Biology subjects

Greathouse, K. L.

Publications and source records attributed to Greathouse, K. L..

6 recordsLinked to original sources

bioRxiv2025

Outer membrane vesicles from Bacteroides fragilis contain coding and non-coding small RNA species that modulate inflammatory signaling in intestinal epithelial cells.

Alterations to the community structure and function of the microbiome are associated with changes to host physiology, including immune responses. However, the contribution of microbe-derived RNAs carried by outer membrane vesicles (OMVs) to host immune responses remain unclear. This study investigated the role of OMVs and OMV-associated small RNA (sRNA) species from pathogenic and commensal Bacteroides fragilis (ETBF and NTBF respectively) in eliciting different immune responses from intestinal epithelial cells. To distinguish the differences in the sRNA profiles of the two strains and their OMVs, RNA-seq, qRT-PCR, and northern blotting were conducted to identify enrichment of discrete sRNA species in OMVs, which were also differentially expressed between the two strains. Specifically, both coding and non-coding RNAs were enriched in OMVs from NTBF and ETBF, with BF9343_RS22680 and BF9343_RS17870 being significantly enriched in ETBF OMVs compared to NTBF. To understand the effects of OMVs on pattern recognition receptors, reporter cells of Toll-like receptor (TLR) activation were treated with OMVs, demonstrating activation of TLRs 2, 3, and 7. Treatment of Caco-2 and HT29-MTX cells with OMVs demonstrated increased expression of IL-8. Surprisingly, we discovered that degradation of RNase-accessible RNAs within ETBF OMVs, but not NTBF OMVs, resulted in vesicles with enhanced capacity to stimulate IL-8 expression, indicating that these extravesicular RNAs exert an immunosuppressive effect. This suggests a dual role for OMV-associated RNAs in modulating host immune responses, with implications for both bacterial pathogenesis and therapeutic applications. Graphical Abstract

molecular biology↗

bioRxiv2024

Targeted therapy of oncomicrobe F. nucleatum with bioengineered probiotic expressing guided antimicrobial peptide (gAMP)

Colorectal cancer (CRC) is a leading cause of cancer-related mortality, with Fusobacterium nucleatum (F. nucleatum) identified as a key contributor to its progression. This study explores a novel targeted therapy using bioengineered probiotics expressing guided antimicrobial peptides (gAMPs) to selectively inhibit F. nucleatum. We engineered Lactococcus lactis MG1363 to express gAMPs derived from Ovispirin and Cathelin-related peptide SCF, linked to a Statherin-derived guide peptide (YQPVPE) that binds specifically to the F. nucleatum membrane porin FomA. Our results demonstrate that the Statherin-derived guide peptide enhances the binding affinity to F. nucleatum, significantly increasing the preferential attachment compared to control peptides. In vitro assays revealed that both unguided and guided AMPs effectively inhibited biofilm formation in F. nucleatum, with gAMPs showing reduced toxicity against non-target bacteria (Bacteroides fragilis and Escherichia coli). The gAMPs were also more effective in modulating growth kinetics, exhibiting selective toxicity towards F. nucleatum at lower concentrations. Co-culture experiments in a simulated human gut microbiome demonstrated that the gAMP probiotic maintained microbial diversity while effectively reducing F. nucleatum abundance. Quantitative PCR and 16S rRNA sequencing confirmed that gAMP treatment preserved the richness of the microbiota, contrasting with significant dysbiosis observed in control samples. These findings support the potential of engineered probiotics as a targeted therapeutic approach to combat F. nucleatum-associated CRC. By leveraging the specificity of Statherin-derived peptides, this strategy not only addresses the pathogenicity of F. nucleatum but also mitigates the adverse effects of traditional antimicrobial therapies on beneficial gut microbiota. Future studies will explore the clinical applicability of this approach in CRC management and its impact on overall gut health.

microbiology↗

bioRxiv2023

Co-enrichment of cancer-associated bacterial taxa is correlated with immune cell infiltrates in esophageal tumor tissue

Esophageal carcinoma (ESCA) is a leading cause of cancer-related death worldwide, and Barretts esophagus (BE) is a strong risk factor along with smoking. Smoking is well-known to induce microbial dysbiosis and we asked if BE and esophageal microbiomes had shared microbial alterations that could provide novel biomarkers. We extracted DNA from BE tissues (n=5) and tumors of 158 patients in the NCI-MD case control study and sequenced the 16S rRNA gene (V3-4), with TCGA ESCA RNAseq (n = 173) and WGS (n = 139) non-human reads used as validation. We identified four taxa, Campylobacter, Prevotella, Streptococcus, and Fusobacterium as highly enriched in esophageal cancer across all cohorts. Using SparCC, we discovered that Fusobacterium and Prevotella were also co-enriched across all cohorts. We then analyzed immune cell infiltration to determine if these dysbiotic taxa were associated with immune signatures. Using xCell to obtain predicted immune infiltrates, we identified a depletion of megakaryocyte-erythroid progenitor (MEP) cells in tumors with presence of any of the four taxa, along with enrichment of platelets in tumors with Campylobactor or Fusobacterium. Taken together, our results suggest that intratumoral presence of these co-occurring bacterial genera may confer tumor promoting immune alternations that allow disease progression in esophageal cancer.

