Biology subjects

Mitchell, T. W.

Publications and source records attributed to Mitchell, T. W..

5 recordsLinked to original sources

bioRxiv2024

Targeting de novo lipogenesis improves gemcitabine efficacy in pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease with few treatment options and poor survivability. In this work we sought to characterise metabolic adaptations to gemcitabine (GEMC)-based chemotherapy exposure to discover new therapeutic targets for improving treatment efficacy. We show that GEMC resistance (GEMR) upregulates de novo lipogenesis in Panc1 and MiaPaCa2 cells through increased activity and expression of acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD1). We also discovered alternate fatty acid desaturase 2 (FADS2) activity in Panc1 cells, which led to the production of sapienic acid (FA 16:1n-10, cis) from palmitic acid (FA 16:0). Knockdown of key lipid synthesis enzymes sensitised cells to GEMC treatment, with FAS (both cell lines), SCD1 (MiaPaCa2 only) and SCD1+FADS2 (Panc1) knockdown showing the greatest reduction in cell growth when combined with GEMC treatment. In Panc1 cells, both desaturases upregulated their activity when the alternate was knocked down, necessitating the need for dual desaturase knockdown in this cell line. PDAC cells attenuated to grow in combination GEMC/paclitaxel (CombAT) also displayed enhanced de novo lipogenesis; however, combination chemotherapy significantly downregulated FADS2 expression and activity in Panc1 CombAT cells rendering them more sensitive to SCD1 knockdown. We conclude that co-targeting lipid synthesis in PDAC could be a viable strategy for improving the efficacy of both GEMC monotherapy and combination GEMC/PTX therapy.

cancer biology↗

bioRxiv2024

Temperature adaptation of yeast phospholipid molecular species at the acyl chain positional level

The budding yeast Saccharomyces cerevisiae is a poikilothermic organism and adapts its lipid composition to the environmental temperature to maintain membrane physical properties. Studies addressing temperature-dependent adaptation of the lipidome in yeast have described changes in the phospholipid composition at the level of sum composition (e.g. PC 32:1) and molecular composition (e.g. PC 16:0_16:1). However, to date, there is no information at the level of positional isomers (e.g. PC 16:0/16:1 versus PC 16:1/16:0). In this study, combined Collision- and Ozone-Induced Dissociation (CID/OzID) mass spectrometry was deployed to investigate homeoviscous adaptation of PC, PE, and PS sn-molecular species composition. We determined the main species to be 16:1/16:1, 16:0/16:1, 16:1/18:1, 16:0/18:1, and 18:0/16:1. In general, at higher culture temperature, the sn-1 position is increased in saturated acyl chains, whereas the sn-2 position mainly is increased in acyl chain length. PC mainly increases in 16:0/16:1 and 16:0/18:1, at the expense of 16:1/16:1, whereas PS and PE increase in 16:1/18:1, at the expense of 16:1/16:1 and 16:0/16:1. Our data suggest distinct adaptation mechanisms of the sn-1 and sn-2 acyl chains, and different manners of sn-molecular species adaptation between PC and PE/PS.

biochemistry↗

bioRxiv2023

A High-Throughput Data-Independent Acquisition Workflow for Deep Characterisation of the sn-Isomer Lipidome

We report a workflow based on ozone-induced dissociation for untargeted characterization of hundreds of sn-resolved glycerophospholipid isomers from biological extracts in under 20 minutes, coupled with an automated data analysis pipeline. It provides an order of magnitude increase in the number of sn-isomer pairs identified compared to previous reports, reveals that sn-isomer populations are tightly regulated and significantly different between cell lines, and enables identification of rare lipids containing ultra-long chain monounsaturated acyl chains.

cell biology↗

bioRxiv2022

High-throughput detection and quantification of phosphatidylcholines and sphingomyelins from single cells by chip-based nanoelectrospray ionisation

Recent advances in single-cell genomics and transcriptomics technologies have transformed our understanding of cellular heterogeneity in growth, development, ageing and disease; however, methods for single-cell lipidomics have comparatively lagged behind in development. We have developed a high-throughput method for the detection and quantification of a wide range of phosphatidylcholine (PC) and sphingomyelin (SM) species from single cells that combines fluorescence-assisted cell sorting (FACS) with automated chip-based nanoelectrospray ionization (nanoESI) and shotgun lipidomics. We show herein that our method is capable of quantifying more than 50 different PC and SM species from single cells and can easily distinguish between cells of different lineages or cells treated with exogenous fatty acids. Moreover, our method can detect more subtle differences in the lipidome between cell lines of the same cancer type. Our approach can be run in parallel with other single-cell technologies to deliver near-complete multi-omics data on cells with a similar phenotype and has the capacity to significantly advance our current knowledge on cellular heterogeneity.

biochemistry↗

bioRxiv2021

Analysis of sex-specific lipid metabolism in P. falciparum gametocytes points to importance of sphingomyelin for gametocytogenesis

Male and female Plasmodium falciparum gametocytes are the parasite lifecycle stage responsible for transmission of malaria from the human host to mosquito vector. Not only are gametocytes able to survive in radically different host environments, but they are also precursors for male and female gametes that reproduce sexually soon after ingestion by the mosquito. Here we investigate the sex-specific lipid metabolism of gametocytes within their host red blood cell and poised for ingestion by the mosquito vector and subsequent sexual reproduction. Comparison of the male and female lipidome identifies cholesteryl esters and dihydrosphingomyelin enrichment in female gametocytes. Chemical inhibition of each of these lipid types in mature gametocytes suggests dihydrosphingomyelin synthesis but not cholesteryl ester synthesis is important for sex-specific gametocyte viability. Genetic disruption of each of the two sphingomyelin synthase gene points towards sphingomyelin synthesis contributing to gametocytogenesis. This study shows that gametocytes are not only distinct from asexual stages, but that the lipid composition is also vastly different between male and female gametocytes, reflecting the different cellular roles these stages play. Together our results highlight the sex-specific nature of gametocyte lipid metabolism that has the potential to be targeted to block malaria transmission.

microbiology↗