Matrin3 binds directly to intronic pyrimidine‐rich sequences and controls alternative splicing

Abstract

Matrin3 is an RNA-binding protein that is localized in the nuclear matrix. Although various roles in RNA metabolism have been reported for Matrin3, in vivo target RNAs to which Matrin3 binds directly have not been investigated comprehensively so far. Here, we show that Matrin3 binds predominantly to intronic regions of pre-mRNAs. Photoactivatable Ribonucleoside-Enhanced Cross-linking and Immunoprecipitation (PAR-CLIP) analysis using human neuronal cells showed that Matrin3 recognized pyrimidine-rich sequences as binding motifs, including the polypyrimidine tract, a splicing regulatory element. Splicing-sensitive microarray analysis showed that depletion of Matrin3 preferentially increased the inclusion of cassette exons that were adjacent to introns that contained Matrin3-binding sites. We further found that although most of the genes targeted by polypyrimidine tract binding protein 1 (PTBP1) were also bound by Matrin3, Matrin3 could control alternative splicing in a PTBP1-independent manner, at least in part. These findings suggest that Matrin3 is a splicing regulator that targets intronic pyrimidine-rich sequences.

Lamin A/C controls nuclear matrin-3 levels and localization, but not alternative splicing of cassette exons

“…We performed RT-PCR from cDNA isolated from matrin-3 depleted U2OS cells using primers in flanking constitutive exons and the percentage exon inclusion was determined (Figure 2). Matrin-3 knock-down in U2OS cells resulted in significant splicing changes in 5 of 6 cassette exons as previously seen in HeLa cells [14, 15]. Matrin-3 knock-down enhanced inclusion of cassette exons in ST7, SETD5, PLEKHA3, PIGX and ACSL3 , while no change in splicing of exon 18 of TCF12 was observed.…”

“…RNA was isolated from U2OS cells and reverse transcribed to cDNA as described above. PCRs for cassette exon splicing were carried out using primer sets in flanking constitutive exons as previously described [14, 15]. All PCRs were carried out using the Jumpstart Taq polymerase (Sigma), and the products were separated and quantified on a QIAXcel capillary electrophoresis system (Qiagen) as previously described[14].…”

RNA processing in skeletal muscle biology and disease

“…TDP43 also plays a role in RNA stability and homeostasis [158], and whole genome RNA instability has been recently demonstrated in fibroblasts from individuals with ALS [159]. TDP43 interacts with other RNA processing factors, such as Matrin-3 (MATR3) and Fused-insarcoma (FUS) [160,161], that are frequently mutated in ALS [161][162][163][164]. While MATR3 is involved in polyadenylation site selection and intron retention via interactions with PABPN1 [162][163][164], FUS is a RBP that colocalizes with wild-type TDP43 in cytoplasmic inclusion bodies in models of ALS [161].…”

“…TDP43 interacts with other RNA processing factors, such as Matrin-3 (MATR3) and Fused-insarcoma (FUS) [160,161], that are frequently mutated in ALS [161][162][163][164]. While MATR3 is involved in polyadenylation site selection and intron retention via interactions with PABPN1 [162][163][164], FUS is a RBP that colocalizes with wild-type TDP43 in cytoplasmic inclusion bodies in models of ALS [161]. However, studies in autopsy human tissues do not support the idea that FUS and TDP43 overlap [165], indicating the need for additional studies to explore the potential interaction between FUS and TDP43 [166].…”

Matrin 3-dependent neurotoxicity is modified by nucleic acid binding and nucleocytoplasmic localization

“…The downstream implications of abnormal LLPS on RNA 55 misprocessing, RBP pathology, and neurodegeneration in ALS are unknown, however. 56Matrin 3 (MATR3) is a DNA-and RNA-binding protein with wide-ranging 57 functions in nucleic acid metabolism including gene transcription, the DNA damage 58 response, splicing, RNA degradation, and the sequestration of hyperedited RNAs 59 (Belgrader et al, 1991;Hibino et al, 2000;Zhang and Carmichael, 2001; Salton et al, 60 2014; Coelho et al, 2015;Rajgor et al, 2016;Uemura et al, 2017). The MATR3 S85C 61 mutation leads to autosomal dominant distal myopathy with vocal cord and pharyngeal 62 4 weakness (Feit et al, 1998;Senderek et al, 2009).…”