cancer biology↗

bioRxiv2022

Acidovorax temperans polarizes T17 cells and skews neutrophil maturation to promote lung adenocarcinoma development

Dysbiosis, or changes within the microbiome, is a common feature of solid tumors, however whether this dysbiosis directly contributes to tumor development is largely unknown. We previously characterized the lung cancer microbiome and identified Acidovorax temperans as enriched in tumors. In this study, we instilled A. temperans in an animal model driven by mutant Kras and Tp53 alleles. This revealed A. temperans accelerates tumor development and burden through infiltration of proinflammatory cells. Neutrophils exposed to A. temperans displayed a mature, pro-tumorigenic genotype with increased cytokine signaling, with a global shift away from IL-1{beta} signaling. Neutrophil to monocyte and macrophage signaling upregulated MHC II to activate CD4+ T cells which polarized to an IL-17A+ phenotype detectable in CD4+ and {gamma}{delta} populations. T17 cells shared a common gene expression program predictive of poor survival in human LUAD. These data indicate dysbiosis promotes tumor growth by modulating inflammation.

cancer biology↗

bioRxiv2018

Gut microbiome meta-analysis reveals dysbiosis is independent of body mass index in predicting risk of obesity-associated CRC

Obesity is a risk factor for colorectal cancer (CRC), accounting for more than 14% of CRC incidence. Microbial dysbiosis and chronic inflammation are common characteristics in both obesity and CRC. Human and murine studies, together, demonstrate the significant impact of the microbiome on governing energy metabolism and CRC development; yet, little is understood about the contribution of the microbiome to development of obesity-associated CRC as compared to non-obese individuals. In this study, we conducted a meta-analysis using five publicly available stool and tissue-based 16S rRNA and whole genome sequencing (WGS) data sets of CRC microbiome studies. High-resolution analysis was employed for 16S rRNA data using Resphera Insight, which allowed us to achieve species-level information to compare with WGS. Characterization of the confounders between studies, 16S rRNA variable region, and sequencing method, did not reveal any significant effect on alpha diversity in CRC prediction. Both 16S rRNA and WGS were equally variable in their ability to predict CRC. Results from community structure and composition analysis confirmed lower diversity in obese individuals without CRC; however, no universal differences were found in diversity between obese and non-obese individuals with CRC. When examining taxonomic differences, the probability of being classified as CRC did not change significantly in obese individuals for all taxa tested. However, random forest classification was able to distinguish CRC and non-CRC stool when body mass index was added to the model. Overall, microbial dysbiosis was not a significant factor in explaining the higher risk of colon cancer among individuals with obesity.

cancer biology↗

bioRxiv2018

Microbiome-TP53 Gene Interaction in Human Lung Cancer

BackgroundLung cancer is the leading cancer diagnosis worldwide and the number one cause of cancer deaths. Exposure to cigarette smoke, the primary risk factor in lung cancer, reduces epithelial barrier integrity and increases susceptibility to infections. Herein, we hypothesized that somatic mutations together with cigarette smoke generate a dysbiotic microbiota that is associated with lung carcinogenesis. Using lung tissue from controls (n=33) and cancer cases (n=143), we conducted 16S rRNA bacterial gene sequencing, with RNA-seq data from lung cancer cases in The Cancer Genome Atlas (n=1112) serving as the validation cohort.\n\nResultsOverall, we demonstrate a lower alpha diversity in normal lung as compared to non-tumor adjacent or tumor tissue. In squamous cell carcinoma (SCC) specifically, a separate group of taxa were identified, in which Acidovorax was enriched in smokers (P =0.0013). Acidovorax temporans was identified by fluorescent in situ hybridization within tumor sections, and confirmed by two separate 16S rRNA strategies. Further, these taxa, including Acidovorax, exhibited higher abundance among the subset of SCC cases with TP53 mutations, an association not seen in adenocarcinomas (AD).\n\nConclusionsThe results of this comprehensive study show both a microbiome-gene and microbiome-exposure interactions in SCC lung cancer tissue. Specifically, tumors harboring TP53 mutations, which can damage epithelial function, have a unique bacterial consortia which is higher in relative abundance in smoking-associated SCC. Given the significant need for clinical diagnostic tools in lung cancer, this study may provide novel biomarkers for early detection.

cancer biology↗