Shiftless Restricts Viral Gene Expression and Influences RNA Granule Formation during KSHV lytic replication

Abstract

Herpesviral infection reflects thousands of years of co-evolution and the constant struggle between virus and host for control of cellular gene expression. During Kaposi's sarcoma-associated herpesvirus (KSHV) lytic replication, the virus rapidly seizes control of host gene expression machinery by triggering a massive RNA decay event via a virally-encoded endoribonuclease, SOX. This virus takeover strategy decimates close to 80% of cellular transcripts, reallocating host resources toward viral replication. The host cell, however, is not entirely passive in this assault on RNA stability. A small pool of host transcripts that actively evade SOX cleavage has been identified over the years. One such "escapee", C19ORF66 (herein referred to as Shiftless - SHFL) encodes a potent anti-viral protein capable of restricting the replication of multiple DNA, RNA, and retroviruses including KSHV. Here, we show that SHFL restricts KSHV replication by targeting the expression of critical viral early genes, including the master transactivator protein, KSHV ORF50, and thus subsequently the entire lytic gene cascade. Consistent with previous reports, we found the SHFL interactome throughout KSHV infection is dominated by RNA-binding proteins that influence both translation and protein stability, including the viral protein ORF57, a crucial regulator of viral RNA fate. We next show that SHFL affects cytoplasmic RNA granule formation, triggering the disassembly of processing bodies. Taken together, our findings provide insights into the complex relationship between RNA stability, RNA granule formation, and the anti-viral response to KSHV infection.

Restriction of Flaviviruses by an Interferon-Stimulated Gene SHFL/C19orf66

“…In this review, we focused only on the inhibitory activity of SHFL against flaviviruses, as the antiviral function of SHFL was first demonstrated in studies of Flaviviridae family viruses (i.e., HCV and DENV studies [ 14 , 15 , 22 ]). However, as Rodriguez and Muller well summarized in their review [ 7 ], the virus inhibitory activity of SHFL is not limited to flaviviruses but extends to many other types of RNA and DNA viruses, and interestingly, various inhibitory mechanisms of action have been proposed for SHFL’s antiviral activities [ 15 , 18 , 22 , 23 , 24 , 26 , 27 , 51 , 52 , 53 , 54 , 55 , 56 ]. Thus, questions remain: (i) Are the different mechanisms seen in the inhibition of various types of viruses the different consequences of one molecular function of SHFL?…”

Shiftless, a Critical Piece of the Innate Immune Response to Viral Infection

“…In nearly every context, this protein was identified as a broad-spectrum anti-viral factor whose expression is upregulated in response to Type I interferon (IFN) induction and generally viral infection. From these studies, several unique identifiers have been attributed to the C19orf66 gene product including FLJ11286 (genomic location), Repressor of Yield of Dengue Virus (RyDEN), Suppressor of Viral Activity-1 (SVA-1), Interferon Regulated Anti-Viral gene (IRAV), and more recently Shiftless (SFL/SHFL) with SHFL finally approved for its official gene symbol [10][11][12][13][14][15][16][17][18][19][20]. SHFL expression can often vary dramatically between cell types as demonstrated by Balinsky and colleagues [11].…”

“…One pattern that emerges among studies of the SHFL interactome is that it is consistently dominated by constituents of phase-separated RNA granules [10,11,14,20]. RNAgranules are membrane-free, phase-separated ribonucleoprotein (RNP) complexes that function in the storage, translational arrest, and/or degradation of RNA throughout the cell [55][56][57][58].…”

“…Recently, our group has investigated the mechanism by which SHFL restricts lytic viral gene expression [20] and explored the interactome of SHFL throughout KSHV infection. In line with previous reports, we have found a number of RNA binding proteins interacting with SHFL and we are now in process of further characterizing the relationship between SHFL and RNA granule formation during herpesviral infection.…”

“…An increasing number of large-scale screens have independently identified SHFL as both an anti-viral factor and downstream effector of the Type I, II, and III interferon response to viral infection [2][3][4][5][6][7][8][9]. Some studies have gone even further to characterize SHFL as a potent, broad-spectrum restriction factor for a growing list of DNA, RNA, and Retroviruses (Summarized in Table 1) [8,[10][11][12][13][14][15][16][17][18][19][20]. Many ISGs have evolved to target viral gene expression as it is often the primary battleground that nearly every viral replication steps depend upon.…”