Grishkovskaya, I. | bioRxiv Science

bioRxiv2020

PHF3 regulates neuronal gene expression through the new Pol II CTD reader domain SPOC

The C-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II) is a regulatory hub for transcription and RNA processing. Here, we identify PHD-finger protein 3 (PHF3) as a new CTD-binding factor that negatively regulates transcription and mRNA stability. The PHF3 SPOC domain preferentially binds to CTD repeats phosphorylated on Serine-2 and PHF3 tracks with Pol II across the length of genes. PHF3 competes with TFIIS for Pol II binding through its TFIIS-like domain (TLD), thus inhibiting TFIIS-mediated rescue of backtracked Pol II. PHF3 knock-out or PHF3 SPOC deletion in human cells result in gene upregulation and a global increase in mRNA stability, with marked derepression of neuronal genes. Key neuronal genes are aberrantly expressed in Phf3 knock-out mouse embryonic stem cells, resulting in impaired neuronal differentiation. Our data suggest that PHF3 is a prominent effector of neuronal gene regulation at the interface of transcription elongation and mRNA decay.

biochemistry↗

bioRxiv2019

Structures of three MORN repeat proteins and a re-evaluation of the proposed lipid-binding properties of MORN repeats

MORN (Membrane Occupation and Recognition Nexus) repeat proteins have a wide taxonomic distribution, being found in both prokaryotes and eukaryotes. Despite this ubiquity, they remain poorly characterised at both a structural and a functional level compared to other common repeats. In functional terms, they are often assumed to be lipid-binding modules that mediate membrane targeting. We addressed this putative activity by focusing on a protein composed solely of MORN repeats - Trypanosoma brucei MORN1. Surprisingly, no evidence for binding to membranes or lipid vesicles by TbMORN1 could be obtained either in vivo or in vitro. Conversely, TbMORN1 did interact with individual phospholipids. High- and low-resolution structures of the MORN1 protein from Trypanosoma brucei and homologous proteins from the parasites Toxoplasma gondii and Plasmodium falciparum were obtained using a combination of macromolecular crystallography, small-angle X-ray scattering, and electron microscopy. This enabled a first structure-based definition of the MORN repeat itself. Furthermore, all three structures dimerised via their C-termini in an antiparallel configuration. The dimers could form extended or V-shaped quaternary structures depending on the presence of specific interface residues. This work provides a new perspective on MORN repeats, showing that they are protein-protein interaction modules capable of mediating both dimerisation and oligomerisation.

biochemistry↗