Protein aggregation is mostly viewed as deleterious and irreversible causing several pathologies. However, reversible protein aggregation has recently emerged as a novel concept for cellular regulation. Here, we characterize stress-induced, reversible aggregation of yeast pyruvate kinase, Cdc19. Aggregation of Cdc19 is regulated by oligomerization and binding to allosteric regulators. We identify a region of low compositional complexity (LCR) within Cdc19 as necessary and sufficient for reversible aggregation. During exponential growth, shielding the LCR within tetrameric Cdc19 or phosphorylation of the LCR prevents unscheduled aggregation, while its dephosphorylation is necessary for reversible aggregation during stress. Cdc19 aggregation triggers its localization to stress granules and modulates their formation and dissolution. Reversible aggregation protects Cdc19 from stress-induced degradation, thereby allowing cell cycle restart after stress. Several other enzymes necessary for G1 progression also contain LCRs and aggregate reversibly during stress, implying that reversible aggregation represents a conserved mechanism regulating cell growth and survival.

Regulation of cell growth by nutrients is governed by highly conserved signaling pathways, yet mechanisms of nutrient sensing are still poorly understood. In yeast, glucose activates both the Ras/PKA pathway and TORC1, which coordinately regulate growth through enhancing translation and ribosome biogenesis and suppressing autophagy. Here, we show that cytosolic pH acts as a cellular signal to activate Ras and TORC1 in response to glucose availability. We demonstrate that cytosolic pH is sensitive to the quality and quantity of the available carbon source (C-source). Interestingly, Ras/PKA and TORC1 are both activated through the vacuolar ATPase (V-ATPase), which was previously identified as a sensor for cytosolic pH in vivo. V-ATPase interacts with two distinct GTPases, Arf1 and Gtr1, which are required for Ras and TORC1 activation, respectively. Together, these data provide a molecular mechanism for how cytosolic pH links C-source availability to the activity of signaling networks promoting cell growth.

Protein aggregates, and in particular amyloids, are generally considered to be inherently irreversible aberrant clumps, and are often associated with pathologies, such as Alzheimer's disease, Parkinson's disease, or systemic amyloidosis. However, recent evidence demonstrates that some aggregates are not only fully reversible, but also perform essential physiological functions. Despite these new findings, very little is known about how these functional protein aggregates are regulated in a physiological context. Here, we take the yeast pyruvate kinase Cdc19 as an example of a protein forming functional, reversible, solid, amyloid-like aggregates in response to stress conditions. Cdc19 aggregation is regulated via an aggregation-prone low complexity region (LCR). In favorable growth conditions, this LCR is prevented from aggregating by phosphorylation or oligomerization, while upon glucose starvation it becomes exposed and allows aggregation. We suggest that LCR phosphorylation, oligomerization or partner-binding may be general and widespread mechanisms regulating LCR-mediated reversible protein aggregation. Moreover, we show that, as predicted by computational tools, Cdc19 forms amyloid-like aggregates in vitro. Interestingly, we also observe striking similarities between Cdc19 and its mammalian counterpart, PKM2. Indeed, also PKM2 harbors a LCR and contains several peptides with high amyloidogenic propensity, which coincide with known phosphorylation sites. Thus, we speculate that the formation of reversible, amyloid-like aggregates may be a general physiological mechanism for cells to adapt to stress conditions, and that the underlying regulatory mechanisms may be conserved from yeast to humans.

Cells respond to stress by blocking translation, rewiring metabolism, and forming transient mRNP assemblies called stress granules (SGs). After stress release, re-establishing homeostasis requires energy-consuming processes. However, the molecular mechanisms whereby cells restore energy production to disassemble SGs and reinitiate growth after stress remain poorly understood. Here we show that, upon stress, the ATP-producing enzyme Cdc19 forms inactive amyloids, and that their rapid re-solubilization is essential to restore energy production and disassemble SGs. Cdc19 re-solubilization is initiated by the glycolytic metabolite fructose-1,6-bisphosphate (FBP), which directly binds Cdc19 amyloids and facilitates conformational changes that allow Hsp104 and Ssa2 chaperone recruitment. FBP then promotes Cdc19 tetramerization, which boosts its activity to further enhance ATP production and SG disassembly. Together, these results describe a molecular mechanism essential for stress recovery, which directly couples metabolism with SG dynamics via regulation of Cdc19 amyloids.