Biology subjects

Bardwell, J. C.

Publications and source records attributed to Bardwell, J. C..

3 recordsLinked to original sources

bioRxiv2024

Molecular insights into the interaction between a disordered protein and a folded RNA

Intrinsically disordered protein regions (IDRs) are well-established as contributors to intermolecular interactions and the formation of biomolecular condensates. In particular, RNA-binding proteins (RBPs) often harbor IDRs in addition to folded RNA-binding domains that contribute to RBP function. To understand the dynamic interactions of an IDR-RNA complex, we characterized the RNA-binding features of a small (68 residues), positively charged IDR-containing protein, SERF. At high concentrations, SERF and RNA undergo charge-driven associative phase separation to form a protein- and RNA-rich dense phase. A key advantage of this model system is that this threshold for demixing is sufficiently high that we could use solution-state biophysical methods to interrogate the stoichiometric complexes of SERF with RNA in the one-phase regime. Herein, we describe our comprehensive characterization of SERF alone and in complex with a small fragment of the HIV-1 TAR RNA (TAR) with complementary biophysical methods and molecular simulations. We find that this binding event is not accompanied by the acquisition of structure by either molecule; however, we see evidence for a modest global compaction of the SERF ensemble when bound to RNA. This behavior likely reflects attenuated charge repulsion within SERF via binding to the polyanionic RNA and provides a rationale for the higher-order assembly of SERF in the context of RNA. We envision that the SERF-RNA system will lower the barrier to accessing the details that support IDR-RNA interactions and likewise deepen our understanding of the role of IDR-RNA contacts in complex formation and liquid-liquid phase separation. SIGNIFICANCESubcellular organization through the formation of biomolecular condensates has emerged as an important contributor to myriad cellular functions, with implications in homeostasis, stress response, and disease. To understand the general and specific principles that support condensate formation, we must interrogate the interactions and assembly of their constituent biomolecules. To this end, this study introduces a simple model system comprised of a small, disordered protein and small RNA that undergo charge-driven, associative phase separation. In addition to extensive biophysical characterization of these molecules and their complex, we also generate new insights into mode of interaction and assembly between an unstructured protein and a structured RNA.

biochemistry↗

bioRxiv2023

SERF2, an RNA G-quadruplex Binding Protein, promotes stress granule formation

Liquid-liquid phase transitions govern a wide range of protein-protein and protein-RNA interactions. Although the importance of multivalency and protein disorder in driving these transitions is clear, there is limited knowledge concerning the structural basis of phase transitions or the conformational changes that accompany this process. In this work, we found that a small human protein, SERF2, is important for the formation of stress granules. We determined the solution NMR structure ensemble of SERF2. We show that SERF2 specifically interacts with non-canonical tetrahelical RNA structures called G-quadruplexes, structures linked to stress granule formation. The biophysical amenability of both SERF2 and RNA G4 quadruplexes have allowed us to characterize the multivalent protein-RNA interactions involved in liquid-liquid phase transitions, the role that protein disorder plays in these transitions, identify the specific contacts involved, and describe how these interactions impact the structural dynamics of the components enabling a detailed understanding of the structural transitions involved in early stages of ribonucleoprotein condensate formation.

cell biology↗

bioRxiv2023

Effects of protein G-quadruplex interactions on phase transitions and protein aggregation

The SERF family of proteins were originally discovered for their ability to accelerate amyloid formation. Znf706 is an uncharacterized protein whose N-terminus is homologous to SERF proteins. We show here that human Znf706 can promote protein aggregation and amyloid formation. Unexpectedly, Znf706 specifically interacts with stable, non-canonical nucleic acid structures known as G-quadruplexes. G-quadruplexes can affect gene regulation and suppress protein aggregation; however, it is unknown if and how these two activities are linked. We find Znf706 binds preferentially to parallel G-quadruplexes with low micromolar affinity, primarily using its N-terminus, and upon interaction, its dynamics are constrained. G-quadruplex binding suppresses Znf706s ability to promote protein aggregation. Znf706 in conjunction with G-quadruplexes therefore may play a role in regulating protein folding. RNAseq analysis shows that Znf706 depletion specifically impacts the mRNA abundance of genes that are predicted to contain high G-quadruplex density. Our studies give insight into how proteins and G-quadruplexes interact, and how these interactions affect both partners and lead to the modulation of protein aggregation and cellular mRNA levels. These observations suggest that the SERF family of proteins, in conjunction with G-quadruplexes, may have a broader role in regulating protein folding and gene expression than previously appreciated.

biochemistry↗