Critical contribution of 3’ non-seed base pairing to the in vivo function of the evolutionarily conserved let-7a microRNA

Base-pairing of the seed region (g2-g8) is essential for microRNA targeting, however, the in vivo functions of the 3’ non-seed region (g9-g22) are less well understood. Here we report the first systematic investigation of the in vivo roles of 3’ non-seed nucleotides in microRNA let-7a, whose entire g9-g22 region is conserved among bilaterians. We found that the 3’ non-seed sequence functionally distinguishes let-7a from its family paralogs. The complete pairing of g11-g16 is essential for let-7a to fully repress multiple key targets, including evolutionarily conserved lin-41, daf-12 and hbl-1. Nucleotides at g17-g22 are less critical but may compensate for mismatches in the g11-g16 region. Interestingly, the 3’ non-seed pairing of let-7a can be critically required even with sites that permit perfect seed pairing. These results provide evidence that the specific configurations of both seed and 3’ non-seed base-pairing can critically influence microRNA-mediated gene regulation in vivo.

种子区域（seed region，g2-g8）的碱基配对（base-pairing）对于微小RNA（microRNA）的靶向调控至关重要，但目前对其3'非种子区域（3' non-seed region，g9-g22）的体内（in vivo）功能仍知之甚少。本研究首次对微小RNA let-7a的3'非种子核苷酸的体内功能开展系统性探究，该微小RNA的完整g9-g22区域在两侧对称动物（bilaterians）中高度保守。研究发现，3'非种子序列可在功能上区分let-7a与其家族旁系同源基因（paralogs）。g11-g16区域的完全碱基配对是let-7a完全抑制多个关键靶基因的必要条件，这些靶基因包括进化保守的lin-41、daf-12与hbl-1。g17-g22区域的核苷酸相对不那么关键，但可弥补g11-g16区域存在的碱基错配。值得注意的是，即便靶位点存在完美的种子区域配对，let-7a的3'非种子碱基配对仍可能发挥关键作用。本研究结果证实，种子区域与3'非种子区域的碱基配对的特定构型均可显著影响微小RNA介导的体内基因调控过程。