scholarly journals Papillomavirus Capsid Protein Expression Level Depends on the Match between Codon Usage and tRNA Availability

1999 ◽  
Vol 73 (6) ◽  
pp. 4972-4982 ◽  
Author(s):  
Jian Zhou ◽  
Wen Jun Liu ◽  
Shi Wen Peng ◽  
Xiao Yi Sun ◽  
Ian Frazer
Keyword(s):  
Gene Expression ◽  
Codon Usage ◽  
Fluorescent Protein ◽  
In Vitro Translation ◽  
Late Gene ◽  
Wild Type ◽  
Late Gene Expression ◽  
Late Genes ◽  
Codon Composition ◽  
L1 And L2

ABSTRACT Translation of mRNA encoding the L1 and L2 capsid proteins of papillomavirus (PV) is restricted in vivo to differentiated epithelial cells, although transcription of the L1 and L2 late genes occurs more widely. The codon composition of PV late genes is quite different from that of most mammalian genes. To test the possibility that PV late gene codon composition determines the efficiency of PV late gene expression in some cell types, synthetic bovine papillomavirus type 1 (BPV1) late genes were constructed with codon composition modified to resemble the typical mammalian gene. Expression of these genes from a strong promoter in Cos-1 cells was compared with expression of wild-type BPV1 late genes from the same promoter. Both unmodified and modified PV late genes were transcribed in Cos-1 cells, but only the codon-modified genes were translated. In vitro translation of wild-type but not synthetic BPV1 L1 mRNA was markedly enhanced by addition of aminoacyl-tRNAs. Codon composition thus limits BPV1 late gene translation in Cos-1 cells, and this limitation can be overcome by modification of the codon composition of the genes or by provision of excess tRNA. Replacement of codons in the green fluorescent protein (gfp) gene with those frequently used in PV late genes did not alter gfp transcription in Cos-1 cells but almost abolished translation, supporting the hypothesis that the observed differences in efficiency of translation of modified and unmodified PV capsid genes were related to codon usage rather than mRNA structure. As tRNA populations vary within and between tissues in the same eukaryotic organism, we speculate that matching of tRNA availability to codon usage may be one determinant of the restriction of expression of PV late genes to differentiated epithelium.

scholarly journals The Interaction between ORF18 and ORF30 Is Required for Late Gene Expression in Kaposi's Sarcoma-Associated Herpesvirus

2018 ◽  
Vol 93 (1) ◽  
Author(s):  
Angelica F. Castañeda ◽  
Britt A. Glaunsinger
Keyword(s):  
Gene Expression ◽  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Late Gene ◽  
Central Component ◽  
Gene Promoters ◽  
Late Gene Expression ◽  
Late Genes ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACTIn the beta- and gammaherpesviruses, a specialized complex of viral transcriptional activators (vTAs) coordinate to direct expression of virus-encoded late genes, which are critical for viral assembly and whose transcription initiates only after the onset of viral DNA replication. The vTAs in Kaposi’s sarcoma-associated herpesvirus (KSHV) are ORF18, ORF24, ORF30, ORF31, ORF34, and ORF66. While the general organization of the vTA complex has been mapped, the individual roles of these proteins and how they coordinate to activate late gene promoters remain largely unknown. Here, we performed a comprehensive mutational analysis of the conserved residues in ORF18, which is a highly interconnected vTA component. Surprisingly, the mutants were largely selective for disrupting the interaction with ORF30 but not the other three ORF18 binding partners. Furthermore, disrupting the ORF18-ORF30 interaction weakened the vTA complex as a whole, and an ORF18 point mutant that failed to bind ORF30 was unable to complement an ORF18 null virus. Thus, contacts between individual vTAs are critical as even small disruptions in this complex result in profound defects in KSHV late gene expression.IMPORTANCEKaposi’s sarcoma-associated herpesvirus (KSHV) is the etiologic agent of Kaposi’s sarcoma and other B-cell cancers and remains a leading cause of death in immunocompromised individuals. A key step in the production of infectious virions is the transcription of viral late genes, which generates capsid and structural proteins and requires the coordination of six viral proteins that form a complex. The role of these proteins during transcription complex formation and the importance of protein-protein interactions are not well understood. Here, we focused on a central component of the complex, ORF18, and revealed that disruption of its interaction with even a single component of the complex (ORF30) prevents late gene expression and completion of the viral lifecycle. These findings underscore how individual interactions between the late gene transcription components are critical for both the stability and function of the complex.

