Identification of the critical regulators of protein synthesis and degradation in human muscle atrophy

Dr. Lionel Tintignac and Dr. Nitish Mittal, University of Basel


The imbalance between protein synthesis and protein degradation is a common pathogenic mechanism underlying muscle diseases. A higher rate of protein synthesis than the rate of protein degradation leads to hypertrophy, while a relatively lower rate of protein synthesis leads to atrophy. Understanding the regulators of these processes is a prerequisite for the development of therapeutic strategies to counteract atrophy. Although the vast majority of studies have focused on deciphering degradation processes that contribute to acute muscle breakdown (wasting), it is only recently that the proper functioning of the protein synthesis machinery in atrophying muscles has started to be questioned and investigated. We previously established that upon atrophy induction protein degradation directly targets protein synthesis. More recently, we identified an opposite link as well, demonstrating that increased protein synthesis initiation through sustained activation of mTORC1 in muscle (TSCmKO mouse model) does not leads to hypertrophy, but rather to activation of the unfolding protein response pathway. To follow up on these observations we have recently established a unique combination of cutting-edge techniques in our laboratories. We here proposed a 2-year project aiming to identify the molecular networks that specifically control protein synthesis and degradation in human myofibers under physiological conditions, and that are perturbed upon atrophic stress. Our strategy will be to integrate genome-wide measurements of 1) transcript levels (RNA-Seq), 2) translation levels (sequencing of Ribosome protected fragments, RPFs) and 3) protein levels (proteomics) from human myofibers. The ribosome footprinting technique will provide us, for the first time, with estimates of protein synthesis rates at individual RNA resolution, thus allowing us to further uncover the impact of these regulatory factors on translation of each gene. We will further apply a powerful technique that we have recently implemented, known as BioID, to map the direct interactome of the two main players we have already identified in atrophy. For this, we will engineer human myoblast cell lines to express endogenously chimeric BioID proteins. This will allow us to determine the direct interactomes of these proteins, as well as to investigate the translation process at extremely high resolution, because we will be able to isolate tagged ribosomes due to their interaction with our BioID chimeras and compare the pool of mRNAs interacting with these ribosomes with that of all ribosome-bound translated mRNAs. With this project we will define the mechanisms by which protein degradation and synthesis dysregulation promote muscle atrophy, which will open new avenues for drug discovery to counteract pathological muscle proteolysis and wasting.