Regulating Protein-RNA Interactions: Advances in Targeting the LIN28/Let-7 Pathway

Abstract

Originally discovered in C. elegans, LIN28 is an evolutionarily conserved zinc finger RNA-binding protein (RBP) that post-transcriptionally regulates genes involved in developmental timing, stem cell programming, and oncogenesis. LIN28 acts via two distinct mechanisms. It blocks the biogenesis of the lethal-7 (let-7) microRNA (miRNA) family, and also directly binds messenger RNA (mRNA) targets, such as IGF-2 mRNA, and alters downstream splicing and translation events. This review focuses on the molecular mechanism of LIN28 repression of let-7 and current strategies to overcome this blockade for the purpose of cancer therapy. We highlight the value of the LIN28/let-7 pathway as a drug target, as multiple oncogenic proteins that the pathway regulates are considered undruggable due to their inaccessible cellular location and lack of cavities for small molecule binding.

Keywords: LIN28; gene therapy; let-7; miRNA; oncogene; small-molecule inhibitor.

Figure 1

LIN28 Inhibits let-7 miRNA Biogenesis and Contributes to Cancer. In the nucleus, human LIN28B sequesters pri-let-7 and as a result, the Drosha-DGCR8 microprocessor fails to generate pre-let-7, thereby arresting let-7 biogenesis. LIN28A (CSD—cold-shock domain; ZKD—zinc knuckle domain) binds to pre-let-7 after its nuclear export. Given the cytoplasmic localization of LIN28B in some cell types, LIN28B inhibition of mature let-7 biogenesis can also occur in the cytoplasm. LIN28A/B recruits TUT4/Zcchc11, which oligouridylates pre-let-7 at its 3′ end. Oligouridylated pre-let-7 then undergoes exonuclease degradation by DIS3L2. LIN28A/B binding to pre-let-7 could interfere with Dicer cleavage, providing the mechanism for TUTase-independent repression. Without this inhibition, pre-let-7 undergoes processing by Dicer to yield mature let-7 miRNA, which constitutes part of RISC and inhibits multiple oncogenes through RNAi.

Figure 2

Involvement of LIN28A CSD and ZKD in let-7 pre-miRNA binding and processing. Using their CSD and ZKD binding elements, let-7 miRNA precursor recruits LIN28A, resulting to their 3ʹuridylation by TUTase and eventual degradation by DIS3L2.

Figure 3

Some effects of the repression of let-7 by LIN28 on cancer cells.

Figure 4

Pharmacological inhibition of LIN28-driven oncogenesis. (A) Domain architecture of human LIN28A and LIN28B, which share an N-terminal CSD (magenta) and a C-terminal ZKD (green) The residue numbers are denoted atop. (B) Let-7 mimics enhance tumor suppression by replenishing let-7, the level of which is decreased by LIN28-mediated oligouridylation and degradation by nuclease DIS3L2 (grey) (PDB ID: 4PMW). (C) Looptomirs hinder (red cross) LIN28 binding to pre-let-7 without blocking downstream pre-let-7 processing by Dicer, thereby promoting let-7 maturation. (D) Small molecules LI71 and TPEN inhibit (red cross) LIN28 by targeting its CSD and ZKD, respectively. Crystal structure of human LIN28A in complex with pre-let-7f-1 (yellow) (PDB ID: 5UDZ). Zn²⁺ ions (blue) are visible near the ZKD. The GGAG motif in pre-let-7f-1 is indicated near its 3′ end.