Membrane association and activity of rough endoplasmic reticulum associated RISC requires farnesylated chaperones. Arabidopsis thaliana strain:Col-0

ARGONAUTE (AGO) proteins affect small RNA-guided gene silencing. Arabidopsis AGO1 mediates microRNA (miRNA) function, and, similar to its animal orthologues, AGO1 requires heat shock proteins for miRNA loading and carries out some silencing functions bound to the rough endoplasmic reticulum (rER). The mevalonate pathway is required for miRNA activity, but precise mechanisms underlying this requirement for isoprenoid metabolites are not known. Here, we show that farnesylation of two Arabidopsis Hsp40 chaperones, J2 and J3, is required for silencing activity of several miRNA and siRNAs. J2/J3 associate with AGO1 in membrane fractions, and their farnesylation is implicated in membrane association of AGO1. Surprisingly, membrane-bound AGO1 is loaded in farnesylation-defective Hsp40 mutants, providing genetic evidence that miRNA activity requires chaperone function after formation of AGO-miRNA complexes.

ARGONAUTE（AGO）蛋白参与调控小RNA介导的基因沉默过程。拟南芥AGO1介导微小RNA（microRNA，miRNA）的功能发挥，且与动物同源物类似，AGO1需要热激蛋白（heat shock proteins）参与miRNA的装载过程，并可结合于粗面内质网（rough endoplasmic reticulum，rER）上执行部分基因沉默功能。甲羟戊酸途径对miRNA的活性不可或缺，但这类类异戊二烯代谢物发挥作用的具体分子机制仍不明晰。本研究发现，拟南芥的两种Hsp40分子伴侣（Hsp40 chaperones）J2与J3的法尼基化修饰，是多种miRNA与小干扰RNA（siRNAs）发挥沉默活性所必需的。J2与J3可在膜组分中与AGO1结合，且它们的法尼基化修饰参与AGO1的膜定位过程。令人意外的是，在法尼基化缺陷的Hsp40突变体中，结合于膜上的AGO1仍可完成miRNA的装载，这为“miRNA活性需要在AGO-miRNA复合物形成后依赖分子伴侣功能”这一结论提供了遗传学证据。