Splicing factors such as BUD31 are identified in a synthetic-lethal screen with cells overexpressing the transcription factor MYC; oncogenic MYC leads to an increase in pre-mRNA synthesis, and spliceosome inhibition impairs the growth and tumorigenicity of MYC-dependent breast cancers, suggesting that spliceosome components may be potential therapeutic targets for MYC-driven cancers. The transcription factor MYC is frequently amplified or overexpressed in cancer and drives increased RNA and protein production. Here, Thomas Westbrook and colleagues identify the splicing factor BUD31 in a synthetic lethal screen with cells overexpressing MYC and show that other splicing factors are also required for cells to tolerate overexpressed MYC. Oncogenic MYC leads to an increase in pre-mRNA synthesis, and inhibition of the spliceosome impairs the growth and tumorigenicity of MYC-dependent breast cancer cells. Spliceosome components may therefore be potential therapeutic targets for MYC-driven cancers. MYC (also known as c-MYC) overexpression or hyperactivation is one of the most common drivers of human cancer. Despite intensive study, the MYC oncogene remains recalcitrant to therapeutic inhibition. MYC is a transcription factor, and many of its pro-tumorigenic functions have been attributed to its ability to regulate gene expression programs1,2,3. Notably, oncogenic MYC activation has also been shown to increase total RNA and protein production in many tissue and disease contexts4,5,6,7. While such increases in RNA and protein production may endow cancer cells with pro-tumour hallmarks, this increase in synthesis may also generate new or heightened burden on MYC-driven cancer cells to process these macromolecules properly8. Here we discover that the spliceosome is a new target of oncogenic stress in MYC-driven cancers. We identify BUD31 as a MYC-synthetic lethal gene in human mammary epithelial cells, and demonstrate that BUD31 is a component of the core spliceosome required for its assembly and catalytic activity. Core spliceosomal factors (such as SF3B1 and U2AF1) associated with BUD31 are also required to tolerate oncogenic MYC. Notably, MYC hyperactivation induces an increase in total precursor messenger RNA synthesis, suggesting an increased burden on the core spliceosome to process pre-mRNA. In contrast to normal cells, partial inhibition of the spliceosome in MYC-hyperactivated cells leads to global intron retention, widespread defects in pre-mRNA maturation, and deregulation of many essential cell processes. Notably, genetic or pharmacological inhibition of the spliceosome in vivo impairs survival, tumorigenicity and metastatic proclivity of MYC-dependent breast cancers. Collectively, these data suggest that oncogenic MYC confers a collateral stress on splicing, and that components of the spliceosome may be therapeutic entry points for aggressive MYC-driven cancers.

Induction of tumor cell death is the therapeutic goal for most anticancer drugs. Yet, a mode of drug-induced cell death, known as immunogenic cell death (ICD), can propagate antitumoral immunity to augment therapeutic efficacy. Currently, the molecular hallmark of ICD features the release of damage-associated molecular patterns (DAMPs) by dying cancer cells. Here, we show that gemcitabine, a standard chemotherapy for various solid tumors, triggers hallmark immunostimualtory DAMP release (e.g., calreticulin, HSP70, and HMGB1); however, is unable to induce ICD. Mechanistic studies reveal gemcitabine concurrently triggers prostaglandin E₂ release as an inhibitory DAMP to counterpoise the adjuvanticity of immunostimulatory DAMPs. Pharmacological blockade of prostaglandin E₂ biosythesis favors CD103⁺ dendritic cell activation that primes a Tc1-polarized CD8⁺ T cell response to bolster tumor rejection. Herein, we postulate that an intricate balance between immunostimulatory and inhibitory DAMPs could determine the outcome of drug-induced ICD and pose COX-2/prostaglandin E₂ blockade as a strategy to harness ICD.

