PUNCH-P for global translatome profiling

In recent years, regulation of mRNA translation has attracted increasing attention as an important determinant of gene expression in health and disease. Such regulation can result in different translation rates for mRNAs of similar abundance; therefore, estimates of steady-state mRNA levels by methods such as microarray or deep-sequencing (RNA-seq) may fail to capture critical aspects of gene expression. Along with advances in system-wide RNA and protein analysis platforms, several methods were developed to better understand the complex regulation of translation. While these methods were successfully employed to study many biological questions, a need still remained for a simple, low-cost technique that would allow direct measurement of system-wide protein synthesis in cells and whole tissues. For this purpose, we developed a novel approach termed PUromycin-associated Nascent CHain Proteomics (PUNCH-P), which directly monitors nascent polypeptide chains by mass-spectrometry without the need for prior metabolic labeling.