Biology subjects

Ziegelbauer, J. M.

Publications and source records attributed to Ziegelbauer, J. M..

5 recordsLinked to original sources

bioRxiv2023

Interferon induced circRNAs escape herpesvirus host shutoff and suppress lytic infection

A first line of defense during infection is expression of interferon (IFN)-stimulated gene products which suppress viral lytic infection. To combat this, herpesviruses express endoribonucleases to deplete host RNAs. Here we demonstrate that IFN-induced circular RNAs (circRNAs) can escape viral-mediated degradation. We performed comparative circRNA expression profiling for representative alpha- (Herpes simplex virus-1, HSV-1), beta- (human cytomegalovirus, HCMV), and gamma-herpesviruses (Kaposi sarcoma herpesvirus, KSHV; murine gamma-herpesvirus 68, MHV68). Strikingly, we found that circRNAs are, as a population, resistant to host shutoff. This observation was confirmed by ectopic expression assays of human and murine herpesvirus endoribonucleases. During primary lytic infection, ten circRNAs were commonly regulated across all subfamilies of human herpesviruses, suggesting a common mechanism of regulation. We tested one such mechanism, namely how interferon-stimulation influences circRNA expression. 67 circRNAs were upregulated by either IFN-{beta} or -{gamma} treatment, with half of these also upregulated during lytic infection. Using gain and loss of function studies we found an interferon-stimulated circRNA, circRELL1, inhibited lytic HSV-1 infection. We have previously reported circRELL1 inhibits lytic KSHV infection, suggesting a pan-herpesvirus antiviral activity. We propose a two-pronged model in which interferon-stimulated genes may encode both mRNA and circRNA with antiviral activity. This is critical in cases of host shutoff, such as alpha- and gamma-herpesvirus infection, where the mRNA products are degraded but circRNAs escape.

microbiology↗

bioRxiv2022

A virus-induced circular RNA maintains latent infection of Kaposi sarcoma herpesvirus

Non-coding RNAs (ncRNAs) play important roles in host-pathogen interactions; oncogenic viruses like Kaposi sarcoma herpesvirus (KSHV) employ ncRNAs to establish a latent reservoir and persist for the life of the host. We previously reported that KSHV infection alters a novel class of RNA, circular RNAs (circRNAs). CircRNAs are alternative splicing isoforms and regulate gene expression but, their importance in infection is largely unknown. Here, we showed that a human circRNA, hsa_circ_0001400, is induced by various pathogenic viruses, namely KSHV, Epstein-Barr virus, and human cytomegalovirus. The induction of circRNAs including circ_0001400 by KSHV is co-transcriptionally regulated, likely at splicing. Consistently, screening for circ_0001400-interacting proteins identified a splicing factor, PNISR. Functional studies using infected primary endothelial cells revealed that circ_0001400 inhibits KSHV lytic transcription and virus production. Simultaneously, the circRNA promoted cell cycle, inhibited apoptosis, and induced immune genes. RNA-pulldown assays identified transcripts interacting with circ_0001400, including TTI1, which is a component of the pro-growth mTOR complexes. We thus identified a circRNA that is pro-growth and anti-lytic replication. These results support a model in which KSHV induces circ_0001400 expression to maintain latency. Since circ_0001400 is induced by multiple viruses, this novel viral strategy may be widely employed by other viruses. AUTHOR SUMMARYCircular RNAs (circRNAs) are single-stranded, closed-circular RNAs. They coincide with linear mRNAs as alternative splice isoforms. CircRNAs binds to other RNAs or proteins to regulate their abundance or functions. We previously showed that specific human circRNAs are differentials regulated upon infection with an oncogenic DNA virus, Kaposis sarcoma herpesvirus (KSHV). Functions of such circRNAs for virus infections were largely elusive. Here, we identified that one of circRNAs, hsa_circ_0001400 controls both viral and human gene expression so that infected human cells have less virus production but better cell growth. KSHV, like other herpesviruses, has two phases: latent and lytic cycle. The functions of circ_0001400 suggest it biases the cells to latent cycle during which, unlike lytic phase, viruses express only limited number of genes to survive and new viruses are not produced. Since oncogenic viruses like KSHV mainly stay at latent phase for the life of the host, our results suggest that the virus utilized circRNAs to switch to latent phase after the infection. Infection of other pathogenic viruses like Epstein-Barr virus and human cytomegalovirus were also found to induce circ_0001400. The circRNA may be thus controlling infection of various viruses.

microbiology↗

bioRxiv2022

Alternative splicing and circRNA biogenesis driven by alpha and gamma herpesviruses

