Biology subjects

King, K. M.

Publications and source records attributed to King, K. M..

4 recordsLinked to original sources

bioRxiv2025

Human Papillomavirus Infection Blunts STING Signaling and Alters Downstream Response

The cGAS/STING pathway is an important innate immune pathway that senses and responds to foreign DNA or damaged host DNA. cGAS recognizes DNA and generates the second messenger cGAMP which activates STING. STING then creates a platform for IRF3 to be phosphorylated before pIRF3 induces a type I interferon response resulting in transcription of anti-viral genes. We examined how HPV18 infection modulates cGAS/STING activation following DNA stimulation in primary human foreskin keratinocytes from three different donors. Following exogenous activation, HPV(+) cells produced higher levels of cGAMP compared to patient-matched HPV(-) cells yet phosphorylation of STING and IRF3 was reduced in the HPV(+) cells. The reduced STING and IRF3 activation corresponded with a selective dampening of type I interferon driven antiviral genes and pro-inflammatory cytokines in HPV(+) cells. Simultaneously, HPV(+) cells had a baseline increase in genes associated with epithelial proliferation and skin development, which remained elevated following cGAMP treatment. We demonstrate that both E6 and E7 are required and sufficient to drive increased cGAMP levels and attenuate STING activation. Our results characterize cGAS/STING pathway activation across time in HPV positive and negative cells and demonstrate that the HPV oncogenes cooperate to suppress early STING activation despite higher levels of cGAMP in these HPV(+) cells. These findings indicate that HPV modulates cGAS/STING signaling to blunt antiviral defenses without impacting other arms of the cGAS/STING pathway response. ImportanceIn this study, we used primary human keratinocytes from multiple donors to track cGAS/STING activity over time following stimulation. We found that although HPV(+) cells produced more cGAMP, these cells showed reduced early activation of STING and IRF3. Downstream of cGAS/STING, many antiviral genes were suppressed by HPV however, epithelial proliferation-related genes had an HPV-mediated increase in baseline expression which remained higher than uninfected cells following cGAMP stimulation. We identified the HPV oncogenes E6 and E7 work together to enhance cGAMP levels and suppress parts of the downstream STING signaling response. These findings reveal that HPV attenuates cGAS/STING signaling and downstream events, selectively suppressing antiviral genes without inhibiting other parts of the pathway. Our results provide new insights into how HPV modulates innate immune responses.

microbiology↗

bioRxiv2024

PRMT1 Modulates Alternative Splicing to Enhance HPV18 mRNA Stability and Promote the Establishment of Infection

Only persistent HPV infections lead to the development of cancer. Thus, understanding the virus-host interplay that influences the establishment of viral infection has important implications for HPV biology and human cancers. The ability of papillomaviruses to establish in cells requires the strict temporal regulation of viral gene expression in sync with cellular differentiation. This control primarily happens at the level of RNA splicing and polyadenylation. However, the details of how this spatio-temporal regulation is achieved still need to be fully understood. Until recently, it has been challenging to study the early events of the HPV lifecycle following infection. We used a single-cell genomics approach to identify cellular factors involved in viral infection and establishment. We identify protein arginine N-methyltransferase 1 (PRMT1) as an important factor in viral infection of primary human cervical cells. PRMT1 is the main cellular enzyme responsible for asymmetric dimethylation of cellular proteins. PRMT1 is an enzyme responsible for catalyzing the methylation of arginine residues on various proteins, which influences processes such as RNA processing, transcriptional regulation, and signal transduction. In this study, we show that HPV18 infection leads to increased PRMT1 levels across the viral lifecycle. PRMT1 is critical for the establishment of a persistent infection in primary cells. Mechanistically, PRMT1 inhibition leads to a highly dysregulated viral splicing pattern. Specifically, reduced PRMT1 activity leads to intron retention and a change in the E6 and E7 expression ratio. In the absence of PRMT1, viral transcripts are destabilized and subject to degradation via the nonsense-mediated decay (NMD) pathway. These findings highlight PRMT1 as a critical regulator of the HPV18 lifecycle, particularly in RNA processing, and position it as a potential therapeutic target for persistent HPV18 infections.

