The human polyomaviruses, BK virus and JC virus, have long been associated with serious diseases including polyomavirus nephropathy and progressive multifocal leukoencephalopathy. Both viruses establish ubiquitous, persistent infections in healthy individuals. Reactivation can occur when the immune system is impaired, leading to disease progression. Recently, the human polyomavirus family has expanded with the identification of three new viruses (KI, WU and Merkel cell polyomavirus), all of which may prove to be involved in human disease. This review describes the general aspects of human polyomavirus infections and pathogenicity. Current topics of investigation and future directions in the field are also discussed.

The bacterial ribosome is an extremely complicated macromolecular complex the in vivo biogenesis of which is poorly understood. Although several bona fide assembly factors have been identified, their precise functions and temporal relationships are not clearly defined. Here we describe the involvement of an Escherichia coli GTPase, CgtA E , in late steps of large ribosomal subunit biogenesis. CgtA E belongs to the Obg/CgtA GTPase subfamily, whose highly conserved members are predominantly involved in ribosome function. Mutations in CgtA E cause both polysome and rRNA processing defects; small-and large-subunit precursor rRNAs accumulate in a cgtA E mutant. In this study we apply a new semiquantitative proteomic approach to show that CgtA E is required for optimal incorporation of certain late-assembly ribosomal proteins into the large ribosomal subunit. Moreover, we demonstrate the interaction with the 50S ribosomal subunits of specific nonribosomal proteins (including heretofore uncharacterized proteins) and define possible temporal relationships between these proteins and CgtA E . We also show that purified CgtA E associates with purified ribosomal particles in the GTP-bound form. Finally, CgtA E cofractionates with the mature 50S but not with intermediate particles accumulated in other large ribosome assembly mutants.Although assembly of prokaryotic ribosomes from purified ribosomal proteins (r-proteins) and rRNAs can occur independently in vitro (51, 52, 75), accumulating evidence suggests that, as in eukaryotes, in vivo prokaryotic ribosome biogenesis depends on the aid of nonribosomal factors. The higher temperature, increased Mg 2ϩ concentration, and longer incubation times necessary for in vitro relative to in vivo conditions (51) imply that the likely role of accessory factors is to expedite ribosome maturation by reducing the activation energies for the rate-limiting reactions. Although not complicated by the involvement of different cellular compartments, the prokaryotic ribosome assembly process is far from simple, requiring coordinated synthesis of 3 rRNAs (5S, 16S, and 23S) and 55 r-proteins, processing and modification of these components, and their appropriate sequential unification to produce mature ribosomes. The details of how this process is controlled temporally, even spatially, in the small bacterial cell are incompletely understood.More than 170 nonribosomal proteins that transiently associate with different preribosomal particles have been identified in Saccharomyces cerevisiae (19,22,38,62), largely due to progress in combining biochemical affinity purification methods with newly developed proteomic techniques (24,25,29,54,58,61). By contrast, only a few such assembly factors have been found in bacteria, and most were identified via conventional genetic methods. These proteins consist of RNA-modifying enzymes such as methyltransferases and pseudouridine synthases, RNA-remodeling proteins such as RNA helicases, chaperones, GTPases, and proteins with unknown functions (1,

