Biology subjects

McComb, J.

Publications and source records attributed to McComb, J..

4 recordsLinked to original sources

bioRxiv2025

Decoding Cellular Stress States for Toxicology Using Single-Cell Transcriptomics

We applied the TempO-LINC(R) platform to generate single-cell transcriptomic (SCTr) profiles of [~]40,000 HepaRG cells exposed to etoposide, brefeldin A, cycloheximide, rotenone, tBHQ, troglitazone, and tunicamycin at three concentrations for 24 hours. SCTr enabled a detailed analysis of adaptive stress response pathways (SRPs), including the unfolded protein response (UPR), oxidative stress response (OSR), heat shock response (HSR), and DNA damage response (DDR). Troglitazone upregulated lipid metabolism genes (PLIN2, ACOX1) along with HSR and UPR activation, with co-expression of DNAJA1, HSP90AA1, and DDIT3 in subsets of cells. Brefeldin A and tunicamycin strongly induced UPR markers (HSPA5, SYVN1, LMF2, PDIA4) in subsets of cells, with some also expressing apoptotic (DDIT3, CASP8) and autophagic (SQSTM1) genes, indicating diverse stress responses. Rotenone activated GDF15, TRIB3, and DDIT3 in a fraction of cells, accompanied by PLIN2 and mild UPR induction, reflecting heterogeneous mitochondrial stress responses. We scored individual cells using literature-derived SRP gene signatures to characterize overall stress phenotypes and clustered them using a generalized Jaccard metric. The clustering revealed five phenotypic groups spanning cell states associated with homeostasis, adaptive responses, terminal outcomes, autophagy, and apoptosis. By systematically analyzing the distributions of cells in different states across treatments, we visualized dynamic shifts in cellular subpopulations responding to chemicals, revealing early stress responses and potential transitions to cell death. Our findings suggest the utility of SCTr in decoding stress states that could provide possible insights into transitions between cellular adaptive and terminal transitions involved in toxicity.

pharmacology and toxicology↗

bioRxiv2024

High-throughput gene expression analysis with TempO-LINC sensitively resolves complex brain, lung and kidney heterogeneity at single-cell resolution

We report the development and performance of a novel genomics platform, TempO-LINC, for conducting high-throughput transcriptomic analysis on single cells and nuclei. TempO-LINC works by adding cell-identifying molecular barcodes onto highly selective and high-sensitivity gene expression probes within fixed cells, without having to first generate cDNA. Using an instrument-free combinatorial-indexing approach, all probes within the same fixed cell receive an identical barcode, enabling the reconstruction of single-cell gene expression profiles across as few as several hundred cells and up to 100,000+ cells per run. The TempO-LINC approach is easily scalable based on the number of barcodes and rounds of barcoding performed; however, for the experiments reported in this study, the assay utilized over 5.3 million unique barcodes. TempO-LINC has a robust protocol for fixing and banking cells and displays high-sensitivity gene detection from multiple diverse sample types. We show that TempO-LINC has an observed multiplet rate of less than 1.1% and a cell capture rate of [~]50%. Although the assay can accurately profile the whole transcriptome (19,683 human or 21,400 mouse genes), it can be targeted to measure only actionable/informative genes and molecular pathways of interest - thereby reducing sequencing requirements. In this study, we applied TempO-LINC to profile the transcriptomes of 89,722 cells across multiple sample types, including nuclei from mouse lung, kidney and brain tissues. The data demonstrated the ability to identify and annotate at least 50 unique cell populations and positively correlate expression of cell type-specific molecular markers within them. TempO-LINC is a robust new single-cell technology that is ideal for large-scale applications/studies across thousands of samples with high data quality.

genomics↗

bioRxiv2022

IRF7 impacts on prostate cancer cell survival in response to radiation

Understanding the impact of radiotherapy on the evolution of treatment resistant prostate cancer is critical for selecting effective treatment combinations. Whilst activation of Type 1 interferon signalling is a hallmark of how cells respond to viral infection, in cancer cells, multiple stresses are known to activate this same response. In this study we have evaluated for the first time the changes in the interferon response induced by culturing prostate cancer cells under sphere- forming conditions and following irradiation. We report a conserved upregulated transcript profile for both conditions that is strongly associated with therapeutic resistance and cell survival in vitro and in vivo. The profile includes and is regulated by the Type 1 interferon master regulator IRF7 which, when depleted, delays tumour re-growth following irradiation. We immuno-stained two independent prostate cohorts for IRF7 and found that increased expression, particularly in cases with low PTEN expression, correlated with poor prognosis. To more comprehensively characterise the impact of IRF7 and radiation on cells, RNA-Seq was performed on IRF7 knockdown cells at different radiation doses. We identified a number of biological processes that were IRF7-dependent, including the formation of stem-like cell populations and also therapeutic vulnerabilities. For example, irradiation sensitised surviving cells to either a combination of an IKK{varepsilon}/TBK1 and a MEK inhibitor or treatment with an inhibitor of IDO1, an IRF7- dependent gene. Translationally our work suggests that IRF7 expression can be used to stratify patients who may not benefit from receiving radiotherapy alone but rather may benefit from treatment combinations. In two cohorts treated with radical intent, strong IRF7 staining was associated with disease-specific death implicating this pathway as a convergence point for therapeutic resistance in prostate and potentially other cancer types.

cancer biology↗

bioRxiv2022

Soil amendments and suppresssion of Phytophthora root rot in avocado (Persea indica)

The ability of microbial or mineral-based soil additives to suppress root rot caused by Phytophthora cinnamomi was assessed. Phosphite and metalaxyl treatments for the control of disease, and glyphosate for weed control were also assessed. A treatment simulating avocado orchard conditions had chicken manure, wood mulch, and mulch from beneath trees in an avocado orchard added to the pots. Soil treatments (three probiotic and two mineral-based) were applied to 9-month-old saplings growing in containers in a glasshouse. After one-month, half of the plants of each treatment were inoculated with the pathogen. Three months after inoculation, plants were harvested and plant growth and root damage were measured. In the first experiment infestation with P. cinnamomi significantly reduced fine root dry weight in all plants except those in soil treated with one silicon-based mineral mulch. Visible root damage was higher in plants treated with probiotics. In this experiment, and in a repeat experiment the reduction of fine root damage achieved by spraying plants with phosphite or addition of a silica based mineral mulch was similar. Phosphite was preferable to metalaxyl as a chemical treatment, as the latter reduced shoot and root growth of non-infected plants. Glyphosate treatment of wheat seedlings growing in the pots with the avocados reduced shoot and fine root growth of both non-infected and infected plants. These observations need to be confirmed under field conditions.

plant biology↗