs) and utilized selectively following maturation and fertilization. The transcription factor Yin Yang 1 (YY1) is associated with cytoplasmic mRNPs in vertebrate oocytes; however, the mechanism by which any of the mRNP proteins associate with mRNA in the oocyte is unknown. Here we demonstrate the mechanism by which YY1 associates with mRNPs depends on its direct RNA binding activity. High affinity binding for U-rich single-stranded RNA and A:U RNA duplexes was observed in the nanomolar range, similar to the affinity for the cognate double-stranded DNA-binding element. Similar RNA binding affinity was observed with endogenous YY1 isolated from native mRNP complexes. In vivo expression experiments reveal epitope-tagged YY1 assembled into high molecular mass mRNPs, and assembly was blocked by microinjection of high affinity RNA substrate competitor. These findings present the first clues to how mRNPs assemble during early development.
脊椎动物发育的早期阶段在很大程度上依赖于对母源转录mRNA的控制,这些mRNA长期储存在称为信使核糖核蛋白颗粒(mRNP)的复合物中,并在成熟和受精后被选择性利用。转录因子阴阳1(YY1)与脊椎动物卵母细胞中的细胞质mRNP相关;然而,卵母细胞中任何mRNP蛋白与mRNA结合的机制尚不清楚。在这里,我们证明了YY1与mRNP结合的机制取决于其直接的RNA结合活性。在纳摩尔范围内观察到对富含U的单链RNA和A:U RNA双链体的高亲和力结合,类似于对同源双链DNA结合元件的亲和力。从天然mRNP复合物中分离出的内源性YY1也观察到类似的RNA结合亲和力。体内表达实验表明,表位标记的YY1组装成高分子量的mRNP,并且通过显微注射高亲和力RNA底物竞争者可阻断组装。这些发现为早期发育过程中mRNP如何组装提供了首个线索。