Influenza Virus NS1 Protein Interacts with the Cellular 30 kDa Subunit of CPSF and Inhibits 3′ End Formation of Cellular Pre-mRNAs

Abstract

Inhibition of the nuclear export of poly(A)-containing mRNAs caused by the influenza A virus NS1 protein requires its effector domain. Here, we demonstrate that the NS1 effector domain functionally interacts with the cellular 30 kDa subunit of CPSF, an essential component of the 3' end processing machinery of cellular pre-mRNAs. In influenza virus-infected cells, the NS1 protein is physically associated with CPSF 30 kDa. Binding of the NS1 protein to the 30 kDa protein in vitro prevents CPSF binding to the RNA substrate and inhibits 3' end cleavage and polyadenylation of host pre-mRNAs. The NS1 protein also inhibits 3' end processing in vivo, and the uncleaved pre-mRNA remains in the nucleus. Via this novel regulation of pre-mRNA 3' end processing, the NS1 protein selectively inhibits the nuclear export of cellular, and not viral, mRNAs.

Protein factors in pre-mRNA 3′-end processing

“…ZF1 and ZF4 of Yth1p interact with the N-terminal region of Brr5p/Ysh1p [125]. Influenza virus attenuates host antiviral response by blocking the function of CPSF-30 with its NS1 protein [127][128][129].…”

Large-scale analysis of influenza A virus sequences reveals potential drug target sites of non-structural proteins

“…The major role of NS1 is to antagonize the antiviral response of the host by preventing the activation of NF-kB and induction of alpha/betainterferon (IFN-a/b) (Wang et al, 2000). However, NS1 is a multifunctional protein that is additionally involved in several processes: (i) inhibiting the pre-mRNA 39-end processing (Fortes et al, 1994) by binding to two 39-end processing factors, namely cleavage and polyadenylation specificity factor and poly(A)-binding protein II (Chen et al, 1999;Nemeroff et al, 1998); (ii) blocking the post-transcriptional processing and nuclear export of cellular mRNA (Fortes et al, 1994); (iii) stimulating the translation of matrix (M1) proteins (Enami et al, 1994;Marió n et al, 1997); (iv) inhibiting the activation of a protein kinase that phosphorylates the elf-2 translation initiation factor by binding to doublestranded (ds)RNA (Lu et al, 1995;Aragó n et al, 2000); (v) induction of the phosphatidylinositol-3-kinase (PI3K)/Akt signalling pathway in order to support virus replication (Ehrhardt et al, 2006(Ehrhardt et al, , 2007. The NS1 protein is 230-237 aa, depending on the strain, and has a molecular mass of approximately 26 kDa (reviewed by Hale et al, 2008b).…”

SUMOylation Affects the Interferon Blocking Activity of the Influenza A Nonstructural Protein NS1 without Affecting Its Stability or Cellular Localization

“…Out of the 10 to 11 viral proteins encoded by influenza A virus, one of the most functionally diverse is the nonstructural viral protein NS1, which has been associated with numerous roles during influenza infection, including the modulation of viral RNA (vRNA) replication (3)(4)(5)(6), general inhibition of the nuclear export of mRNAs carrying polyadenylated tails (7,8), inhibition of the transcriptional elongation of host genes (9), regulation of host and viral protein synthesis (recently reviewed by Yanguez and Nieto [10]), and the neutralization of the activity of some of the interferon (IFN)-induced antiviral proteins, such as 2=,5=-oligoadenylate synthetase (OAS)/RNase L (11) and the double-stranded RNA (dsRNA)-dependent protein kinase R (PKR) (12)(13)(14)(15)(16)(17)(18). However, the main function attributed to NS1 is the neutralization of the initial signaling pathway leading to the production of type I IFN (reviewed by Krug et al and Hale et al [19,20]).…”

“…Whereas the latter function is heavily dependent on NS1's RNA binding properties (21), the ability of this protein to interact with cellular and viral proteins also contributes to its IFN-blocking activity and constitutes the main determinant of its other numerous functions (19). For example, NS1's interactions with both the 30-kDa subunit of the cleavage and polyadenylation specificity factor protein (CPSF30) and polyadenylate binding protein 2 (PABP2) are thought to mediate NS1's ability to decrease the processing and maturation of cellular mRNAs, therefore leading to a substantial decrease in host protein synthesis (7,22). Similarly, NS1's ability to inhibit the activation of the viral RNA sensor RIG-I is mediated by its ability to bind the tripartite motif protein TRIM25, a RING domain ubiquitin E3 ligase, therefore blocking its multimerization and in turn inhibiting its ability to ubiquitinate the CARD motif in RIG-I (23), a requirement to allow RIG-I interaction with its downstream effector MAVS/VISA/IPS-1/Cardif (24).…”