Neuropathological aggregates of the intrinsically disordered microtubule-associated protein Tau are hallmarks of Alzheimer’s disease, with decades of research devoted to studying the protein’s aggregation properties both in vitro and in vivo. Recent demonstrations that Tau is capable of undergoing liquid-liquid phase separation (LLPS) reveal the possibility that protein-enriched phase separated compartments could serve as initiation sites for Tau aggregation, as shown for other amyloidogenic proteins, such as the Fused in Sarcoma protein (FUS) and TAR DNA-binding protein-43 (TDP-43). Although truncation, mutation, and hyperphosphorylation have been shown to enhance Tau LLPS and aggregation, the effect of hyperacetylation on Tau aggregation remains unclear. Here, we investigate how the acetylation of Tau affects its potential to undergo phase separation and aggregation. Our data show that the hyperacetylation of Tau by p300 histone acetyltransferase (HAT) disfavors LLPS, inhibits heparin-induced aggregation, and impedes access to LLPS-initiated microtubule assembly. We propose that Tau acetylation prevents the toxic effects of LLPS-dependent aggregation but, nevertheless, contributes to Tau loss-of-function pathology by inhibiting Tau LLPS-mediated microtubule assembly.

In quantitative mass spectrometry, the method by which peptides are grouped into proteins can have dramatic effects on downstream analyses. Here we describe gpGrouper, an inference and quantitation algorithm that offers an alternative method for assignment of protein groups by gene locus and improves pseudo-absolute iBAQ quantitation by weighted distribution of shared peptide areas. We experimentally show that distributing shared peptide quantities based on unique peptide peak ratios improves quantitation accuracy compared with conventional winner-take-all scenarios. Furthermore, gpGrouper seamlessly handles two-species samples such as patient-derived xenografts (PDXs) without ignoring the host species or species-shared peptides. This is a critical capability for proper evaluation of proteomics data from PDX samples, where stromal infiltration varies across individual tumors. Finally, gpGrouper calculates peptide peak area (MS1) based expression estimates from multiplexed isobaric data, producing iBAQ results that are directly comparable across label-free, isotopic, and isobaric proteomics approaches. Graphical Abstract Highlights Gene-centric inference algorithm with classification for distinguishable groups. Shared peptides are split proportionally to corresponding unique peptide ratios. iBAQ values are calculated for label-free, isotopic or isobaric labeling methods. Universally handles single or mixed species PDX data with accurate deconvolution.

Stress granules are large macromolecular aggregates that contain translation initiation complexes and mRNAs. Stress granule formation coincides with translational repression, and stress granules actively signal to mediate cell fate decisions by a) signaling to the translation apparatus to maintain translational repression, b) mount various transcriptional responses including innate immunity, and c) to repress apoptosis. Previous work showed G3BP1 is phosphorylated at serine 149, which regulates G3BP1 oligomerization, stress granule assembly, and RNase activity intrinsic to G3BP1. However, the kinase that phosphorylates G3BP1 was not identified, leaving a key step in stress granule regulation uncharacterized. Here we show that Casein Kinase 2 promotes stress granule dynamics using chemical inhibition, genetic depletion, and overexpression experiments. These results link CK2 activity with SG disassembly. We also show that Casein Kinase 2 phosphorylates G3BP1 at serine 149 in vitro and in cells. These data support a role for casein kinase 2 in regulation of protein synthesis by down-regulating stress granule formation through G3BP1.

Neuronal activity drives alterations in gene expression within neurons, yet how it directs transcriptional and epigenomic changes in neighboring astrocytes in functioning circuits is unknown. We found that neuronal activity induces widespread transcriptional up-regulation and down-regulation in astrocytes, highlighted by the identification of Slc22a3 as an activity-inducible astrocyte gene that encodes neuromodulator transporter Slc22a3 and regulates sensory processing in the mouse olfactory bulb. Loss of astrocytic Slc22a3 reduced serotonin levels in astrocytes, leading to alterations in histone serotonylation. Inhibition of histone serotonylation in astrocytes reduced the expression of γ-aminobutyric acid (GABA) biosynthetic genes and GABA release, culminating in olfactory deficits. Our study reveals that neuronal activity orchestrates transcriptional and epigenomic responses in astrocytes while illustrating new mechanisms for how astrocytes process neuromodulatory input to gate neurotransmitter release for sensory processing.