Nuclear envelope transmembrane proteins involved in genome organization are misregulated in myotonic dystrophy type 1 muscle

Abstract

Myotonic dystrophy type 1 is a multisystemic disorder with predominant muscle and neurological involvement. Despite a well described pathomechanism, which is primarily a global missplicing due to sequestration of RNA-binding proteins, there are still many unsolved questions. One such question is the disease etiology in the different affected tissues. We observed alterations at the nuclear envelope in primary muscle cell cultures before. This led us to reanalyze a published RNA-sequencing dataset of DM1 and control muscle biopsies regarding the misregulation of NE proteins. We could identify several muscle NE protein encoding genes to be misregulated depending on the severity of the muscle phenotype. Among these misregulated genes were NE transmembrane proteins (NETs) involved in nuclear-cytoskeletal coupling as well as genome organization. For selected genes, we could confirm that observed gene-misregulation led to protein expression changes. Furthermore, we investigated if genes known to be under expression-regulation by genome organization NETs were also misregulated in DM1 biopsies, which revealed that misregulation of two NETs alone is likely responsible for differential expression of about 10% of all genes being differentially expressed in DM1. Notably, the majority of NETs identified here to be misregulated in DM1 muscle are mutated in Emery-Dreifuss muscular dystrophy or clinical similar muscular dystrophies, suggesting a broader similarity on the molecular level for muscular dystrophies than anticipated. This shows not only the importance of muscle NETs in muscle health and disease, but also highlights the importance of the NE in DM1 disease progression.

Keywords: genome organization; muscle biopsy; myotonic dystrophy type 1; nuclear envelope; nuclear envelope transmembrane proteins.

FIGURE 1

Muscle nuclear envelope (NE) proteins are differential expressed and spliced in DM1. (A) Gene expression of 386 muscle NE proteins for proto-DM1, DM1, and severe DM1 based on dorsiflexion strength. (B) Number of genes being differential expressed and the proportion of NE transmembrane proteins (NETs). (C) GO-term enrichment for the differential expressed muscle NE proteins. (D) Number of genes being differential spliced and the proportion of NETs. (E) GO-term enrichment for the differential spliced muscle NE proteins.

FIGURE 2

LINC (linker of nucleo- and cytoskeleton) complex protein expression and splicing is altered in DM1. (A) SYNE1 gene expression is inversely correlated with dorsiflexion strength (left panel), there is a switch in isoform expression (second panel) and the muscle-specific DV23 exon is preferentially spliced out in DM1 (third panel). Western blot analysis confirms the expression changes for short nesprin isoforms (fourth panel). (B) SYNE3 gene expression is inversely correlated with dorsiflexion strength (left panel), there is a switch in isoform expression (right panel). (C) SYNE2 gene expression slightly correlates with dorsiflexion strength (left panel), there appears to be a general slight downregulation of several isoforms (right panel). (D) SUN2 gene expression correlates with dorsiflexion strength (left panel), the main muscle isoform is downregulated (right panel). (E) SUN1 gene expression is unaffected (left panel), but there is a switch in isoform expression (right panel). (F) TMEM201 gene expression correlates with dorsiflexion strength (left panel), the main muscle isoform is downregulated (right panel).

FIGURE 3

Genome organizing muscle nuclear envelope transmembrane protein (NET) expression is altered in DM1 and affects gene expression. (A) PLPP7 gene expression is correlated with dorsiflexion strength (left panel), the main muscle isoform is downregulated (middle panel). Western blot analysis confirms the expression changes (right panel). (B) TMEM38A gene expression is correlated with dorsiflexion strength (left panel), the main muscle isoform is downregulated (middle panel). Western blot analysis confirms the expression changes (right panel). (C) TOR1AIP1 gene expression is correlated with dorsiflexion strength (left panel), several isoforms are downregulated (right panel). (D) EMD gene expression is correlated with dorsiflexion strength (left panel), several isoforms only different in the UTR-region are downregulated (right panel). (E) Overlap between Plpp7 and Tmem38a regulated genes and DM1 differential expressed genes for proto-DM1, DM1, and severe DM1 based on dorsiflexion strength. (F) GO-term enrichment for the Plpp7 and Tmem38a regulated and DM1 differential expressed genes.