Penaeus monodon TATA Box-Binding Protein Interacts with the White Spot Syndrome Virus Transactivator IE1 and Promotes Its Transcriptional Activity

Abstract

We show here that the white spot syndrome virus (WSSV) immediate-early protein IE1 interacts with the Penaeus monodon TATA box-binding protein (PmTBP) and that this protein-protein interaction occurs in the absence of any other viral or cellular proteins or nucleic acids, both in vitro and in vivo. Mapping studies using enhanced green fluorescent protein (EGFP) fusion proteins containing truncations of IE1 and PmTBP delimited the interacting regions to amino acids (aa) 81 to 180 in IE1 and, except for aa 171 to 230, to aa 111 to 300 in PmTBP. A WSSV IE1 transactivation assay showed that large quantities (>800 ng) of the GAL4-IE1 plasmid caused "squelching" of the GAL4-IE1 activity and that this squelching effect was alleviated by the overexpression of PmTBP. Gene silencing of WSSV ie1 and PmTBP by pretreatment with double-stranded RNAs (dsRNAs) prior to WSSV challenge showed that the expression of these two target genes was specifically inhibited by their corresponding dsRNAs 72 and 96 h after dsRNA treatment. dsRNA silencing of ie1 and PmTBP expression also significantly reduced WSSV replication and the expression of the viral early gene dnapol (DNA polymerase gene). These results suggest that WSSV IE1 and PmTBP work cooperatively with each other during transcription initiation and, furthermore, that PmTBP is an important target for WSSV IE1's transactivation activity that can enhance viral gene expression and help in virus replication.White spot syndrome virus (WSSV), the type species of the genus Whispovirus, family Nimaviridae (42), has a wide host range and is a lethal agent infecting penaeid shrimp (35). WSSV has spread globally and has caused huge economic losses to the shrimp farming industry (10,27,28,35). WSSV is a large double-stranded DNA (dsDNA) virus (43), and although the complete sequence of the WSSV genome has been known for several years (7,40,45), knowledge of the biological functions of the viral proteins is still quite poor. During infection by large DNA viruses, such as baculoviruses and herpesviruses, gene expression is regulated such that the immediate-early (IE) genes are transcribed first, followed by the expression of the early and late genes (1,2,11,14). To date, 18 WSSV IE genes have been identified (20,25). Although the functions of most of the corresponding WSSV IE proteins have not yet been studied, many viral IE genes are known to encode multifunctional transcriptional regulators that both positively and negatively modulate viral early and late gene expression (17,37,44). Once expressed, the IE gene products may then function as regulatory trans-acting factors and may serve to initiate viral replicative events during infection. In the cascade of viral regulatory events, successive stages of virus replication are dependent on the proper expression of the genes in the preceding stage. Thus, viral IE genes are critically important in the virus infection cycle.Upon infection, expression of the WSSV genes can be divided into immediate-early, early, and late phases (25,29). However, th...

Molecular Mechanisms of White Spot Syndrome Virus Infection and Perspectives on Treatments

“…After expression of ie1 , WSSV IE1 protein (a viral transcription factor) is then able to facilitate expression of the viral early genes. WSSV IE1 has been shown to accomplish this through cooperation with P. monodon TATA box-binding proteins (PmTBP) in transcription initiation [ 116 ].…”

White Spot Syndrome Virus IE1 and WSV056 Modulate the G 1 /S Transition by Binding to the Host Retinoblastoma Protein

“…Based on detailed mapping of IE1 promoters, progress has been made in understanding the cellular regulation of IE1 expression. Several transcription factors have been found to affect IE transcription, including shrimp homologs of STAT (7,8), NF-B (9,10), and a TATA box binding protein (11). JAK-STAT and NF-B pathways are believed to be critical for antiviral defense (12,13); therefore, the findings for WSSV indicate that this virus takes advantage of the host immune system to assist its own proliferation.…”

“…JAK-STAT and NF-B pathways are believed to be critical for antiviral defense (12,13); therefore, the findings for WSSV indicate that this virus takes advantage of the host immune system to assist its own proliferation. The biological significance of IE1 during WSSV infection was demonstrated by RNA interference experiments which revealed that depletion of IE1 strongly inhibits the replication of WSSV (11). A recent report showed that the shrimp thioredoxin PmTrx can bind to IE1 and restore its DNAbinding activity under oxidizing conditions, indicating a role for IE1 in WSSV pathogenicity (14).…”

Penaeus monodonThioredoxin Restores the DNA Binding Activity of Oxidized White Spot Syndrome Virus IE1

“…The extracted lysate was centrifuged at 12,000 g for 10 min, and the supernatant was dried using a centrifugal evaporator (CVE-2000, EYELA). The pellet was suspended in a volume of deionized water that was equal WSSV copy number quantification WSSV copy number quantification was performed using the commercial real-time PCR kit for viral load quantification (GeneReach Biotechnology Corp.) as described previously (22). Results were presented as meansstandard deviation.…”