microbiology↗

bioRxiv2021

Insight into Cross-Amyloid Interactions and Morphologies: Molecular Dynamics Simulations of Model Peptide Fragments of Amyloid-β (Aβ16-22) and Islet Amyloid Polypeptide (IAPP20-29)

Amyloid-beta (A{beta}) and islet amyloid polypeptide (IAPP) are small peptides, classified as amyloids, that have the potential to self-assemble and form cytotoxic species, such as small soluble oligomers and large insoluble fibrils. The formation of A{beta} aggregates facilitates the progression of Alzheimers disease (AD), while IAPP aggregates induce pancreatic {beta}-cell apoptosis, leading to exacerbation of Type 2 diabetes (T2D). Cross-amyloid interactions between A{beta} and IAPP have been described both in vivo and in vitro, implying the role of A{beta} or IAPP as modulators of cytotoxic self-aggregation of each peptide, and suggesting that A{beta}-IAPP interactions are a potential molecular link between AD and T2D. Using molecular dynamics simulations, "hot spot" regions of the two peptides were studied to understand the formation of hexamers in a heterogenous and homogenous peptide-containing environment. Systems of only A{beta}(16-22) peptides formed antiparallel, {beta}-barrel-like structures, while systems of only IAPP(20-29) peptides formed stacked, parallel beta strands and had relatively unstable aggregation structures after 2 s of simulation time. Systems containing both A{beta} and IAPP (1:1 ratio) hexamers showed antiparallel, {beta}-barrel-like structures, with an interdigitated arrangement of A{beta}(16-22) and IAPP(20-29). These {beta}-barrel structures have features of cytotoxic amyloid species identified in previous literature. Ultimately, this work seeks to provide atomistic insight into both the mechanism behind cross-amyloid interactions and structural morphologies of these toxic amyloid species. Statement of SignificanceMolecular knowledge, biophysical characterization, structural morphologies, and formation pathways of amyloid oligomers - specifically low-molecular weight, cross-amyloid oligomers - remain preliminary and undefined. Characterizing interactions between homogenous and heterogenous amyloid oligomers is of great interest given that certain oligomer morphologies contribute to cytotoxicity, eventually resulting in comorbid diseases such as Alzheimers disease (AD) and Type 2 Diabetes Mellitus (T2DM). Utilizing model systems (e.g., fragments of full-length peptides) and molecular dynamics (MD) simulations to probe the biophysical underpinnings of cross-amyloid oligomer structures is the first step in understanding the dynamics, stability, and potential modes of cytotoxicity of these species, providing important insights into targetable biomolecular structures.

biochemistry↗

bioRxiv2021

Co-evolutionary analysis suggests a role for TLR9 in papillomavirus restriction

A.Upon infection, DNA viruses can be sensed by pattern recognition receptors (PRRs) leading to the activation of type I and III interferons, aimed at blocking infection. Therefore, viruses must inhibit these signaling pathways, avoid being detected, or both. Papillomavirus virions are trafficked from early endosomes to the Golgi apparatus and wait for the onset of mitosis to complete nuclear entry. This unique subcellular trafficking strategy avoids detection by cytoplasmic PRRs, a property that may contribute to establishment of infection. However, as the capsid uncoats within acidic endosomal compartments, the viral DNA may be exposed to detection by toll-like receptor (TLR) 9. In this study we characterize two new papillomaviruses from bats and use molecular archeology to demonstrate that their genomes altered their nucleotide composition to avoid detection by TLR9, providing evidence that TLR9 acts as a PRR during papillomavirus infection. Furthermore, we demonstrate that TLR9, like other components of the innate immune system, is under evolutionary selection in bats, providing the first direct evidence for co-evolution between papillomaviruses and their hosts.

microbiology↗