TY - JOUR
T1 - Real-time analysis of single influenza virus replication complexes reveals large promoter-dependent differences in initiation dynamics
AU - Robb, Nicole C.
AU - te Velthuis, Aartjan J.W.
AU - Fodor, Ervin
AU - Kapanidis, Achillefs N.
N1 - Funding Information:
Royal Society Dorothy Hodgkin Research Fellowship [DKR00620 to N.C.R.]; joint Wellcome Trust and Royal Society grant [206579/Z/17/Z]; Marmaduke Shield Fund award and Isaac Newton Trust grant [17.37(r) to A.t.V.]; Medical Research Council grants [MR/K000241/1 and MR/R009945/1 to E.F. and MR/N010744/1 to A.N.K and E.F.]; Wellcome Trust grant [110164/Z/15/Z to A.N.K.]. Funding for open access charge: Oxford’s RCUK Open Access Block Grant. Conflict of interest statement. None declared.
Publisher Copyright:
© The Author(s) 2019
PY - 2019/7/9
Y1 - 2019/7/9
N2 - The viral RNA (vRNA) genome of influenza viruses is replicated by the RNA-dependent RNA polymerase (RNAP) via a complementary RNA (cRNA) intermediate. The vRNA promoter can adopt multiple conformations when bound by the RNAP. However, the dynamics, determinants, and biological role of these conformations are unknown; further, little is known about cRNA promoter conformations. To probe the RNA conformations adopted during initial replication, we monitored single, surface-immobilized vRNA and cRNA initiation complexes in real-time. Our results show that, while the 3' terminus of the vRNA promoter exists in dynamic equilibrium between pre-initiation and initiation conformations, the cRNA promoter exhibited very limited dynamics. Two residues in the proximal 3' region of the cRNA promoter (residues absent in the vRNA promoter) allowed the cRNA template strand to reach further into the active site, limiting promoter dynamics. Our results highlight promoter-dependent differences in influenza initiation mechanisms, and advance our understanding of virus replication.
AB - The viral RNA (vRNA) genome of influenza viruses is replicated by the RNA-dependent RNA polymerase (RNAP) via a complementary RNA (cRNA) intermediate. The vRNA promoter can adopt multiple conformations when bound by the RNAP. However, the dynamics, determinants, and biological role of these conformations are unknown; further, little is known about cRNA promoter conformations. To probe the RNA conformations adopted during initial replication, we monitored single, surface-immobilized vRNA and cRNA initiation complexes in real-time. Our results show that, while the 3' terminus of the vRNA promoter exists in dynamic equilibrium between pre-initiation and initiation conformations, the cRNA promoter exhibited very limited dynamics. Two residues in the proximal 3' region of the cRNA promoter (residues absent in the vRNA promoter) allowed the cRNA template strand to reach further into the active site, limiting promoter dynamics. Our results highlight promoter-dependent differences in influenza initiation mechanisms, and advance our understanding of virus replication.
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U2 - 10.1093/NAR/GKZ313
DO - 10.1093/NAR/GKZ313
M3 - Article
C2 - 31032520
AN - SCOPUS:85069294819
SN - 0305-1048
VL - 47
SP - 6466
EP - 6477
JO - Nucleic acids research
JF - Nucleic acids research
IS - 12
ER -