Impact of the mitogen‐activated protein kinase pathway on the subproteome of detergent‐resistant microdomains of colon carcinoma cells

Abstract

Lipid rafts play a key role in the regulation of fundamentally important cellular processes, including cell proliferation, differentiation, and survival. The composition of such detergent-resistant microdomains (DRMs) is altered under pathologic conditions, including cancer. Although DRMs have been analyzed in colorectal carcinoma little information exists about their composition upon treatment with targeted drugs. Hence, a quantitative proteomic profiling approach was performed to define alterations within the DRM fraction of colorectal carcinoma cells upon treatment with the drug U0126, an inhibitor of the mitogen-activated protein kinase pathway. Comparative expression profilings resulted in the identification of 300 proteins, which could partially be linked to key oncogenic signaling pathways and tumor-related cellular features, such as cell proliferation, adhesion, motility, invasion, and apoptosis resistance. Most of these proteins were downregulated upon inhibitor treatment. In addition, quantitative proteomic profilings of cholesterol-depleted versus intact lipid rafts were performed to define, which U0126-regulated target structures represent bona fide raft proteins. Selected differentially abundant raft proteins were validated at the mRNA and/or protein level using U0126- or Trametinib-treated cells. The presented data provide insights into the molecular mechanisms associated with the response to the treatment with MEK inhibitors and might also lead to novel candidates for therapeutic interventions.

Dances with Membranes: Breakthroughs from Super-resolution Imaging

“…Many experimental approaches are controversial in their ability to distinguish membrane domains. For example, cold low density detergent-insoluble fractions of lysed cell membranes have been extensively used as a determinant of lipid rafts (from ( Brown & Rose, 1992 ) to ( Recktenwald et al., 2015 )), however their representation of domains in native cell membranes is tenuous and yet to be verified ( Edidin, 2003 , Kraft, 2013 , Lichtenberg et al., 2005 ).…”

Comparative membrane proteomics: a technical advancement in the search of renal cell carcinoma biomarkers

“…[11][12][13] Recent data on the quantitative proteomics of MD -lipid rafts or caveolaehave underlined the puzzling role of various signaling proteins in cancer development, but these studies were mainly performed on neoplastic cell lines. [14][15][16][17] However, procedures commonly used for cell cultures, such as metabolic labelling, are not suitable for the study of human tissue MD, making label-free quantification a valid choice to achieve differential expression profiles. Moreover, the protocol needs to be adapted to small amount of samples, as feasible when working with human surgical specimens.…”