Biology subjects

Wang, K.-H.

Publications and source records attributed to Wang, K.-H..

4 recordsLinked to original sources

bioRxiv2023

Prolonged Kaposi's Sarcoma-associated Herpesvirus vIL-6 Exposure Enhances Inflammatory Responses by Epigenetic Reprogramming

Kaposi sarcoma-associated herpesvirus (KSHV) inflammatory cytokine syndrome (KICS) is a newly described chronic inflammatory disease condition caused by KSHV infection and is characterized by high KSHV viral load and sustained elevations of serum KSHV-encoded IL-6 (vIL-6) and human IL-6 (hIL-6). KICS has significant immortality and possesses greater risks of having other complications, which include malignancies. Although prolonged inflammatory vIL-6 exposure by persistent KSHV infection is expected to have key roles in subsequent disease development, the biological effects of prolonged vIL-6 exposure remain elusive. Using thiol-Linked Alkylation for the Metabolic Sequencing and Cleavage Under Target & Release Using Nuclease analysis, we studied the effect of prolonged vIL-6 exposure in chromatin landscape and resulting cytokine production. The studies showed that prolonged vIL-6 exposure increased Bromodomain containing 4 (BRD4) and histone H3 lysine 27 acetylation co-occupancies on chromatin, and the recruitment sites were frequently co-localized with poised RNAPII with associated enzymes. Increased BRD4 recruitment on promoters was associated with increased and prolonged NF-KB p65 binding after the lipopolysaccharide stimulation. The p65 binding resulted in quicker and sustained transcription bursts from the promoters; this mechanism increased total amounts of hIL-6 and IL-10 in tissue culture. Pretreatment with the BRD4 inhibitor, OTX015, eliminated the enhanced inflammatory cytokine production. These findings suggest that persistent vIL-6 exposure may establish a chromatin landscape favorable for the reactivation of inflammatory responses in monocytes. This epigenetic memory may explain the greater risk of chronic inflammatory disease development in KSHV-infected individuals. Author summaryCombined and continuous cytokine stimulation triggers transcription reprogramming and is often used for specific tissue development. Continuous vIL-6 exposure occurs in KSHV-infected patients and leads to inflammatory cytokine storm with high mortality. However, possible epigenetic reprogramming by the vIL-6 and its association with pathogenesis remain unclear. Here we demonstrate the establishment of a new chromatin landscape mediated by BRD4 through prolonged vIL-6 exposure which contributes to more robust and rapid transcription and increased cytokines production. Inhibition of BRD4 suppressed this inflammatory response. Our results indicate that targeting the epigenetic effect of viral cytokines may lead to novel therapies for KSHV-induced inflammatory cytokine storms.

microbiology↗

bioRxiv2022

Exploring Taxonomic and Functional Microbiome of Hawaiian Stream and Spring Irrigation Water Systems Using Illumina and Oxford Nanopore Sequencing Platforms

Irrigation water is a potential source of contamination that carries plant and foodborne human pathogens and provides a niche for survival and proliferation of microbes in agricultural settings. This project investigated bacterial communities and their functions in the irrigation water from wetland taro farms on Oahu, Hawaii using different DNA sequencing platforms. Irrigation water samples (stream, spring, and tank stored water) were collected from North, East, and West sides of Oahu and subjected to high quality DNA isolation, library preparation and sequencing of the V3-V4 region, full length 16S rRNA, and shotgun metagenome sequencing using Illumina iSeq100, Oxford Nanopore MinION and Illumina NovaSeq, respectively. Illumina reads provided the most comprehensive taxonomic classification at the phylum level where Proteobacteria was identified as the most abundant phyla in river stream source and associated wet taro field water samples. Cyanobacteria was also a dominant phylum from tank and spring water, whereas Bacteroidetes were most abundant in wetland taro fields irrigated with spring water. However, over 50% of the valid short amplicon reads remained unclassified and inconclusive at the species level. Whereas samples sequenced for full length 16S rRNA and shotgun metagenome, clearly illustrated that Oxford Nanopore MinION is a better choice to classify the microbes to the genus and species levels. In terms of functional analyses, only 12% of the genes were shared by two consortia. Total 95 antibiotic resistant genes (ARGs) were detected with variable relative abundance. Description of microbial communities and their functions are essential for the development of better water management strategies to produce safer fresh produce and to protect plant, animal, human and environmental health. This project identified analytical tools to study microbiome of irrigation water.