Herpesviruses rely on the host transcriptional machinery for expression of their >100 different transcripts. To prioritize viral transcripts, herpesviruses affect significant changes in all stages of gene expression. Herein, we examined how herpesviruses alter circular RNA (circRNA) synthesis resulting in distinct characteristics for those expressed from the host or viral genome. CircRNA were identified with our Circrnas in Host And viRuses anaLysis pIpEline (CHARLIE) capable of de novo annotation using aggregated calls from five distinct tools. Comparative profiling for Herpes Simplex Virus-1, Kaposi sarcoma-associated herpesvirus, and murine gammaherpesvirus 68 identified thousands of back splicing variants, including circRNA species commonly expressed in all phases of infection. CircRNA expression was highest during lytic infection, with a transcriptional density 500-fold greater than the host. We characterized cis- and trans-elements controlling back splicing and found viral circRNAs are generally resistant to spliceosome inhibition or depletion, with >90% lacking canonical splice donor-acceptor sites. Using eCLIP and 4sU-Sequencing, we determined that the viral RNA binding protein, ORF57, enhanced synthesis for a subset of viral and host circRNAs. Our work elucidates a unique splicing mechanism driven by late lytic replication and identifies a class of transcripts with potential to function in replication, persistence, or tumorigenesis.

microbiology↗

bioRxiv2022

Transcriptional analysis identifies overlapping and tissue-distinct profiles between Kaposi sarcoma tumors of the skin and gastrointestinal tract

Kaposi sarcoma (KS), caused by Kaposi sarcoma herpesvirus (KSHV), is a multicentric tumor characterized by abnormal vasculature and proliferation of KSHV-infected spindle cells. KS commonly involves the skin but in severe cases KS can also involve the gastrointestinal tract (GI). Here, we sought to compare the cellular and KSHV gene expression signatures of skin and GI KS lesions. Skin and GI KS were compared to normal matched samples using bulk RNA sequencing.Twenty-two paired samples of KS and normal tissue were obtained (skin (10 pairs) and GI (12 pairs)) from 19 patients with KS of whom 17 had concurrent HIV infection. Seven paired samples were from patients who had received prior KS therapy. Three patients provided both skin and GI samples at the same timepoint. These analyses identified 370 differentially expressed genes unique to cutaneous KS and 58 DEGs unique to GI KS compared to normal skin or GI tissues. Twenty-six differentially expressed genes overlapped between skin and GI KS, which included FLT4, which encodes for a VEGF-C and VEGF-D receptor, and STC1. KSHV infection of primary lymphatic endothelial cells (LECs) resulted in increased angiogenesis, and repression of STC1 or FLT4 inhibited angiogenesis. The analyses of KSHV expression from KS lesions identified certain lytic genes, specifically ORF75, that were consistently expressed, and these expression patterns differed from laboratory infection of LECs with KSHV and KSHV gene expression in PEL cell lines. This study demonstrates that complex patterns of gene expression are found in KS tissue that differ from the canonical latent/lytic programs seen in KSHV cell lines and also demonstrates differences in viral gene and clinically relevant host gene expression in skin and GI KS that may offer insights into the pathogenesis of these forms of KS. One sentence summaryKaposi sarcoma that manifests in the skin and gastrointestinal tracts differ by human and viral gene expression.

microbiology↗

bioRxiv2019

Oncogenic herpesvirus engages the endothelial transcription factors SOX18 and PROX1 to increase viral genome copies and virus production

Kaposi sarcoma (KS) is a tumour of endothelial origin caused by KS herpesvirus (KSHV) infection and suggested to originate from lymphatic endothelial cells (LECs). While KSHV establishes latency in virtually all susceptible cell types, LECs support a spontaneous lytic gene expression program with high viral genome copies and release of infectious virus. Here, we investigated the role of PROX1, SOX18 and COUPTF2, drivers of lymphatic endothelial fate during embryogenesis, in this unique KSHV infection program. We found that these factors were co-expressed in KS tumours with the viral lytic marker K8.1, and that SOX18 and PROX1 regulate KSHV infection via two independent mechanisms. SOX18 binds to the viral origins of replication and its depletion or chemical inhibition significantly reduced the KSHV genome copies in LECs. PROX1 interacts with ORF50, the initiator of the lytic cascade, increases lytic gene expression and virus production and its depletion reduces KSHV spontaneous lytic reactivation. Upon lytic replication, PROX1 binds to the KSHV genome in the promoter region of ORF50 and enhances its transactivation activity. These results demonstrate the importance of two endothelial transcription factors in the regulation of the KSHV life cycle and introduce SOX18 inhibition as a potential, novel therapeutic modality for KS.

microbiology↗