Genome organization around nuclear speckles drives mRNA splicing efficiency.

The nucleus is highly organized, such that factors involved in the transcription and processing of distinct classes of RNA are confined within specific nuclear bodies1,2. One example is the nuclear speckle, which is defined by high concentrations of protein and noncoding RNA regulators of pre-mRNA splicing3. What functional role, if any, speckles might play in the process of mRNA splicing is unclear4,5. Here we show that genes localized near nuclear speckles display higher spliceosome concentrations, increased spliceosome binding to their pre-mRNAs and higher co-transcriptional splicing levels than genes that are located farther from nuclear speckles. Gene organization around nuclear speckles is dynamic between cell types, and changes in speckle proximity lead to differences in splicing efficiency. Finally, directed recruitment of a pre-mRNA to nuclear speckles is sufficient to increase mRNA splicing levels. Together, our results integrate the long-standing observations of nuclear speckles with the biochemistry of mRNA splicing and demonstrate a crucial role for dynamic three-dimensional spatial organization of genomic DNA in driving spliceosome concentrations and controlling the efficiency of mRNA splicing.

DHX15-independent roles for TFIP11 in U6 snRNA modification, U4/U6.U5 tri-snRNP assembly and pre-mRNA splicing fidelity.

The U6 snRNA, the core catalytic component of the spliceosome, is extensively modified post-transcriptionally, with 2'-O-methylation being most common. However, how U6 2'-O-methylation is regulated remains largely unknown. Here we report that TFIP11, the human homolog of the yeast spliceosome disassembly factor Ntr1, localizes to nucleoli and Cajal Bodies and is essential for the 2'-O-methylation of U6. Mechanistically, we demonstrate that TFIP11 knockdown reduces the association of U6 snRNA with fibrillarin and associated snoRNAs, therefore altering U6 2'-O-methylation. We show U6 snRNA hypomethylation is associated with changes in assembly of the U4/U6.U5 tri-snRNP leading to defects in spliceosome assembly and alterations in splicing fidelity. Strikingly, this function of TFIP11 is independent of the RNA helicase DHX15, its known partner in yeast. In sum, our study demonstrates an unrecognized function for TFIP11 in U6 snRNP modification and U4/U6.U5 tri-snRNP assembly, identifying TFIP11 as a critical spliceosome assembly regulator.

Pathological tau drives ectopic nuclear speckle scaffold protein SRRM2 accumulation in neuron cytoplasm in Alzheimer's disease.

Several conserved nuclear RNA binding proteins (sut-1, sut-2, and parn-2) control tau aggregation and toxicity in C. elegans, mice, and human cells. MSUT2 protein normally resides in nuclear speckles, membraneless organelles composed of phase-separated RNAs and RNA-binding proteins that mediate critical steps in mRNA processing including mRNA splicing. We used human pathological tissue and transgenic mice to identify Alzheimer's disease-specific cellular changes related to nuclear speckles. We observed that nuclear speckle constituent scaffold protein SRRM2 is mislocalized and accumulates in cytoplasmic lesions in AD brain tissue. Furthermore, progression of tauopathy in transgenic mice is accompanied by increasing mislocalization of SRRM2 from the neuronal nucleus to the soma. In AD brain tissue, SRRM2 mislocalization associates with increased severity of pathological tau deposition. These findings suggest potential mechanisms by which pathological tau impacts nuclear speckle function in diverse organisms ranging from C. elegans to mice to humans. Future translational studies aimed at restoring nuclear speckle homeostasis may provide novel candidate therapeutic targets for pharmacological intervention.

USP42 drives nuclear speckle mRNA splicing via directing dynamic phase separation to promote tumorigenesis.

Liquid-liquid phase separation is considered a generic approach to organize membrane-less compartments, enabling the dynamic regulation of phase-separated assemblies to be investigated and pivotal roles of protein posttranslational modifications to be demonstrated. By surveying the subcellular localizations of human deubiquitylases, USP42 was identified to form nuclear punctate structures that are associated with phase separation properties. Bioinformatic analysis demonstrated that the USP42 C-terminal sequence was intrinsically disordered, which was further experimentally confirmed to confer phase separation features. USP42 is distributed to SC35-positive nuclear speckles in a positively charged C-terminal residue- and enzymatic activity-dependent manner. Notably, USP42 directs the integration of the spliceosome component PLRG1 into nuclear speckles, and its depletion interferes with the conformation of SC35 foci. Functionally, USP42 downregulation deregulates multiple mRNA splicing events and leads to deterred cancer cell growth, which is consistent with the impact of PLRG1 repression. Finally, USP42 expression is strongly correlated with that of PLRG1 in non-small-cell lung cancer samples and predicts adverse prognosis in overall survival. As a deubiquitylase capable of dynamically guiding nuclear speckle phase separation and mRNA splicing, USP42 inhibition presents a novel anticancer strategy by targeting phase separation.

Relationship between the infarct localization and left ventricular rotation parameters following acute ST-segment elevation myocardial infarction.

OBJECTIVE: This study was an investigation of the role of left ventricular (LV) apical rotation seen in the early period after myocardial infarction (MI) in predicting infarct localization. METHODS: A total of 124 patients with a ST-Segment elevation myocardial infarction (STEMI) diagnosis who underwent primary percutaneous coronary intervention (PCI) and 50 healthy volunteers with similar demographic characteristics were included in the study. The relationship between 2-dimenstional speckle tracking echocardiography (STE)-guided LV apical rotation angle measurements and technetium-99m sestamibi-single-photon emission computed tomography (SPECT)-guided infarct localization was evaluated. Conventional echocardiography and STE were performed on average 2 days after PCI, and gated SPECT myocardial perfusion imaging (MPI) was performed within an average of 60 days. RESULTS: The apical rotation angle was lower in patients with an anterior MI compared with those who had an inferior MI and the control group (AntMI-InfMI: 6.51±2.4°, AntMI-Control: 13.20±2.5°, InfMI-Control: 14.3±2.1°; p value: 0.00, 0.00, 0.15, respectively). SPECT MPI analysis revealed the presence of an LV apical scar in all patients with acute anterior MI, but only 14 of those with inferior MI group (usually the inferoapical wall). The apical rotation angle recorded in patients with apical scar was lower than that of the patients without apical scar (7.6±2.8° and 14.5±2°, respectively; p=0.00). Receiver operating characteristic curve analysis yielded an area under the curve for apical rotation of 0.799 (p<0.01). The optimal cutoff value of 12.1° had a sensitivity of 78.3% and a specificity of 68.2% for predicting LV apical scar following STEMI. CONCLUSION: Detection of apical rotation angle decrease in the early period after STEMI may be useful in predicting extension of infarct scarring to the LV apex.