scholarly journals A Dominant-Negative Herpesvirus Protein Inhibits Intranuclear Targeting of Viral Proteins: Effects on DNA Replication and Late Gene Expression

2000 ◽  
Vol 74 (21) ◽  
pp. 10122-10131 ◽  
Author(s):  
Elizabeth E. McNamee ◽  
Travis J Taylor ◽  
David M. Knipe
Keyword(s):  
Gene Expression ◽  
Dna Replication ◽  
Viral Replication ◽  
Gene Transcription ◽  
Dominant Negative ◽  
Late Gene ◽  
Late Gene Expression ◽  
Viral Dna ◽  
Late Genes ◽  
Infected Cells

ABSTRACT The d105 dominant-negative mutant form of the herpes simplex virus 1 (HSV-1) single-stranded DNA-binding protein, ICP8 (d105 ICP8), inhibits wild-type viral replication, and it blocks both viral DNA replication and late gene transcription, although to different degrees (M. Gao and D. M. Knipe, J. Virol. 65:2666–2675, 1991; Y. M. Chen and D. M. Knipe, Virology 221:281–290, 1996). We demonstrate here that this protein is also capable of preventing the formation of intranuclear prereplicative sites and replication compartments during HSV infection. We defined three patterns of ICP8 localization using indirect immunofluorescence staining of HSV-1-infected cells: large replication compartments, small compartments, and no specific intranuclear localization of ICP8. Cells that form large replication compartments replicate viral DNA and express late genes. Cells that form small replication compartments replicate viral DNA but do not express late genes, while cells without viral replication compartments are incapable of both DNA replication and late gene expression. The d105 ICP8 protein blocks formation of prereplicative sites and large replication compartments in 80% of infected cells and formation of large replication compartments in the remaining 20% of infected cells. The phenotype ofd105 suggests a correlation between formation of large replication compartments and late gene expression and a role for intranuclear rearrangement of viral DNA and bound proteins in activation of late gene transcription. Thus, these results provide evidence for specialized machinery for late gene expression within replication compartments.

scholarly journals Identification of heterogenous nuclear ribonucleoproteins (hnRNPs) and serine- and arginine-rich (SR) proteins that induce human papillomavirus type 16 late gene expression and alter L1 mRNA splicing

2021 ◽  
Author(s):  
Chengyu Hao ◽  
Lijing Gong ◽  
Xiaoxu Cui ◽  
Johanna Jönsson ◽  
Yunji Zheng ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Papillomavirus ◽  
Sr Proteins ◽  
Mrna Splicing ◽  
Late Gene ◽  
Mrna Levels ◽  
Late Gene Expression ◽  
Human Papillomavirus Type 16 ◽  
Hnrnp Proteins ◽  
L1 And L2

AbstractWe have determined the effect of seven serine- and arginine-rich (SR) proteins and 15 heterogenous nuclear ribonucleoproteins (hnRNPs) on human papillomavirus type 16 (HPV16) late gene expression. Of the seven SR proteins analyzed here, SRSF1, SRSF3, and SRSF9 induced HPV16 late gene expression, and five of the SR proteins affected HPV16 L1 mRNA splicing. Of the 15 hnRNP proteins analyzed here, hnRNP A2, hnRNP F, and hnRNP H efficiently induced HPV16 late gene expression, and all of the hnRNPs affected HPV16 L1 mRNA levels or mRNA splicing. Thus, the majority of SR proteins and hnRNPs have the potential to regulate HPV16 L1 mRNA splicing. Strict control of the expression of the immunogenic L1 and L2 capsid proteins may contribute to the ability of HPV16 to cause persistence.

