If we looked a protein as a human, we can also observed the birth, illness, death. Classical biochemistry approaches analysis one single protein life cycle. Here, we combine measurements of protein production and degradation and mRNA dynamics to build a quantitative genomic model of the differential regulation of gene expression in lipopolysaccharide-stimulated mouse dendritic cells.
To monitor protein production and degradation during a dynamic response, we used a modified pulsed-SILAC (stable isotope labeling with amino acids in cell culture) approach. Newly synthesized proteins were thus labeled with heavy amino acids, serving as a proxy for protein synthesis, whereas proteins with medium heavy amino acids decayed over time, reflecting cellular half-lives.
We quantified 6079 proteins by means of liquid chromatography–tandem mass spectrometry (LC-MS/MS) in at least one sample and 2288 proteins in all samples.
Next, we determined the contribution of each regulatory step to protein fold changes at 12 hours. We determined the contribution of changes in mRNA levels, protein synthesis, and protein degradation rates during a dynamic response and found that changes in mRNA levels dominate relative fold changes.
In the future, Enhanced methods will help to identify new key regulators of these responses.