Transcriptional and Metabolic Investigation in 5′-Nucleotidase Deficient Cancer Cell Lines

Abstract

Enzymes of nucleoside and nucleotide metabolism regulate important cellular processes with potential impacts on nucleotide-unrelated parameters. We have used a set of CRISPR/Cas9-modified cell models expressing both, one, or none of the 5′-nucleotidases cN-II and CD73, together with RNA sequencing and targeted metabolomics, to decipher new regulatory roles of these proteins. We observed important transcriptional modifications between models as well as upon exposure to adenosine. Metabolite content varied differently between cell models in response to adenosine exposure but was rather similar in control conditions. Our original cell models allowed us to identify a new unobvious link between proteins in the nucleotide metabolism and other cellular pathways. Further analyses of our models, including additional experiments, could help us to better understand some of the roles played by these enzymes.

A Proposed Methodology to Deal with the Impact of In Vitro Cellular Matrix on the Analytical Performances of a Targeted Metabolomic LC-HRMS Method

“…Although the ME is not considered, the main drawback of this approach is that it is necessary to adapt calibration curves for each metabolite studied to the area of expected concentration. Another approach is based on the peak area ratio of a metabolite and its corresponding IS [ 23 , 24 ]. This method was selected in the present work.…”