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Regulation of human neurotropic JC virus replication by alternative splicing factor SF2/ASF in glial cells.

ABSTRACT: The human neurotropic virus, JC virus (JCV), is the etiologic agent of the fatal demyelinating disease of the central nervous system, Progressive Multifocal Leukoencephalopathy (PML) that is seen primarily in immunodeficient individuals. Productive infection of JCV occurs only in glial cells, and this restriction is, to a great extent, due to the activation of the viral promoter that has cell type-specific characteristics. Earlier studies led to the hypothesis that glial-specific activation of the JCV promoter is mediated through positive and negative transcription factors that control reactivation of the JCV genome under normal physiological conditions and suppress its activation in non-glial cells.Using a variety of virological and molecular biological approaches, we demonstrate that the alternative splicing factor SF2/ASF has the capacity to exert a negative effect on transcription of the JCV promoter in glial cells through direct association with a specific DNA sequence within the viral enhancer/promoter region. Our results show that down-regulation of SF2/ASF in fetal and adult glial cells increases the level of JCV gene expression and its replication indicating that negative regulation of the JCV promoter by SF2/ASF may control reactivation of JCV replication in brain.Our results establish a new regulatory role for SF2/ASF in controlling gene expression at the transcriptional level.

SUBMITTER: Sariyer IK

PROVIDER: S-EPMC3031499 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Regulation of human neurotropic JC virus replication by alternative splicing factor SF2/ASF in glial cells.

Sariyer Ilker Kudret IK Khalili Kamel K

PloS one 20110131 1

<h4>Background</h4>The human neurotropic virus, JC virus (JCV), is the etiologic agent of the fatal demyelinating disease of the central nervous system, Progressive Multifocal Leukoencephalopathy (PML) that is seen primarily in immunodeficient individuals. Productive infection of JCV occurs only in glial cells, and this restriction is, to a great extent, due to the activation of the viral promoter that has cell type-specific characteristics. Earlier studies led to the hypothesis that glial-speci ...[more]

PMID: 21297941

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Project description:BackgroundPatients undergoing immune modulatory therapies for the treatment of autoimmune diseases such as multiple sclerosis, and individuals with an impaired-immune system, most notably AIDS patients, are in the high risk group of developing progressive multifocal leukoencephalopathy (PML), a fatal demyelinating disease of the white matter caused by human neurotropic polyomavirus, JC virus. It is now widely accepted that pathologic strains of JCV shows unique rearrangements consist of deletions and insertions within viral NCCR. While these kinds of rearrangements are related to viral tropism and pathology of the disease, their roles in molecular regulation of JCV gene expression and replication are unclear. We have previously identified SF2/ASF as a negative regulator of JCV gene expression in glial cells. This negative impact of SF2/ASF was dependent on its ability to bind a specific region mapped to the tandem repeat within viral promoter. In this report, functional role of SF2/ASF binding region in viral gene expression and replication was investigated by using deletion mutants of viral regulatory sequences.ResultsThe second 98-base-pair tandem repeat on Mad1 strain was first mutated by deletion and named Mad1-(1X98). In addition to this mutant, the CR3 region which served the binding side for SF2/ASF was also mutated and named Mad1-ΔCR3 (1X73). Both mutations were tested for SF2/ASF binding by ChIP assay. While SF2/ASF was associated with Mad1-WT and Mad1-(1X98), its interaction was completely abolished on Mad1-ΔCR3 (1X73) construct as expected. Surprisingly, reporter gene analysis of Mad1-(1X98) and Mad1-ΔCR3 (1X73) early promoter sequences showed two and three fold increase in promoter activities, respectively. The impact of "CR3" region on JCV propagation was also tested on the viral background. While replication of Mad1-(1X98) strain in glial cells was similar to Mad1-WT strain, propagation of Mad1-ΔCR3 (1X73) was less productive. Further analysis of the transcription mediated by Mad1-ΔCR3 (1X73) NCCR revealed that late gene expression was significantly affected.ConclusionsThe results of this study reveal a differential role of CR3 region within JCV NCCR in expression of JCV early and late genes.

Project description:Our previous study revealed that microRNA (miR) ‑30c represents a potential tumor suppressor gene, the expression of which is associated with decreased oncogenic potential in prostate cancer (PCa) cell lines. However, the functional role and underlying mechanisms of miR‑30c in PCa remain to be fully elucidated. Reverse transcription‑quantitative polymerase chain reaction and immunohistochemical analysis were used to detect the expression levels of alternative splicing factor/splicing factor 2 (ASF/SF2) in PCa tissues. A luciferase reporter assay was used to investigate whether ASF/SF2 may be a direct target gene of miR‑30c. In addition, the effects of miR‑30c on the proliferation and apoptosis of PCa cell lines were examined, following transfection with miR‑30c mimics. Furthermore, correlation analysis was performed to investigate the relationship between the expression of miR‑30c and ASF/SF2 and various clinicopathological parameters of patients with PCa. The present results demonstrated that PCa tissues exhibited higher levels of alternative splicing factor/splicing factor 2 (ASF/SF2), compared with normal tissues. In addition, miR‑30c was revealed to targete the 3'‑untranslated region of the ASF/SF2 gene, causing a decrease in the mRNA and protein levels of ASF/SF2. Furthermore, miR‑30c was reported to decrease cell proliferation, increase the percentage of cells in the G1 cell cycle phase, and promote apoptosis through the inhibition of ASF/SF2. Following correlation analysis using patient samples, the expression of ASF/SF2 was revealed to be tightly correlated with the pathological stage of PCa and biochemical recurrence (BCR). In addition, patients with PCa exhibiting low expression levels of miR‑30c and high expression of ASF/SF2 had significantly lower rates of BCR‑free survival. In conclusion, the present study suggested that the tumor suppressor miR‑30c may be involved in PCa tumorigenesis, possibly via targeting ASF/SF2. The combined analysis of the expression of ASF/SF2 and miR‑30c may be a valuable tool for early prediction of BCR in patients with PCa following radical prostatectomy.

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Regulation of human neurotropic JC virus replication by alternative splicing factor SF2/ASF in glial cells.

Publications

Regulation of human neurotropic JC virus replication by alternative splicing factor SF2/ASF in glial cells.

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