Fig. 4: Dynamic interplay between 5′-end counting and motif-driven cleavage rules.
From: DICER cleavage fidelity is governed by 5′-end binding pockets
a, Structural comparison of the DICER–26S-UG and DICER–26S-GU complexes. RMSD heat maps show minor structural differences in the 5′ binding pocket, PAZ domain and RIIIDa near the 3′ cleavage site. b, Alignment of the 26S-UG and 26S-GU RNA structures with their corresponding structures from AlphaFold3 (AF3)-predicted models. c, RNA flexibility and DICER’s dsRBD interactions: the 26S-GU complex exhibits distortion of the RNA backbone, aligning DC22 with the catalytic centre. d, Pre-dicing state of DICER bound to pre-mir-517a_GU. Cryo-EM density shows the PAZ domain (gold) engaging the duplex end and the dsRBD (gold) positioned along the RNA duplex (green), with other domains in grey. e, Dicing state. The duplex (green) docks into RIIIDa and RIIIDb (RIIIDa/b) as the PAZ domain and dsRBD (red) reposition to clamp the RNA for cleavage. f, Model of domain motion. Arrows indicate the movements of the dsRBD and the PAZ domain during activation. g, Density view of the end of the RNA duplex, showing G1 and opposing U59. h, The RNA duplex 5′-end occupies the DC21 pocket; the boundary loop (red) lies between DC21 and DC22. Open circles indicate pocket positions. i, Close-up view of the 5′-end region, showing G1 and the 5′-phosphate (PO4) coordinated near residues D991 and H992. j, RNA helical geometry comparison, highlighting the duplex distortions in 26S-GU compared with AlphaFold3 and experimental models of pre-mir-517a_GU. k, Proposed mechanism. 5′-U anchor: the 5′-U pocket aligns the mWCU–YCR motif at DC22 for cleavage. 5′-G anchor: the 5′-G pocket favours DC21 cleavage, misaligned with the mWCU–YCR motif at DC22. Thus, motif-driven RNA conformational changes override the 5′-G rule, distort the RNA backbone and ensure DC22 cleavage.
