Coordination of protein synthesis and degradation in mammalian cells

The cellular protein levels are determined by protein synthesis and turnover rates. Two processes are involved in the proteome's turnover in proliferating cells: protein degradation and dilution. In theory, maintaining the cellular proteome concentration, which is imperative to proper cellular function, requires the coordination between protein synthesis and turnover. Although the relationship between protein synthesis, degradation, and dilution has been studied in bacteria and yeasts, little is known about how mammalian cells can balance these rates to uphold proteostasis. The peculiarity of the mammalian cells is that the dilution rate, the main counterpart to protein synthesis in bacteria or yeasts, can be highly restrictive in many cases, giving greater necessity to the coordination between synthesis and degradation. Despite evidence to support the existence of such coordination, there is little information about the mechanism behind such cooperation. Previous studies that attempted to disentangle the relationship between protein synthesis and degradation relied on using high concentrations of inhibitors that irreversible disrupt the equilibrium proteostasis.

The thesis addressed these questions by creating reversible static or dynamic proteostasis states where the protein synthesis rate was manipulated. Meanwhile, the dynamics in protein synthesis, degradation, and dilution were quantified with high temporal resolution. The tool that enables the measurements is the tandem fluorescent timer (tFT, or Timer) reporter, engineered to measure both ubiquitin-dependent and ubiquitin-independent degradation rates by the proteasome. Complementary to the tFT measurements was a superstatistical Bayesian inference algorithm employed to calculate timer evolving the synthesis and degradation rates from the live cell time-lapse microscopy.

We discovered that the change in protein synthesis was counterbalanced by the change in both protein degradation and dilution, which buffered the protein concentration but could not completely compensate. Such counteracting of degradation towards synthesis was also observed in non-dividing cells. The recapitulation of protein degradation adaption dynamics revealed a delay in the action of degradation machinery in a time range of several hours. We discussed the effect of change in degradation and dilution on proteome stoichiometry. The transcriptome and in vitro assays further concluded that neither the differential expression nor proteasome content could fully explain the change in the degradation adaption. Finally, we proposed a passive adaption model to explain the degradation adaption to synthesis, which matched the observation from our Timer reporter.