Expression of Simian Virus 40 Large T-Antigen: A Case Control Study of Non-Hodgkin Lymphoma

Background: Cancer such as neoplasma lymphoma is related to the immune system. Of all lymphoma cases, about 90% are Non-Hodgkin Lymphoma (NHL). In the United Kingdom, NHL is among the five most frequently diagnosed cancer cases. In Indonesia, based on statistical data from the Dharmais Cancer Center Hospital in 2006, NHL was in the ten most commonly diagnosed cancers. The global incidence of NHL continues to increase annually in western countries, especially in Caucasian populations, men and the elderly. The etiology of NHL is uncertain. Viral infection is thought to be associated with the pathogenesis of NHL. One of them is the Simian 40 virus (SV 40) infection. In some studies, positive SV40 sequencing was found in the majority of NHL cases, but other research found no significant relationship between SV40 infection and NHL. Objective: This study aimed to identify the frequency of SV40 infection in NHL cases in Yogyakarta and to determine the relationship between SV40 infection in NHL cases and clinical patients. Methods: This study examined 102 samples of paraffin blocks from subjects diagnosed with NHL in the Anatomical Pathology Laboratory of Dr. Sardjito Hospital from 2014 through 2016. DNA extraction was done and the results were used for PCR. SV40 genome primer sets were used for the PCR process to identify any positive cases. Results: In this study, all 102 subjects did not indicate genomic L-Tag positive cases of NHL. Conclusion: It cannot be proved that SV40 has a role in the pathogenesis of NHL in Indonesia.

Differential SP1 Interactions in SV40 Chromatin from Virions and Minichromosomes

ABSTRACTSP1 binding in SV40 chromatin in vitro and in vivo was characterized in order to better understand its role during the initiation of early transcription. We observed that chromatin from disrupted virions, but not minichromosomes, was efficiently bound by HIS-tagged SP1 in vitro, while the opposite was true for the presence of endogenous SP1 introduced in vivo. Using ChIP-Seq to compare the location of SP1 to nucleosomes carrying modified histones, we found that SP1 could occupy its whole binding site in virion chromatin but only the early side of its binding site in most of the minichromosomes carrying modified histones due to the presence of overlapping nucleosomes. The results suggest that during the initiation of an SV40 infection, SP1 binds to an open region in SV40 virion chromatin but quickly triggers chromatin reorganization and its own removal by a hit and run mechanism.

BK polyomavirus evades innate immune sensing by disrupting the mitochondrial network and membrane potential and by promoting mitophagy

Immune escape contributes to viral persistence, yet little is known about human polyomaviruses. BK-polyomavirus (BKPyV) asymptomatically infects 90% of the human population, but causes early allograft failure in 10% of kidney transplants. Despite inducing potent virus-specific T-cells and neutralizing antibodies, BKPyV persists in the kidneys and regularly escapes from immune control as indicated by urinary shedding in immunocompetent individuals. Here, we report that BKPyV disrupts the mitochondrial network and its membrane potential when expressing the 66aa-long agnoprotein during late replication. Agnoprotein impairs nuclear IRF3-translocation, interferon-β expression, and promotes p62-mitophagy in vitro and in kidney transplant biopsies. Agnoprotein-mutant viruses unable to disrupt mitochondria show reduced replication, which can be rescued by type-I-interferon-blockade, TBK1-inhibition, or CoCl2 treatment. Agnoprotein is necessary and sufficient, using its amino-terminal and central domain for mitochondrial targeting and disruption, respectively. JCPyV- and SV40-infection similarly disrupt the mitochondrial network indicating a conserved mechanism facilitating polyomavirus persistence and post-transplant disease.

