Characterization of gene regulation and protein interaction networks for Matrin 3 encoding mutations linked to amyotrophic lateral sclerosis and myopathy

M Carolina Gallego Iradi,Judy C Triplett,James D Thomas,Anthony M Crown,David R Borchelt,Hilda Brown,Guilian Xu,Jada Lewis,Maurice S Swanson,Edgardo Rodriguez-Lebron,Rachel Davila

doi:10.1038/s41598-018-21371-4

Abstract

To understand how mutations in Matrin 3 (MATR3) cause amyotrophic lateral sclerosis (ALS) and distal myopathy, we used transcriptome and interactome analysis, coupled with microscopy. Over-expression of wild-type (WT) or F115C mutant MATR3 had little impact on gene expression in neuroglia cells. Only 23 genes, expressed at levels of >100 transcripts showed ≥1.6-fold changes in expression by transfection with WT or mutant MATR3:YFP vectors. We identified ~123 proteins that bound MATR3, with proteins associated with stress granules and RNA processing/splicing being prominent. The interactome of myopathic S85C and ALS-variant F115C MATR3 were virtually identical to WT protein. Deletion of RNA recognition motif (RRM1) or Zn finger motifs (ZnF1 or ZnF2) diminished the binding of a subset of MATR3 interacting proteins. Remarkably, deletion of the RRM2 motif caused enhanced binding of >100 hundred proteins. In live cells, MATR3 lacking RRM2 (ΔRRM2) formed intranuclear spherical structures that fused over time into large structures. Our findings in the cell models used here suggest that MATR3 with disease-causing mutations is not dramatically different from WT protein in modulating gene regulation or in binding to normal interacting partners. The intra-nuclear localization and interaction network of MATR3 is strongly modulated by its RRM2 domain.

Highlights

To understand how mutations in Matrin 3 (MATR3) cause amyotrophic lateral sclerosis (ALS) and distal myopathy, we used transcriptome and interactome analysis, coupled with microscopy
There were no obvious examples of genes that were dysregulated by the over-expression of F115C-MATR3 within these criteria
To determine whether disease-causing mutations in MATR3 may disrupt its normal interaction network, we conducted affinity-capture proteomic studies in HEK293 cells to identify proteins interacting with WT, S85C, and F115C MATR3

Summary

Introduction

To understand how mutations in Matrin 3 (MATR3) cause amyotrophic lateral sclerosis (ALS) and distal myopathy, we used transcriptome and interactome analysis, coupled with microscopy. Our findings in the cell models used here suggest that MATR3 with disease-causing mutations is not dramatically different from WT protein in modulating gene regulation or in binding to normal interacting partners. A more recent study of MATR3 interactomes included analysis of WT and mutant MATR3 in murine NSC-34 cells[19] This later study, identified proteins of the TRanscription and EXport (TREX) complex as interacting partners that may be influenced by disease-causing mutations. We have used three complementary approaches to characterize the impact that disease-causing mutations in MATR3 may have on its ability to regulate gene expression, interact with other proteins, or properly localize within the cell. Through analysis of MATR3 proteins with deletions in conserved RNA recognition motifs (RRMs) and Zn finger motifs (ZnFs), we revealed a crucial role of the RRM2 motif in MATR3 in regulating its interaction with other nuclear proteins and maintaining its intranuclear localization

Methods

Results

Conclusion