Learning the chemical grammar of biomolecular condensates

Abstract

Biomolecular condensates compartmentalize and regulate assemblies of biomolecules engaged in vital physiological processes in cells. Specific proteins and nucleic acids engaged in shared functions occur in any one kind of condensate, suggesting that these compartments have distinct chemical specificities. Indeed, some small-molecule drugs concentrate in specific condensates due to chemical properties engendered by particular amino acids in the proteins in those condensates. Here we argue that the chemical properties that govern molecular interactions between a small molecule and biomolecules within a condensate can be ascertained for both the small molecule and the biomolecules. We propose that learning this 'chemical grammar', the rules describing the chemical features of small molecules that engender attraction or repulsion by the physicochemical environment of a specific condensate, should enable design of drugs with improved efficacy and reduced toxicity.

Figure 1

Small molecules can concentrate selectively within specific biomolecular condensates, and may do so both through interactions with the chemical environment of the condensate and through interactions with target proteins or nucleic acids within the condensate. Condensates are mesoscopic bodies and interaction of a molecule with the bulk solvation and microscopic chemical environments is determined by a condensate’s chemical specificity. This includes both specific interactions with target binding sites and interactions with other parts of biomolecules not constituting the target site that may contribute to the concentration of small molecules in a condensate. Left panel: small molecules (red spheres) that enter cells can be distributed unequally among diverse membrane-bound and non-membrane compartments. Middle panel: small molecules (red spheres) concentrating in a transcriptional condensate consisting of transcription apparatus assembled at a DNA locus; this can occur when the interaction of the small molecule with the chemical environment is favored over its interaction with the environment outside the condensate. Right panel: interaction of the small molecule is depicted with a specific biomolecular target, shown here: THZ1 bound to CDK7, PDB ID: 6xd3.

Figure 2

Chemical mechanisms in biomolecular condensate assembly. A) The stickers and spacers model suggests how polymers may associate to form condensates; amino acid side chains, nucleobases, and folded domains may produce interactions that qualify as stickers or spacers. ^{–, –, , , –, –}Examples of noncovalent interactions mediating sticker interactions is displayed with electrostatic surface potentials, computed at the M06-2x/6-311+g(d,p) level of theory. B) Condensates form droplets due to the association of biopolymers through noncovalent interactions between sticker regions which are separated by spacer regions in the same biopolymer. In the model system Fused in Sarcoma (FUS), glutamine rich spacers produced more solid-like behavior compared to the liquid-like character of glycine rich spacer regions.^, Noncovalent interactions between amino acids with π-systems, and other π-systems or cationic amino acids influence the formation and dissolution of FUS and other protein condensates by creating ‘sticker’ domains. ^{–, –, , , –, –} These interactions are a consequence of a molecular grammar, rules by which specific amino acids influence condensate formation and behavior.

Figure 3

Biomolecular condensates composed of different protein components have different chemical environments that engender selective partitioning of biomolecules. A) Condensates involved in different functions can be visualized in cells by imaging proteins that are specific to these bodies (e.g., MED1 in transcriptional activation, FIB1 in nucleolar ribosome biosynthesis, HP1a in heterochromatic gene silencing, and SRSF2 in RNA splicing). Images of murine embryonic stem cells with GFP-tagged proteins (green) and Hoechst staining (blue) acquired with a DeltaVision-OM Super resolution microscope. B) Model illustrating how chemical modification of a protein molecule can cause that molecule to change its condensate partitioning behavior. The RNA polymerase II C-Terminal Domain (CTD) becomes hyperphosphorylated during the transition to elongation, reducing the enzyme’s affinity for transcriptional condensates and increasing its affinity for splicing condensates. The RNA polymerase II CTD kinases CDK7 and CDK9 play well-established regulatory roles in transcription. C) Energy diagram showing how posttranslational modifications such as phosphorylation can alter the strength of noncovalent interactions among amino acid side chains. For example, phosphorylation of tyrosine residues will increase the interaction potential with an arginine residue.

Figure 4

Chemical specificity for small molecules in condensates. A) Cisplatin-TR selectively concentrates in specific condensates in droplet partitioning assays. B) Changes in amino acid composition of a condensate forming protein can abrogate small molecule partitioning behavior without affecting condensate formation. The ability of droplets formed by the MED1 protein of the Mediator complex to concentrate cisplatin is reduced with the replacement of aromatic residues with alanine residues.. C) Model depicting how specific local chemical environments within a condensate may influence cisplatin partitioning and concentration within a transcriptional condensate.

Figure 5

Cisplatin concentrates in large transcriptional condensates at driver oncogenes, where it selectively platinates oncogene regulatory DNA. Transcriptional condensates have been shown to occur at loci that contain clusters of enhancer regulatory elements called super-enhancers.^, In normal cells, super-enhancers typically span 5–20kb of DNA, but in metastatic tumor cells, driver oncogenes acquire super-enhancers that can span as much as 500 kb.^– The larger super-enhancers are associated with larger amounts of assembled transcription apparatus, and thus larger condensates. Larger transcriptional condensates have longer half-lives and produce more transcription from their associated genes.^– Thus, the continuous high concentration of cisplatin within the more long-lived oncogenic transcriptional condensates leads to robust DNA-platination at tumor-specific oncogenes, and ultimately this permanently disrupts the condensate at the oncogene, leading to tumor cell death. In contrast, the smaller short-lived condensates at normal genes accumulate far less cisplatin and suffer far less DNA damage.

Figure 6

Small molecules and peptides may be designed to have chemical properties that interact with and influence condensates to improve therapeutic efficacy. A) It should be possible to endow different classes of small molecules and peptides with features that concentrate these molecules in condensates where their targets occur and cause them to avoid partitioning into condensates where toxic effects might be obtained. B) Alteration of a condensate’s condensed fraction and phase behavior with a positive (increasing) or negative (decreasing) phase modulator. C) Material properties of condensate may be altered with small molecules; viscosity, viscoelasticity, surface tension, and diffusivity of molecules may be augmented, corrected, or depleted by drugs. D) Cyclopamine is an example of a small molecule that can induce changes in respiratory syncytial virus condensates by hardening (decreased viscoelasticity).