Biology subjects

Manogaran, A. L.

Publications and source records attributed to Manogaran, A. L..

4 recordsLinked to original sources

bioRxiv2025

Intracellular aggregation of the transthyretin protein is limited by Hsp70 chaperones

Extracellular amyloid deposits are a hallmark feature of systemic protein aggregation diseases such as transthyretin amyloidosis (ATTR). However, emerging evidence suggests that extracellular transthyretin (TTR) aggregates are internalized and result in an intracellular stress response, including elevated Hsp70 levels. While ATTR research has predominantly focused on extracellular TTR amyloid, our understanding of TTR aggregation inside the cell is poorly explored. To better understand how intracellular chaperones impact intracellular TTR, we used a yeast model that expresses TTR fused eGFP (TTR-eGFP) within the cytoplasm. Since the Hsp70 chaperone family, and co-chaperones the J-domain proteins (JDPs) and Hsp110, act as a disaggregase in vitro, we asked how these molecular chaperones impact TTR aggregation intracellularly in vivo. TTR-eGFP forms detergent-soluble high molecular weight (HMW) aggregates in yeast that have biochemistry profiles similar to human patient TTR. While knockdown of the JDP, Sis1, and deletion of the Hsp110, Sse1, appear to slightly increase TTR-eGFP aggregation, the loss of two major yeast Hsp70s, Ssa1 and Ssa2, lead to a significant increase in the size of HMW species. Taken together, our data suggest that Hsp70s limit the formation of HMW TTR aggregates in the intracellular environment. Based on our results, it is possible that the age-related decline of protein homeostasis, including Hsp70s, may promote the intracellular aggregation of TTR.

biochemistry↗

bioRxiv2025

Stress granules and protein aggregates reveal intracellular resource competition

Stress granules are biomolecular condensates that form in response to environmental stress and disassemble once normal conditions are restored. However, when disassembly fails, stress granules can persist and solidify. While stress granule solidification has been well documented, the cellular mechanisms underlying the transition from reversible to persistent stress granules remain unclear. Persistent stress granules can seed the formation of pathological aggregates, such as TDP-43 in amyotrophic lateral sclerosis1, 2. Although amyloid and tau aggregates are hallmarks of Alzheimers disease, a subset of patients also develop TDP-43 deposits, suggesting a possible role for stress granule solidification in Alzheimers disease progression3-5. Despite theoretical models explaining why persistence and ensuing solidification occurs, strong in vivo evidence is lacking6. Here we show that competition for limited chaperone resources drive stress granule persistence. In the presence of TDP-43 aggregates or yeast amyloid proteins called prions, stress granule disassembly is slowed or halted disassembly. Using yeast prions as a model, we show that the addition of chaperones, specifically the AAA+ ATPase molecular chaperone, Hsp104, resulted in resumption of stress granule disassembly. Our results demonstrate that the competition for shared resources, such as molecular chaperones, can limit stress granule disassembly. We suspect that the presence of pathological aggregates results in resource competition within the aging brain, contributing to the persistence of stress granules and their subsequent solidification and aggregation.

cell biology↗

bioRxiv2025

Hsp70 chaperones, Ssa1 and Ssa2, limit poly(A) binding protein aggregation

Molecular chaperones play a central role in maintaining protein homeostasis. The highly conserved Hsp70 family of chaperones have major functions in folding of nascent peptides, protein refolding, and protein aggregate disassembly. In yeast, loss of two Hsp70 proteins, Ssa1 and Ssa2, is associated with decreased cellular growth and shortened lifespan. While heterologous or mutant temperature sensitive proteins form anomalous large cytoplasmic inclusions in ssa1{Delta}ssa2{Delta} strains, it is unclear how endogenous wildtype proteins behave and are regulated in the presence of limiting Hsp70s. Using the wildtype yeast Poly A binding protein (Pab1), which is involved in mRNA binding and forms stress granules (SGs) upon heat shock, Pab1 forms large inclusions in approximately half of ssa1{Delta}ssa2{Delta} cells in the absence of stress. Overexpression of Ssa1, Hsp104, and Sis1 almost completely limits the formation of these large inclusions in ssa1{Delta}ssa2{Delta}, suggesting that excess Ssa1, Hsp104 and Sis1 can each compensate for the lower levels of Ssa proteins. Upon heat shock, SGs also form in cells whether large Pab1 inclusions are present or not. Surprisingly, cells containing only SGs disassemble faster than wildtype, whereas cells with both large inclusions disassemble slower albeit completely. We suspect that disassembly of these large inclusions is linked to the elevated heat shock response and elevated Hsp104 and Sis1 levels in ssa1{Delta}ssa2{Delta} strains. We also observed that wildtype cultures grown to saturation also form large Pab1-GFP inclusions. These inclusions can be partially rescued by overexpression of Ssa1. Taken together, our data suggests that Hsp70 not only plays a role in limiting unwanted protein aggregation in normal cells, but as cells age, the depletion of active Hsp70 possibly underlies the age-related aggregation of endogenous proteins.

cell biology↗

bioRxiv2017

Actin Impacts The Late Stages Of Prion Formation And Prion Propagation

In yeast, the [PSI+] and [PIN+] prions are aggregated forms of the Sup35 and Rnq1 proteins, respectively. The cellular mechanisms that underlie the formation and propagation of these prion states are not clearly understood. Our previous work suggested that actin networks play a role in early and late steps of the formation of [PSI+]. To further explore how actin impacts yeast prions, we turned to a set of actin point mutants. We found that the disruption of actin cables, either by an actin destabilizing drug or the act1-101 mutant, can enhance prion formation during the later stages of prion formation. Our data suggest that under normal conditions, actin cables play a role in limiting the inheritance of newly made prion particles to daughter cells. We also found actin can impact prion propagation. The act1-122 mutant, which contains a substitution in the fimbrin binding region, destabilized the [PIN+] prion over time. This is the first evidence that actin has a role in [PIN+] propagation. Taken together, our findings reveal novel roles for actin in the formation and propagation of prions.

cell biology↗