BK virus (BKV) is a nonenveloped, ubiquitous human polyomavirus that establishes a persistent infection in healthy individuals. It can be reactivated, however, in immunosuppressed patients and cause severe diseases, including polyomavirus nephropathy. The entry and disassembly mechanisms of BKV are not well defined. In this report, we characterized several early events during BKV infection in primary human renal proximal tubule epithelial (RPTE) cells, which are natural host cells for BKV. Our results demonstrate that BKV infection in RPTE cells involves an acidic environment relatively early during entry, followed by transport along the microtubule network to reach the endoplasmic reticulum (ER). A distinct disulfide bond isomerization and cleavage pattern of the major capsid protein VP1 was observed, which was also influenced by alterations in pH and disruption of trafficking to the ER. A dominant negative form of Derlin-1, an ER protein required for retro-translocation of certain misfolded proteins, inhibited BKV infection. Consistent with this, we detected an interaction between Derlin-1 and VP1. Finally, we show that proteasome function is also linked to BKV infection and capsid rearrangement. These results indicate that BKV early entry and disassembly are highly regulated processes involving multiple cellular components.BK virus (BKV) is a member of the Polyomaviridae family, which also includes the well-studied simian virus 40 (SV40) and human JC virus (JCV) (21). Like other members of this family, BKV has a small (40 to 45 nm in diameter), nonenveloped, icosahedral capsid that contains an ϳ5-kb circular double-stranded DNA genome (5). BKV infection is ubiquitous in the human population and occurs during early childhood (24). Primary infection with BKV is followed by dissemination to the kidney and urinary tract, in particular to kidney tubule epithelial cells and urinary tract epithelial cells, where the virus establishes a lifelong persistent infection (9). This infection remains asymptomatic in immunocompetent individuals, but under conditions of immunosuppression, BKV can undergo reactivation resulting in viral shedding in the urine and may eventually lead to severe diseases, such as polyomavirus nephropathy in renal transplant patients and hemorrhagic cystitis in bone marrow transplant recipients (2). The incidence of BKV-related disease has greatly increased recently due to the introduction of new and more potent immunosuppressive agents and the increase in the number of transplants performed (21). No specific antiviral drugs for BKV infection are currently available, and the most common approach to control BKV reactivation is to reduce the immunosuppression, which might leave patients at risk for graft rejection (53). In addition, the immune components that are involved in controlling BKV persistence and reactivation are not well defined. Therefore, a better understanding of the BKV life cycle is warranted to aid in the design of novel, more-efficient antiviral strategies.Based on cryo-electron mic...

The APOBEC3 family of DNA cytosine deaminases has important roles in innate immunity and cancer. It is unclear how DNA tumor viruses regulate these enzymes and how these interactions, in turn, impact the integrity of both the viral and cellular genomes. Polyomavirus (PyVs) are small DNA pathogens that contain oncogenic potentials. In this study, we examined the effects of PyV infection on APOBEC3 expression and activity. We demonstrate that APOBEC3B is specifically upregulated by BK polyomavirus (BKPyV) infection in primary kidney cells and that the upregulated enzyme is active. We further show that the BKPyV large T antigen, as well as large T antigens from related polyomaviruses, is alone capable of upregulating APOBEC3B expression and activity. Furthermore, we assessed the impact of A3B on productive BKPyV infection and viral genome evolution. Although the specific knockdown of APOBEC3B has little short-term effect on productive BKPyV infection, our informatics analyses indicate that the preferred target sequences of APOBEC3B are depleted in BKPyV genomes and that this motif underrepresentation is enriched on the nontranscribed stand of the viral genome, which is also the lagging strand during viral DNA replication. Our results suggest that PyV infection upregulates APOBEC3B activity to influence virus sequence composition over longer evolutionary periods. These findings also imply that the increased activity of APOBEC3B may contribute to PyV-mediated tumorigenesis. IMPORTANCEPolyomaviruses (PyVs) are a group of emerging pathogens that can cause severe diseases, including cancers in immunosuppressed individuals. Here we describe the finding that PyV infection specifically induces the innate immune DNA cytosine deaminase APOBEC3B. The induced APOBEC3B enzyme is fully functional and therefore may exert mutational effects on both viral and host cell DNA. We provide bioinformatic evidence that, consistent with this idea, BK polyomavirus genomes are depleted of APOBEC3B-preferred target motifs and enriched for the corresponding predicted reaction products. These data imply that the interplay between PyV infection and APOBEC proteins may have significant impact on both viral evolution and virusinduced tumorigenesis. P olyomaviruses (PyVs) are a family of small nonenveloped viruses containing an ϳ5-kb circular double-stranded DNA genome. Most human PyVs establish a subclinical persistent infection in healthy individuals (1). These viruses can reactivate under various immunosuppression conditions and cause a variety of severe diseases, including cancers (2). Among them, BK polyomavirus (BKPyV) reactivation is a major concern in kidney and bone marrow transplant patients due to the possibility of development of polyomavirus-associated nephropathy and hemorrhagic cystitis, respectively (3). Recently, there have also been increasing reports demonstrating an association between BKPyV infection and the occurrence of renourinary tumors (4). JC polyomavirus (JCPyV) reactivation can lead to progressive multifocal leu...

