Biology subjects

Calloni, G.

Publications and source records attributed to Calloni, G..

4 recordsLinked to original sources

bioRxiv2020

RNA-Binding and Prion Domains: The Yin and Yang of Phase Separation

Biomolecular condensates are membrane-less organelles mainly composed of proteins and RNAs that organize intracellular spaces and regulate biochemical reactions. The ability of proteins and RNAs to phase separate is encoded in their sequences, yet it is still unknown which domains drive the process and what are their specific roles. Here, we systematically investigated the human and yeast proteomes to find regions promoting biomolecular condensation. Using advanced computational models to predict the phase separation propensity of proteins, we designed a set of experiments to investigate the contributions of Prion-Like Domains (PrLDs) and RNA-Binding Domains (RBDs). We found that while just one PrLD is sufficient to drive protein condensation, multiple RBDs are needed to modulate the dynamicity of the assemblies. In the case of stress granule protein Pub1 we show that the PrLD promotes sequestration of protein partners and the RBD confers liquid-like behaviour to the condensate. Our work sheds light on the fine interplay between RBDs and PrLD to regulate formation of membrane-less organelles, opening up the avenue for their manipulation.

molecular biology↗

bioRxiv2019

N1-methyladenine safeguards against aberrant RNA-protein association during proteostasis stress

Post-transcriptional modifications of nucleotide bases can effect several aspects of mRNA function. For example, the recent work has established the role of m6A in the coordinated regulation of transcriptome turnover and translation during cellular differentiation and tumorigenesis. The levels of m1A in mRNAs reach up to 10% of those of m6A, yet the functional consequences of this modification are much less clear. Here we show that N1-methyladenine protects mRNAs against aberrant interactions during heat shock and amyloidogenesis in mammalian cells. The m1A methylation motif correlated with the enhanced sequestration of transcripts in stress granules (SG). The cognate methyltransferase TRMT6/61A accumulated and m1A was enriched in SG. Downregulation of the catalytic subunit TRMT61A enhanced amyloidogenesis in the cytosol and increased bystander protein and RNA co-aggregation with A{beta} aggregates. Faulty granulation of mutant RNAs has been implicated in pathogenesis of protein aggregation disorders. Our results demonstrate that also normal mRNAs succumb to co-aggregation with proteins if RNA dynamics during stress is disturbed due to the insufficient N1-adenine methylation.

cell biology↗

bioRxiv2019

Active role of free RNA during the early phase of proteostasis stress

Transient sequestration of proteins and RNA is an essential principle of cellular reaction to stress. Compared to polypeptides, less is known about the role of RNA released from polysomes during acute proteostasis stress. Using quantitative mass spectrometry, we identified a set of proteins assembled by free RNA in the heat-shocked mammalian cytosol. RNA-associated proteins displayed higher disorder and larger size, which supports the role of multivalent interactions during the initial phase of the RNA granule formation. Structural features of the free RNA interactors defined them as a subset of RNA-binding proteins. The interactome contained preferentially the active form of eIF2. The interaction between assembled proteins in vivo required RNA. The reconstitution of the association process in vitro indicated to the multimolecular basis for the increased binding to RNA upon heat shock in the cytosol. Our results reveal how free RNA can participate in reorganization of cellular functions during proteostasis stress.

cell biology↗

bioRxiv2018

Insights into the structure-driven protein interactivity of RNA molecules

The combination of high-throughput sequencing and in vivo crosslinking approaches leads to the progressive uncovering of the complex interdependence between cellular transcriptome and proteome. Yet the molecular determinants that govern interactions in protein-RNA networks are poorly known at present. Here we used the most recent experimental data to investigate the relationship between RNA structure and protein interactions. Our results show that, independently of the particular technique, the amount of structure in RNA molecules correlates with the capacity of binding to proteins in vitro and in vivo. To validate this observation, we generated an in vitro network that mimics the composition of phase-separated RNA granules. We observed that RNA, when structured, competes with protein binding and can rearrange the interaction network. The simplicity of the principle bears great potential to boost the understanding and modelling of cellular processes involving RNA-protein interactions.

molecular biology↗