Molecular Cell最新文献

In this issue of Molecular Cell, Kral et al.¹ identify a targetable, novel mechanism of pancreatic ductal adenocarcinoma (PDAC) tumorigenesis via SRSF1 splicing-mediated regulation of an Alu-derived exon in Aurora kinase A (AURKA).

The ribosome-associated quality control (RQC) pathway resolves stalled ribosomes. As part of RQC, stalled nascent polypeptide chains (NCs) are appended with CArboxy-Terminal amino acid tails (CAT tails) in an mRNA-free, non-canonical elongation process. The relationship between CAT tail composition (alanine [Ala] and threonine [Thr] in yeast) and function has remained unknown. Using biochemical approaches in yeast, we discovered that mechanical forces on the NC regulate CAT tailing. We propose that CAT tailing initially operates in "extrusion mode," which increases NC lysine accessibility for on-ribosome ubiquitylation. Thr in CAT tails prevents the formation of polyalanine, which forms α-helices that lower extrusion efficiency and disrupt termination of CAT tailing. After NC ubiquitylation, pulling forces on the NC switch CAT tailing to an Ala-only "release mode," which facilitates NC release and degradation. Failure to switch from extrusion to release mode leads to the accumulation of NCs on large ribosomal subunits and proteotoxic aggregation of Thr-rich CAT tails.

The cytoplasm of eukaryotic cells is crowded with macromolecules. In principle, this crowding could have either a positive or a negative effect on the rates of biochemical reactions. Here, we review two commonly invoked theories to account for these possible effects then survey recent experimental work in cells and extracts that measures the effects. The evidence so far suggests that the effective second-order rate constants (a measure of the speed of a reaction for a given concentration of reactants) for reactions in vivo generally go down when crowding increases due to the slowing of diffusion. If the evidence presented so far proves to be general, it would have important implications for how we view the trade-offs that determine the biochemical dynamics of the cytoplasm.

Pancreatic ductal adenocarcinoma (PDAC) remains a highly lethal malignancy, driven by oncogenic KRAS mutations and dysregulated oncogenes, including SRSF1, MYC, and Aurora kinase A (AURKA). Although KRAS-targeted therapies are in development, resistance mechanisms underscore the need to identify alternative vulnerabilities. Here, we uncover an SRSF1-AURKA-MYC oncogenic circuit, wherein SRSF1 regulates AURKA 5' UTR alternative splicing, enhancing AURKA protein expression; AURKA positively regulates SRSF1 and MYC post-translationally, independently of its kinase activity; and MYC transcriptionally upregulates both SRSF1 and AURKA. Elevated SRSF1 in tumor cells promotes inclusion of an Alu-derived exon in the AURKA 5' UTR, resulting in splicing-dependent mRNA accumulation and exon-junction-complex deposition. Modulating 5' UTR splicing with splice-switching antisense oligonucleotides (ASOs) collapses the oncogenic circuit, reducing PDAC cell viability and triggering apoptosis. Our findings identify AURKA alternative splicing as a critical regulatory node and highlight a potential therapeutic strategy that simultaneously targets SRSF1, AURKA, and MYC oncogenes.

RNA-binding proteins (RBPs) with prion-like domains (PrLDs), such as FUS and TDP-43, condense into functional liquids, which can transform into pathological fibrils that underpin fatal neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS)/frontotemporal dementia (FTD). Here, we define short RNAs that prevent FUS fibrillization by promoting liquid phases and distinct short RNAs that prevent and reverse FUS condensation and fibrillization. These activities require interactions with multiple RNA-binding domains of FUS and are encoded by RNA sequence, length, and structure. We define a short RNA that dissolves cytoplasmic FUS aggregates, restores nuclear FUS, and mitigates FUS toxicity in optogenetic models and ALS patient-derived motor neurons. Another short RNA dissolves cytoplasmic TDP-43 aggregates, restores nuclear TDP-43, and mitigates TDP-43 toxicity. Since short RNAs can be effectively delivered to the human brain, these oligonucleotides could have utility for ALS/FTD and related disorders.

Despite progress in understanding pre-mRNA splicing, the regulatory mechanisms controlling most alternative splicing events remain unclear. We developed CRASP-seq (CRISPR-based identification of regulators of alternative splicing with phenotypic sequencing), a method that integrates pooled CRISPR-based genetic perturbations with deep sequencing of splicing reporters, to quantitatively assess the impact of all human genes on alternative splicing from a single RNA sample. CRASP-seq identified both known and untested regulators, enriched for proteins involved in RNA splicing and metabolism. As a proof-of-concept, CRASP-seq analysis of the LMNA cryptic splicing event linked to progeria uncovered ZNF207, primarily known for mitotic spindle assembly, as a regulator of progerin splicing. ZNF207 depletion enhances canonical LMNA splicing and decreases progerin protein levels in patient-derived cells. We further show that ZNF207's zinc-finger domain broadly impacts alternative splicing through direct interactions with U1 small nuclear ribonucleoprotein (snRNP) components. These findings position ZNF207 as a U1 snRNP auxiliary factor and demonstrate the power of CRASP-seq to uncover key regulators and domains of alternative splicing.