BK virus (BKV) is a polyomavirus that ubiquitously infects the human population. Following a typically subclinical primary infection, BKV establishes a lifelong persistent infection in the kidney and urinary tract. BKV is known to reactivate and cause severe disease in immunosuppressed patients, particularly renal and bone marrow transplant patients. Infection of BKV in rodent animal models or cells in culture often results in tumor formation or transformation, respectively. When co-expressed with activated oncogenes, BKV large tumor antigen drives the transformation of primary human cells. An etiological role of BKV in human cancer, however, remains controversial. Multiple reports have demonstrated conflicting results in regards to the presence of BKV sequences and/or proteins in various tumor types. This review compiles the most recent findings of BKV detection in a number of human cancers. Due to the lack of conclusive causality data from these studies, there does not appear to be a definitive association between BKV and human cancers.

To probe the cellular phenotype and biochemical function associated with the G domains of Escherichia coli EngA (YfgK, Der), mutations were created in the phosphate binding loop of each. Neither an S16A nor an S217A variant of G domain 1 or 2, respectively, was able to support growth of an engA conditional null. Polysome profiles of EngA-depleted cells were significantly altered, and His 6 -EngA was found to cofractionate with the 50S ribosomal subunit. The variants were unable to complement the abnormal polysome profile and were furthermore significantly impacted with respect to in vitro GTPase activity. Together, these observations suggest that the G domains have a cooperative function in ribosome stability and/or biogenesis.

The bacterial response to stress is controlled by two proteins, RelA and SpoT. RelA generates the alarmone (p)ppGpp under amino acid starvation, whereas SpoT is responsible for (p)ppGpp hydrolysis and for synthesis of (p)ppGpp under a variety of cellular stress conditions. It is widely accepted that RelA is associated with translating ribosomes. The cellular location of SpoT, however, has been controversial. SpoT physically interacts with the ribosome-associated GTPase CgtA, and we show here that, under an optimized salt condition, SpoT is also associated with a pre-50S particle. Analysis of spoT and cgtA mutants and strains overexpressing CgtA suggests that the ribosome associations of SpoT and CgtA are mutually independent. The steady-state level of (p)ppGpp is increased in a cgtA mutant, but the accumulation of (p)ppGpp during amino acid starvation is not affected, providing strong evidence that CgtA regulates the (p)ppGpp level during exponential growth but not during the stringent response. We show that CgtA is not associated with pre-50S particles during amino acid starvation, indicating that under these conditions in which (p)ppGpp accumulates, CgtA is not bound either to the pre-50S particle or to SpoT. We propose that, in addition to its role as a 50S assembly factor, CgtA promotes SpoT (p)ppGpp degradation activity on the ribosome and that the loss of CgtA from the ribosome is necessary for maximal (p)ppGpp accumulation under stress conditions. Intriguingly, we found that in the absence of spoT and relA, cgtA is still an essential gene in Escherichia coli.

BK polyomavirus (BKPyV) is an emerging pathogen whose reactivation causes severe disease in transplant patients. Unfortunately, there is no specific anti-BKPyV treatment available, and host cell components that affect the infection outcome are not well characterized. In this report, we examined the relationship between BKPyV productive infection and the activation of the cellular DNA damage response (DDR) in natural host cells. Our results showed that both the ataxia-telangiectasia mutated (ATM)- and ATM and Rad-3-related (ATR)-mediated DDR were activated during BKPyV infection, accompanied by the accumulation of polyploid cells. We assessed the involvement of ATM and ATR during infection using small interfering RNA (siRNA) knockdowns. ATM knockdown did not significantly affect viral gene expression, but reduced BKPyV DNA replication and infectious progeny production. ATR knockdown had a slightly more dramatic effect on viral T antigen (TAg) and its modified forms, DNA replication, and progeny production. ATM and ATR double knockdown had an additive effect on DNA replication and resulted in a severe reduction in viral titer. While ATM mainly led to the activation of pChk2 and ATR was primarily responsible for the activation of pChk1, knockdown of all three major phosphatidylinositol 3-kinase-like kinases (ATM, ATR, and DNA-PKcs) did not abolish the activation of γH2AX during BKPyV infection. Finally, in the absence of ATM or ATR, BKPyV infection caused severe DNA damage and aberrant TAg staining patterns. These results indicate that induction of the DDR by BKPyV is critical for productive infection, and that one of the functions of the DDR is to minimize the DNA damage which is generated during BKPyV infection.