microbiology↗

bioRxiv2022

An atypical F-actin capping protein modulates cytoskeleton behaviors crucial to colonization of Trichomonas vaginalis

Cytoadherence and consequential migration are crucial for pathogens to establish colonization in the host. In contrast to the nonadherent isolate of Trichomonas vaginalis, the adherent one expresses more actin-related machinery proteins with more active flagellate-amoeboid morphogenesis, amoeba migration, and cytoadherence, activities that were abrogated by an actin assembly blocker. By immunoprecipitation coupled with label-free quantitative proteomics, an F-actin capping protein (TvFACP) was identified from the actin-centric interactome, with an atypically greater binding preference to G-actin than F-actin. TvFACP partially colocalized with F-actin at the parasite pseudopodia protrusion and formed the protein complexes with -actin through its c-terminal domain. Meanwhile, TvFACP overexpression suppresses F-actin polymerization, amoeboid morphogenesis, and cytoadherence in this parasite. Ser2 phosphorylation of TvFACP enriched in the amoeboid stage of adhered trophozoites was reduced by a CKII inhibitor. The site-directed mutagenesis and CKII inhibitor treatment revealed that Ser2 phosphorylation acts as a switching signal to alter TvFACP actin-binding activity and consequent actin cytoskeleton behaviors. Through CKII signaling, TvFACP also controls the conversion of adherent trophozoite from amoeboid migration to flagellate form with axonemal motility. Together, CKII-dependent Ser2 phosphorylation regulates TvFACP binding actin to fine-tune cytoskeleton dynamics and drive crucial behaviors underlying host colonization of T. vaginalis.

microbiology↗

bioRxiv2020

Proximity Biotin Labeling Reveals KSHV Interferon Regulatory Factor Networks

Studies on "HIT&RUN" effects by viral protein are difficult when using traditional affinity precipitation-based techniques under dynamic conditions, because only proteins interacting at a specific instance in time can be precipitated by affinity purification. Recent advances in proximity labeling (PL) have enabled study of both static and dynamic protein-protein interactions. Here we applied PL method with recombinant Kaposis sarcoma-associated herpesvirus (KSHV). KSHV, a gamma-herpesvirus, uniquely encodes four interferon regulatory factors (IRFs 1-4) in the genome, and we identified KSHV vIRF-1 and vIRF-4 interacting proteins during reactivation. Fusion of mini-TurboID with vIRF-1 or vIRF-4 did not interfere with KSHV gene expression, DNA replication, or de novo infections. PL identified 213 and 70 proteins for vIRF-1 and vIRF-4 respectively, which possibly interact during KSHV reactivation, and 47 of those were shared between the two vIRFs; the list also includes three viral proteins, ORF17, thymidine kinase, and vIRF-4. Functional annotation of respective interacting proteins showed highly overlapping biological functions such as mRNA processing and transcriptional regulation by TP53. Involvement of commonly interacting 44 cellular proteins in innate immune regulation were examined by siRNAs, and we identified that splicing factor 3B (SF3B) family proteins were clearly involved in interferons transcription and suppressed KSHV reactivation. We propose that recombinant TurboID-KSHV is a powerful tool to probe key cellular proteins that play a role in KSHV replication, and selective splicing factors may have a function beyond connecting two exon sequences to regulate innate immune responses. ImportanceViral protein interaction with a host protein shows at least two sides: (i) taking host protein functions for its own benefit and (ii) disruption of existing host protein complex formation to inhibit undesirable host responses. Due to use of affinity-precipitation approaches, the majority of our studies focused on how the virus takes advantage of the newly-formed protein interactions for its own replication. Proximity labeling (PL) however, can also highlight the transient and negative effects - those interactions which lead to dissociation from the existing protein complex. Here we highlight the power of PL in combination with recombinant KSHV to study viral host interactions.

microbiology↗