Correlated fluorescence microscopy and multi-ion beam secondary ion mass spectrometry imaging reveals phosphatidylethanolamine increases in the membrane of cancer cells over-expressing the molecular chaperone subunit CCT\u03b4

John S Fletcher,Sanna Sämfors,Julie Grantham,Josefine Vallin,Andreas Svanström

doi:10.1007/s00216-020-03013-9

Abstract

Changes in the membrane composition of sub-populations of cells can influence different properties with importance to tumour growth, metastasis and treatment efficacy. In this study, we use correlated fluorescence microscopy and ToF-SIMS with C60+ and (CO2)6k+ ion beams to identify and characterise sub-populations of cells based on successful transfection leading to over-expression of CCTδ, a component of the multi-subunit molecular chaperone named chaperonin-containing tailless complex polypeptide 1 (CCT). CCT has been linked to increased cell growth and proliferation and is known to affect cell morphology but corresponding changes in lipid composition of the membrane have not been measured until now. Multivariate analysis of the surface mass spectra from single cells, focused on the intact lipid ions, indicates an enrichment of phosphatidylethanolamine species in the transfected cells. While the lipid changes in this case are driven by the structural changes in the protein cytoskeleton, the consequence of phosphatidylethanolamine enrichment may have additional implications in cancer such as increased membrane fluidity, increased motility and an ability to adapt to a depletion of unsaturated lipids during cancer cell proliferation. This study demonstrates a successful fluorescence microscopy-guided cell by cell membrane lipid analysis with broad application to biological investigation.Graphical abstract

Highlights

Inspection of the cells by fluorescent microscopy indicated the successful transfection of approximately 70% of the cells based on the fluorescent signal visible from the expression of GFP-CCTδ (160 cells were counted from a total of four fields of view)
The increase in PE lipids in the transfected cells can be explained based on the conical shape of the PE lipid, with its relatively small head group, which facilitates the formation of highly curved structures and results in an increase in membrane fluidity
In the case of the GFP-CCTδ transfection, it is expected that this change in lipid composition can facilitate the phenotypic changes in the cell such as the formation of tubulated membrane structures

Summary

Introduction

While different organs within an organism comprise a range of different cell types with specific function, there is increasing interest in the diversity within the population of each cell type. Phenotypic changes, often with a change in metabolic/lipid profile, can occur within a population of cells leading to disfunction or, in the case of cancer, altering the potential response to treatment or the ability of the cell to infiltrate surrounding tissue leading to metastasis [1]. The plasma membrane, together with the underlying membrane-associated cytoskeletal networks, acts as a sensor for responding to extracellular cues, for example from growth factors. Numerous processes such as endocytosis, exocytosis and filopodia growth involve the dynamic formation of high curvature regions within the plasma membrane at specific locations. Curvature of the plasma membrane is a consequence of lipid composition, occurrence of curvature-inducing lipid-binding proteins, protein insertion into the lipid bilayer and cytoskeletoninduced tension [2]

Methods

Results

Conclusion