scholarly journals In Vivo and In Vitro Analysis of Baculovirusie-2 Mutants

1999 ◽  
Vol 73 (3) ◽  
pp. 2460-2468 ◽  
Author(s):  
Elena A. Prikhod’ko ◽  
Albert Lu ◽  
Joyce A. Wilson ◽  
Lois K. Miller
Keyword(s):  
Gene Expression ◽  
Cell Line ◽  
Late Gene ◽  
Wild Type ◽  
Late Gene Expression ◽  
Specific Effects ◽  
Viral Promoters ◽  
Occlusion Bodies ◽  
A Cell ◽  
Budded Virus

ABSTRACT Upon transient expression in cell culture, the ie-2gene of Autographa californica nuclear polyhedrosis virus (AcMNPV) displays three functions: transactivation of viral promoters, direct or indirect stimulation of virus origin-specific DNA replication, and arrest of the cell cycle. The ability of IE2 to trans stimulate DNA replication and coupled late gene expression is observed in a cell line derived fromSpodoptera frugiperda but not in a cell line derived fromTrichoplusia ni. This finding suggested that IE-2 may exert cell line-specific or host-specific effects. To examine the role of ie-2 in the context of infection and its possible influence on the host range, we constructed recombinants of AcMNPV containing deletions of different functional regions within ie-2 and characterized them in cell lines and larvae of S. frugiperda and T. ni. Theie-2 mutant viruses exhibited delays in viral DNA synthesis, late gene expression, budded virus production, and occlusion body formation in SF-21 cells but not in TN-5B1-4 cells. In TN-5B1-4 cells, the ie-2 mutants produced more budded virus and fewer occlusion bodies but the infection proceeded without delay. Examination of the effects of ie-2 and the respective mutants on immediate-early viral promoters in transient expression assays revealed striking differences in the relative levels of expression and differences in responses to ie-2 and its mutant forms in different cell lines. In T. ni and S. frugiperda larvae, the infectivities of the occluded form ofie-2 mutant viruses by the normal oral route of infection was 100- and 1,000-fold lower, respectively, than that of wild-type AcMNPV. The reduction in oral infectivity was traced to the absence of virions within the occlusion bodies. The infectivity of the budded form of ie-2 mutants by hemocoelic injection was similar to that of wild-type virus in both species. Thus,ie-2 mutants are viable but exhibit cell line-specific effects on temporal regulation of the infection process. Due to its effect on virion occlusion, mutants of IE-2 were essentially noninfectious by the normal route of infection in both species tested. However, since budded viruses exhibited normal infectivity upon hemocoelic injection, we conclude that ie-2 does not affect host range per se. The possibility that IE-2 exerts tissue-specific effects has not been ruled out.

scholarly journals A Downstream Polyadenylation Element in Human Papillomavirus Type 16 L2 Encodes Multiple GGG Motifs and Interacts with hnRNP H

2005 ◽  
Vol 79 (14) ◽  
pp. 9254-9269 ◽  
Author(s):  
Daniel Öberg ◽  
Joanna Fay ◽  
Helen Lambkin ◽  
Stefan Schwartz
Keyword(s):  
Gene Expression ◽  
Human Papillomavirus ◽  
Polyadenylation Signal ◽  
Late Gene ◽  
Coding Region ◽  
Late Gene Expression ◽  
Hpv 16 ◽  
Virus Particles ◽  
Human Papillomavirus Type 16 ◽  
L1 And L2