Interaction between Simian Virus 40 Major Capsid Protein VP1 and Cell Surface Ganglioside GM1 Triggers Vacuole Formation

ABSTRACT Simian virus 40 (SV40), a polyomavirus that has served as an important model to understand many aspects of biology, induces dramatic cytoplasmic vacuolization late during productive infection of monkey host cells. Although this activity led to the discovery of the virus in 1960, the mechanism of vacuolization is still not known. Pentamers of the major SV40 capsid protein VP1 bind to the ganglioside GM1, which serves as the cellular receptor for the virus. In this report, we show that binding of VP1 to cell surface GM1 plays a key role in SV40 infection-induced vacuolization. We previously showed that SV40 VP1 mutants defective for GM1 binding fail to induce vacuolization, even though they replicate efficiently. Here, we show that interfering with GM1-VP1 binding by knockdown of GM1 after infection is established abrogates vacuolization by wild-type SV40. Vacuole formation during permissive infection requires efficient virus release, and conditioned medium harvested late during SV40 infection rapidly induces vacuoles in a VP1- and GM1-dependent fashion. Furthermore, vacuolization can also be induced by a nonreplicating SV40 pseudovirus in a GM1-dependent manner, and a mutation in BK pseudovirus VP1 that generates GM1 binding confers vacuole-inducing activity. Vacuolization can also be triggered by purified pentamers of wild-type SV40 VP1, but not by GM1 binding-defective pentamers or by intracellular expression of VP1. These results demonstrate that SV40 infection-induced vacuolization is caused by the binding of released progeny viruses to GM1, thereby identifying the molecular trigger for the activity that led to the discovery of SV40. IMPORTANCE The DNA tumor virus SV40 was discovered more than a half century ago as a contaminant of poliovirus vaccine stocks, because it caused dramatic cytoplasmic vacuolization of permissive host cells. Although SV40 played a historically important role in the development of molecular and cellular biology, restriction mapping, molecular cloning, and whole-genome sequencing, the basis of this vacuolization phenotype was unknown. Here, we show that SV40-induced vacuolization is triggered by the binding of the major viral capsid protein, VP1, to a cell surface ganglioside receptor, GM1. No other viral proteins or virus replication is required for vacuole formation. Other polyomaviruses utilize different ganglioside receptors, but they do not induce vacuolization. This work identifies the molecular trigger for the phenotype that led to the discovery of this important virus and provides the first molecular insight into an unusual and enigmatic cytopathic effect due to virus infection.

The Endoplasmic Reticulum Membrane J Protein C18 Executes a Distinct Role in Promoting Simian Virus 40 Membrane Penetration

ABSTRACTThe nonenveloped simian virus 40 (SV40) hijacks the three endoplasmic reticulum (ER) membrane-bound J proteins B12, B14, and C18 to escape from the ER into the cytosol en route to successful infection. How C18 controls SV40 ER-to-cytosol membrane penetration is the least understood of these processes. We previously found that SV40 triggers B12 and B14 to reorganize into discrete puncta in the ER membrane called foci, structures postulated to represent the cytosol entry site (C. P. Walczak, M. S. Ravindran, T. Inoue, and B. Tsai, PLoS Pathog10:e1004007, 2014). We now find that SV40 also recruits C18 to the virus-induced B12/B14 foci. Importantly, the C18 foci harbor membrane penetration-competent SV40, further implicating this structure as the membrane penetration site. Consistent with this, a mutant SV40 that cannot penetrate the ER membrane and promote infection fails to induce C18 foci. C18 also regulates the recruitment of B12/B14 into the foci. In contrast to B14, C18's cytosolic Hsc70-binding J domain, but not the lumenal domain, is essential for its targeting to the foci; this J domain likewise is necessary to support SV40 infection. Knockdown-rescue experiments reveal that C18 executes a role that is not redundant with those of B12/B14 during SV40 infection. Collectively, our data illuminate C18's contribution to SV40 ER membrane penetration, strengthening the idea that SV40-triggered foci are critical for cytosol entry.IMPORTANCEPolyomaviruses (PyVs) cause devastating human diseases, particularly in immunocompromised patients. As this virus family continues to be a significant human pathogen, clarifying the molecular basis of their cellular entry pathway remains a high priority. To infect cells, PyV traffics from the cell surface to the ER, where it penetrates the ER membrane to reach the cytosol. In the cytosol, the virus moves to the nucleus to cause infection. ER-to-cytosol membrane penetration is a critical yet mysterious infection step. In this study, we clarify the role of an ER membrane protein called C18 in mobilizing the simian PyV SV40, a PyV archetype, from the ER into the cytosol. Our findings also support the hypothesis that SV40 induces the formation of punctate structures in the ER membrane, called foci, that serve as the portal for cytosol entry of the virus.