title: Recognition of cap structure by influenza B virus RNA polymerase is less dependent on the methyl residue than recognition by influenza A virus polymerase.
abstract: The cap-dependent endonuclease activity of the influenza virus RNA-dependent RNA polymerase cleaves host mRNAs to produce capped RNA fragments for primers to initiate viral mRNA synthesis. The influenza A virus (FluA) cap-dependent endonuclease preferentially recognizes the cap1 structure (m(7)GpppNm). However, little is known about the substrate specificity of the influenza B virus (FluB) endonuclease. Here, we determined the substrate specificity of the FluB polymerase using purified viral RNPs and (32)P-labeled polyribonucleotides containing a variety of cap structures (m(7)GpppGm, m(7)GpppG, and GpppG). We found that the FluA polymerase cleaves m(7)G-capped RNAs preferentially. In contrast, the FluB polymerase could efficiently cleave not only m(7)G-capped RNAs but also unmethylated GpppG-RNAs. To identify a key amino acid(s) related to the cap recognition specificity of the PB2 subunit, the transcription activity of FluB polymerases containing mutated cap-binding domains was examined by use of a minireplicon assay system. In the case of FluA PB2, Phe323, His357, and Phe404, which stack the m(7)GTP, and Glu361 and Lys376, which make hydrogen bonds with a guanine base, were essential for the transcription activity. In contrast, in the case of FluB PB2, the stacking interaction of Trp359 with a guanine base and putative hydrogen bonds using Gln325 and Glu363 were enough for the transcription activity. Taking these results together with the result for the cap-binding activity, we propose that the cap recognition pocket of FluB PB2 does not have the specificity for m(7)G-cap structures and thus is more flexible to accept various cap structures than FluA PB2.
authors: Department of Infection Biology, Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575, Japan.
authors: Chitose
authors: C
authors: Wakai
authors: Minako
authors: M
authors: Iwama
authors: Kiyohisa
authors: K
authors: Mizumoto
authors: Kyosuke
authors: K
authors: Nagata
references: Methods Enzymol. 1996;275:90-9
references: 9026662
references: Proc Natl Acad Sci U S A. 1999 Jun 22;96(13):7149-54
references: 10377383
references: Nat Struct Mol Biol. 2008 May;15(5):500-6
references: 18454157
references: J Mol Biol. 2002 Jun 7;319(3):615-35
references: 12054859
references: EMBO J. 2002 Oct 15;21(20):5548-57
references: 12374755
references: Proc Natl Acad Sci U S A. 1980 Jul;77(7):3952-6
references: 6933444
references: Proc Natl Acad Sci U S A. 1978 Oct;75(10):4886-90
references: 283399
references: Nature. 2008 Aug 28;454(7208):1127-31
references: 18660801
references: J Virol. 2006 Aug;80(16):7789-98
references: 16873236
references: Nucleic Acids Res. 2004 Jan 29;32(2):643-52
references: 14752052
references: Biochem J. 2002 Mar 15;362(Pt 3):539-44
references: 11879179
references: Cell. 1983 Sep;34(2):609-18
references: 6616622
references: J Virol. 1998 Jul;72(7):5493-501
references: 9621005
references: EMBO J. 2009 Jun 17;28(12):1803-11
references: 19461581
references: J Virol. 2005 Jan;79(2):732-44
references: 15613301
references: Nat Struct Biol. 1997 May;4(5):350-4
references: 9145102
references: Rev Med Virol. 2006 Sep-Oct;16(5):329-45
references: 16933365
references: Nature. 2009 Apr 16;458(7240):914-8
references: 19194459
references: J Mol Biol. 2003 Apr 25;328(2):365-83
references: 12691746
references: J Virol. 2008 Dec;82(23):11880-8
references: 18815312
references: Proc Natl Acad Sci U S A. 1999 Aug 3;96(16):9345-50
references: 10430945
references: J Virol. 1978 Oct;28(1):75-83
references: 702657
references: Virus Res. 2008 Jul;135(1):161-5
references: 18455827
references: Nucleic Acids Res. 1994 Nov 25;22(23):5047-53
references: 7800498
references: J Virol. 1979 Dec;32(3):895-904
references: 513205
references: Nature. 2008 Aug 28;454(7208):1123-6
references: 18615018
references: Nature. 2009 Apr 16;458(7240):909-13
references: 19194458
references: Nucleic Acids Res. 1988 Nov 11;16(21):9909-16
references: 2461550
references: Virology. 1994 Jul;202(1):477-9
references: 8009859
references: Virology. 2007 Jun 5;362(2):271-82
references: 17270230
references: J Virol. 2002 Sep;76(18):8989-9001
references: 12186883
references: J Cell Biol. 1992 Sep;118(6):1287-95
references: 1522107
references: J Biol Chem. 2005 Feb 11;280(6):4429-35
references: 15574411
references: J Virol. 1977 Sep;23(3):816-9
references: 561196
references: Vaccine. 1998 Oct;16(16):1544-9
references: 9711802