ABSTRACT Production of human papillomavirus type 16 (HPV-16) virus particles is totally dependent on the differentiation-dependent induction of viral L1 and L2 late gene expression. The early polyadenylation signal in HPV-16 plays a major role in the switch from the early to the late, productive stage of the viral life cycle. Here, we show that the L2 coding region of HPV-16 contains RNA elements that are necessary for polyadenylation at the early polyadenylation signal. Consecutive mutations in six GGG motifs located 174 nucleotides downstream of the polyadenylation signal resulted in a gradual decrease in polyadenylation at the early polyadenylation signal. This caused read-through into the late region, followed by production of the late mRNAs encoding L1 and L2. Binding of hnRNP H to the various triple-G mutants correlated with functional activity of the HPV-16 early polyadenylation signal. In addition, the polyadenylation factor CStF-64 was also found to interact specifically with the region in L2 located 174 nucleotides downstream of the early polyadenylation signal. Staining of cervix epithelium with anti-hnRNP H-specific antiserum revealed high expression levels of hnRNP H in the lower layers of cervical epithelium and a loss of hnRNP H production in the superficial layers, supporting a model in which a differentiation-dependent down regulation of hnRNP H causes a decrease in HPV-16 early polyadenylation and an induction of late gene expression.

scholarly journals Inhibition of the Ubiquitin-Proteasome System Prevents Vaccinia Virus DNA Replication and Expression of Intermediate and Late Genes

2009 ◽  
Vol 83 (6) ◽  
pp. 2469-2479 ◽  
Author(s):  
P. S. Satheshkumar ◽  
Luis C. Anton ◽  
Patrick Sanz ◽  
Bernard Moss
Keyword(s):  
Gene Expression ◽  
Vaccinia Virus ◽  
Proteasome Inhibitors ◽  
Ubiquitin Proteasome System ◽  
Late Gene ◽  
Late Gene Expression ◽  
Viral Dna ◽  
Late Genes ◽  
Ubiquitin Proteasome

ABSTRACT The ubiquitin-proteasome system has a central role in the degradation of intracellular proteins and regulates a variety of functions. Viruses belonging to several different families utilize or modulate the system for their advantage. Here we showed that the proteasome inhibitors MG132 and epoxomicin blocked a postentry step in vaccinia virus (VACV) replication. When proteasome inhibitors were added after virus attachment, early gene expression was prolonged and the expression of intermediate and late genes was almost undetectable. By varying the time of the removal and addition of MG132, the adverse effect of the proteasome inhibitors was narrowly focused on events occurring 2 to 4 h after infection, the time of the onset of viral DNA synthesis. Further analyses confirmed that genome replication was inhibited by both MG132 and epoxomicin, which would account for the effect on intermediate and late gene expression. The virus-induced replication of a transfected plasmid was also inhibited, indicating that the block was not at the step of viral DNA uncoating. UBEI-41, an inhibitor of the ubiquitin-activating enzyme E1, also prevented late gene expression, supporting the role of the ubiquitin-proteasome system in VACV replication. Neither the overexpression of ubiquitin nor the addition of an autophagy inhibitor was able to counter the inhibitory effects of MG132. Further studies of the role of the ubiquitin-proteasome system for VACV replication may provide new insights into virus-host interactions and suggest potential antipoxviral drugs.

scholarly journals Extended Function of Plasmid Partition Genes: the Sop System of Linear Phage-Plasmid N15 Facilitates Late Gene Expression

2008 ◽  
Vol 190 (10) ◽  
pp. 3538-3545 ◽  
Author(s):  
Nikolai V. Ravin ◽  
Jérôme Rech ◽  
David Lane
Keyword(s):  
Gene Expression ◽  
Late Gene ◽  
Northern Hybridization ◽  
Partition Functions ◽  
Wild Type ◽  
Late Promoter ◽  
Late Gene Expression ◽  
Reading Frame ◽  
Partition System ◽  
Plasmid Partition

