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

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.

Chromosome territories and the global regulation of the genome

“…A recent transcriptome and interactome analysis revealed over 100 proteins that bind matrin 3 (prominently including proteins associated with RNA processing/splicing). Deletion of a RNA recognition motif (RRM1) or Zn finger motifs (ZnF1 or ZnF2) diminished the binding of a subset of matrin 3 interacting proteins and altered its intranuclear organization . This study also uncovered a crucial role of the RRM2 motif of matrin 3 for regulating interactions with other nuclear proteins and maintaining its characteristic intranuclear localization.…”

“…120,121 Recent proteomic approaches have further confirmed the S/MAR binding activities of SAF-A, matrin 3, and nuclear lamins. [122][123][124] CTCF and cohesin have been identified as key players in the arrangement of chromatin into higher order domains termed Topologically Associated Domains (TADs), which mediate long range intrachromosomal and interchromosomal interactions within the nucleus. 15,106,[125][126][127][128][129][130][131][132][133][134] Recently, HnRNP-U (SAF-A) has been demonstrated to be integral to maintaining genome organization at TADs and higher levels of chromatin.…”

“…Based on these findings and similar properties of these two proteins, it was proposed that SAF‐A and matrin 3 are major components of a nuclear scaffolding structure that serve as dynamic assembly centers for the organization of functional neighborhoods of genomic function in the cell nucleus . The potential physiological roles that matrin 3 plays in nuclear organization and function are further emphasized by the recent findings connecting mutations in the matrin 3 gene (MATR3) to the diseases ALS, distal myopathy, and left ventricular outflow tract obstruction (LVOTO), a common congenital heart defect …”

“…These include topoisomerase II, CTCF, cohesin, matrin 3 (also called as P‐130 nuclear scaffold protein), HnRNP‐U (also called as SAF‐A, for Scaffold/Matrix attachment region factor), SAF‐B, ARBP (Attachment Region Binding Protein), SAT‐B, and nuclear lamins . Recent proteomic approaches have further confirmed the S/MAR binding activities of SAF‐A, matrin 3, and nuclear lamins . CTCF and cohesin have been identified as key players in the arrangement of chromatin into higher order domains termed Topologically Associated Domains (TADs), which mediate long range intrachromosomal and interchromosomal interactions within the nucleus .…”

“…Detailed visualization studies including computer‐assisted analysis demonstrated that matrin 3 forms a network‐like structure in the nucleus, which permeates the interior of CT and is in close proximity to functional domains containing active sites of transcription, RNA polymerase II, RNA splicing, and other functionally related factors . While the potential role of matrin 3 as a major component of an overall scaffolding network in the cell nucleus remains to be resolved, it is interesting to note that deletion mutations in the RRM2 domain resulted in major alterations of the network‐like organization of matrin 3 into large nuclear body‐like structures and protein‐protein interactions . Moreover, colocalization studies demonstrated a similar network organization and close 3‐D proximity of the S/MAR binding protein SAF‐A (hnRNP‐U) with matrin 3 including penetration into the interior of CT .…”