The SPOC proteins DIDO3 and PHF3 co-regulate gene expression and neuronal differentiation.

Transcription is regulated by a multitude of activators and repressors, which bind to the RNA polymerase II (Pol II) machinery and modulate its progression. Death-inducer obliterator 3 (DIDO3) and PHD finger protein 3 (PHF3) are paralogue proteins that regulate transcription elongation by docking onto phosphorylated serine-2 in the C-terminal domain (CTD) of Pol II through their SPOC domains. Here, we show that DIDO3 and PHF3 form a complex that bridges the Pol II elongation machinery with chromatin and RNA processing factors and tethers Pol II in a phase-separated microenvironment. Their SPOC domains and C-terminal intrinsically disordered regions are critical for transcription regulation. PHF3 and DIDO exert cooperative and antagonistic effects on the expression of neuronal genes and are both essential for neuronal differentiation. In the absence of PHF3, DIDO3 is upregulated as a compensatory mechanism. In addition to shared gene targets, DIDO specifically regulates genes required for lipid metabolism. Collectively, our work reveals multiple layers of gene expression regulation by the DIDO3 and PHF3 paralogues, which have specific, co-regulatory and redundant functions in transcription.

SPOC domain proteins in health and disease.

Since it was first described >20 yr ago, the SPOC domain (Spen paralog and ortholog C-terminal domain) has been identified in many proteins all across eukaryotic species. SPOC-containing proteins regulate gene expression on various levels ranging from transcription to RNA processing, modification, export, and stability, as well as X-chromosome inactivation. Their manifold roles in controlling transcriptional output implicate them in a plethora of developmental processes, and their misregulation is often associated with cancer. Here, we provide an overview of the biophysical properties of the SPOC domain and its interaction with phosphorylated binding partners, the phylogenetic origin of SPOC domain proteins, the diverse functions of mammalian SPOC proteins and their homologs, the mechanisms by which they regulate differentiation and development, and their roles in cancer.

The SPOC domain is a phosphoserine binding module that bridges transcription machinery with co- and post-transcriptional regulators.

The heptad repeats of the C-terminal domain (CTD) of RNA polymerase II (Pol II) are extensively modified throughout the transcription cycle. The CTD coordinates RNA synthesis and processing by recruiting transcription regulators as well as RNA capping, splicing and 3'end processing factors. The SPOC domain of PHF3 was recently identified as a CTD reader domain specifically binding to phosphorylated serine-2 residues in adjacent CTD repeats. Here, we establish the SPOC domains of the human proteins DIDO, SHARP (also known as SPEN) and RBM15 as phosphoserine binding modules that can act as CTD readers but also recognize other phosphorylated binding partners. We report the crystal structure of SHARP SPOC in complex with CTD and identify the molecular determinants for its specific binding to phosphorylated serine-5. PHF3 and DIDO SPOC domains preferentially interact with the Pol II elongation complex, while RBM15 and SHARP SPOC domains engage with writers and readers of m6A, the most abundant RNA modification. RBM15 positively regulates m6A levels and mRNA stability in a SPOC-dependent manner, while SHARP SPOC is essential for its localization to inactive X-chromosomes. Our findings suggest that the SPOC domain is a major interface between the transcription machinery and regulators of transcription and co-transcriptional processes.

RNA editing of Filamin A pre-mRNA regulates vascular contraction and diastolic blood pressure.

Epitranscriptomic events such as adenosine-to-inosine (A-to-I) RNA editing by ADAR can recode mRNAs to translate novel proteins. Editing of the mRNA that encodes actin crosslinking protein Filamin A (FLNA) mediates a Q-to-R transition in the interactive C-terminal region. While FLNA editing is conserved among vertebrates, its physiological function remains unclear. Here, we show that cardiovascular tissues in humans and mice show massive editing and that FLNA RNA is the most prominent substrate. Patient-derived RNA-Seq data demonstrate a significant drop in FLNA editing associated with cardiovascular diseases. Using mice with only impaired FLNA editing, we observed increased vascular contraction and diastolic hypertension accompanied by increased myosin light chain phosphorylation, arterial remodeling, and left ventricular wall thickening, which eventually causes cardiac remodeling and reduced systolic output. These results demonstrate a causal relationship between RNA editing and the development of cardiovascular disease indicating that a single epitranscriptomic RNA modification can maintain cardiovascular health.