ABSTRACT The mitotic stability of the linear plasmid-prophage N15 of Escherichia coli depends on a partition system closely related to that of the F plasmid SopABC. The two Sop systems are distinguished mainly by the arrangement of their centromeric SopB-binding sites, clustered in F (sopC) and dispersed in N15 (IR1 to IR4). Because two of the N15 inverted repeat (IR) sites are located close to elements presumed (by analogy with phage λ) to regulate late gene expression during the lytic growth of N15, we asked whether Sop partition functions play a role in this process. In N15, a putative Q antiterminator gene is located 6 kb upstream of the probable major late promoter and two intrinsic terminator-like sequences, in contrast to λ, where the Q gene is adjacent to the late promoter. Northern hybridization and lacZ reporter activity confirmed the identity of the N15 late promoter (p52), demonstrated antiterminator activity of the Q analogue, and located terminator sequences between p52 and the first open reading frame. Following prophage induction, N15 mutated in IR2 (downstream from gene Q) or IR3 (upstream of p52) showed a pronounced delay in lysis relative to that for wild-type N15. Expression of ir3 −-p52::lacZ during N15 wild-type lytic growth was strongly reduced relative to the equivalent ir3 + fusion. The provision of Q protein and the IR2 and SopAB proteins in trans to ir3 +-p52::lacZ increased expression beyond that seen in the absence of any one of these factors. These results indicate that the N15 Sop system has a dual role: partition and regulation of late gene transcription during lytic growth.

scholarly journals Analyses of Single-Amino-Acid Substitution Mutants of Adenovirus Type 5 E1B-55K Protein

2001 ◽  
Vol 75 (9) ◽  
pp. 4297-4307 ◽  
Author(s):  
Yuqiao Shen ◽  
Galila Kitzes ◽  
Julie A. Nye ◽  
Ali Fattaey ◽  
Terry Hermiston
Keyword(s):  
Gene Expression ◽  
Amino Acid ◽  
Human Cancer ◽  
Adenovirus Type ◽  
Single Amino Acid ◽  
Late Gene ◽  
Wild Type ◽  
Late Gene Expression ◽  
Human Cancer Cells ◽  
Adenovirus Type 5

ABSTRACT The E1B-55K protein plays an important role during human adenovirus type 5 productive infection. In the early phase of the viral infection, E1B-55K binds to and inactivates the tumor suppressor protein p53, allowing efficient replication of the virus. During the late phase of infection, E1B-55K is required for efficient nucleocytoplasmic transport and translation of late viral mRNAs, as well as for host cell shutoff. In an effort to separate the p53 binding and inactivation function and the late functions of the E1B-55K protein, we have generated 26 single-amino-acid mutations in the E1B-55K protein. These mutants were characterized for their ability to modulate the p53 level, interact with the E4orf6 protein, mediate viral late-gene expression, and support virus replication in human cancer cells. Of the 26 mutants, 24 can mediate p53 degradation as efficiently as the wild-type protein. Two mutants, R240A (ONYX-051) and H260A (ONYX-053), failed to degrade p53 in the infected cells. In vitro binding assays indicated that R240A and H260A bound p53 poorly compared to the wild-type protein. When interaction with another viral protein, E4orf6, was examined, H260A significantly lost its ability to bind E4orf6, while R240A was fully functional in this interaction. Another mutant, T255A, lost the ability to bind E4orf6, but unexpectedly, viral late-gene expression was not affected. This raised the possibility that the interaction between E1B-55K and E4orf6 was not required for efficient viral mRNA transport. Both R240A and H260A have retained, at least partially, the late functions of wild-type E1B-55K, as determined by the expression of viral late proteins, host cell shutoff, and lack of a cold-sensitive phenotype. Virus expressing R240A (ONYX-051) replicated very efficiently in human cancer cells, while virus expressing H260A (ONYX-053) was attenuated compared to wild-type virus dl309 but was more active than ONYX-015. The ability to separate the p53-inactivation activity and the late functions of E1B-55K raises the possibility of generating adenovirus variants that retain the tumor selectivity of ONYX-015 but can replicate more efficiently than ONYX-015 in a broad spectrum of cell types.

scholarly journals On the control of late gene expression in Kaposi’s sarcoma-associated herpesvirus (human herpesvirus-8)

