Structural insights into the dual activities of the two-barrel RNA polymerase QDE-1

Author:

Cui Ruixue1,Li Hao1,Zhao Jin1,Li Xuhang1,Gan Jianhua2ORCID,Ma Jinbiao1

Affiliation:

1. Huashan Hospital affiliated to Fudan University, State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Multiscale Research Institute of Complex Systems, Department of Biochemistry and Biophysics, School of Life Sciences, Fudan University , Shanghai 200438, China

2. Shanghai Public Health Clinical Center, State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry and Biophysics, School of Life Sciences, Fudan University , Shanghai 200438, China

Abstract

Abstract Neurospora crassa protein QDE-1, a member of the two-barrel polymerase superfamily, possesses both DNA- and RNA-dependent RNA polymerase (DdRP and RdRP) activities. The dual activities are essential for the production of double-stranded RNAs (dsRNAs), the precursors of small interfering RNAs (siRNAs) in N. crassa. Here, we report five complex structures of N-terminal truncated QDE-1 (QDE-1ΔN), representing four different reaction states: DNA/RNA-templated elongation, the de novo initiation of RNA synthesis, the first step of nucleotide condensation during de novo initiation and initial NTP loading. The template strand is aligned by a bridge-helix and double-psi beta-barrels 2 (DPBB2), the RNA product is held by DPBB1 and the slab domain. The DNA template unpairs with the RNA product at position –7, but the RNA template remains paired. The NTP analog coordinates with cations and is precisely positioned at the addition site by a rigid trigger loop and a proline-containing loop in the active center. The unique C-terminal tail from the QDE-1 dimer partner inserts into the substrate-binding cleft and plays regulatory roles in RNA synthesis. Collectively, this work elucidates the conserved mechanisms for DNA/RNA-dependent dual activities by QDE-1 and other two-barrel polymerase superfamily members.

Funder

National Natural Science Foundation of China

Publisher

Oxford University Press (OUP)

Subject

Genetics

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