2000 ◽  
Vol 81 (8) ◽  
pp. 2039-2047 ◽  
Author(s):  
Jean Chang ◽  
Don Ganem
Keyword(s):  
Gene Expression ◽  
Human Herpesvirus ◽  
Lytic Replication ◽  
Human Herpesvirus 8 ◽  
Late Gene ◽  
Late Gene Expression ◽  
Viral Dna ◽  
Herpesvirus 8 ◽  
Late Genes ◽  
In Cis

Herpesvirus late genes require viral DNA replication for maximal expression. Although late gene expression appears to require DNA replication in cis in alphaherpesviruses, studies in Epstein–Barr virus (EBV) suggest that this cis-requirement might not pertain to the gammaherpesviruses. Based on these findings, a system was created to investigate the elements required for the regulation of Kaposi’s sarcoma-associated herpesvirus (KSHV; human herpesvirus-8) late gene expression. The transcript of a classic late gene encoding the viral assembly protein was characterized and reporter genes driven by the assembly protein promoter region were constructed. Unlike the EBV case, expression of a reporter gene under the control of the assembly protein promoter did not display authentic regulation when removed from the context of the viral genome. Although reporter expression rose in cells displaying lytic replication, this expression was not diminished by specific inhibitors of viral DNA synthesis. Minimal core promoters were similarly unable to reproduce late gene regulation. These results suggest that proper KSHV late gene expression is likely to be dependent upon virus lytic replication in cis and indicate that the regulation of KSHV late genes more closely resembles that observed in herpes simplex virus than that described for EBV.

scholarly journals The Repressor Function of the Chlamydia Late Regulator EUO Is Enhanced by the Plasmid-Encoded Protein Pgp4

2020 ◽  
Vol 202 (8) ◽  
Author(s):  
Qiang Zhang ◽  
Christopher J. Rosario ◽  
Lauren M. Sheehan ◽  
Syed M. Rizvi ◽  
Julie A. Brothwell ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Negative Regulator ◽  
Late Gene ◽  
Late Gene Expression ◽  
New Host ◽  
Content Type ◽  
Late Genes ◽  
Developmental Form

ABSTRACT A critical step in intracellular Chlamydia infection is the production of infectious progeny through the expression of late genes. This differentiation step involves conversion from a reticulate body (RB), which is the replicating form of the bacterium, into an elementary body (EB), which is the developmental form that spreads the infection to a new host cell. EUO is an important chlamydial transcription factor that controls the expression of late genes, but the mechanisms that regulate EUO are not known. We report that a plasmid-encoded protein, Pgp4, enhanced the repressor activity of EUO. Pgp4 did not function as a transcription factor because it did not bind or directly modulate transcription of its target promoters. Instead, Pgp4 increased the ability of EUO to bind and repress EUO-regulated promoters in vitro and physically interacted with EUO in pulldown assays with recombinant proteins. We detected earlier onset of EUO-dependent late gene expression by immunofluorescence microscopy in Pgp4-deficient C. trachomatis and C. muridarum strains. In addition, the absence of Pgp4 led to earlier onset of RB-to-EB conversion in C. muridarum. These data support a role for Pgp4 as a negative regulator of chlamydial transcription that delays late gene expression. Our studies revealed that Pgp4 also has an EUO-independent function as a positive regulator of chlamydial transcription. IMPORTANCE Chlamydia trachomatis is an important human pathogen that causes more than 150 million active cases of genital and eye infection in the world. This obligate intracellular bacterium produces infectious progeny within an infected human cell through the expression of late chlamydial genes. We showed that the ability of a key chlamydial transcription factor, EUO, to repress late genes was enhanced by a plasmid-encoded protein, Pgp4. In addition, studies with Chlamydia Pgp4-deficient strains provide evidence that Pgp4 delays late gene expression in infected cells. Thus, Pgp4 is a novel regulator of late gene expression in Chlamydia through its ability to enhance the repressor function of EUO.

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