Macroscopic and microscopic diversity of missplicing in the central nervous system of patients with myotonic dystrophy type 1

Abstract

Myotonic dystrophy type I (DM1) is a multiorgan disease caused by CTG-repeat expansion in the DMPK gene. Sequestration of the splicing factor MBNL1 results in aberrant splicing in many genes in DM1 skeletal muscle, whereas MBNL2 plays a leading role in missplicing in the central nervous system (CNS) of patients with DM1. Splicing misregulation of most MBNL2-regulated genes occurs in the temporal cortex but not in the cerebellum of autopsied patients with DM1. To understand the diversity at macroscopic and microscopic levels in CNS of patients with DM1. Using autopsied brain tissues, we examined alternative splicing ratios of MBNL2-regulated genes and expression levels of potential splicing factors. We found differences in splicing abnormalities among tested regions of the CNS from patients with DM1. In the frontal and temporal cortices and the hippocampus, many genes were aberrantly spliced, but severity differed among the brain regions. By contrast, there were no significant differences in the ratio of splicing variants for most of the genes in the cerebellar cortex and spinal cord between DM1 and control samples. We failed to find any change in the amount of potential factors (MBNL and CUGBP proteins and DMPK mRNA) which explain the modest missplicing in the cerebellum. LASER capture microdissection demonstrated splicing misregulation in the molecular layer of the cerebellum but not in the granular layer. This is the first study to reveal missplicing in a functional cell layer of DM1 and to compare splicing misregulation in a wide region of the CNS using statistical analysis.

Differences in splicing defects between the grey and white matter in myotonic dystrophy type 1 patients

“…We have previously examined the mis-splicing in DM1 patients for each area of the brain and found that mis-splicing in the cerebellum is less apparent than the other areas [3]. We also demonstrated the different degrees of mis-splicing that occurs among cell layers of the cerebellum [7]. These results clearly showed the heterogeneity of the brain and show that detailed splicing analysis for each cell or region is essential to clarify the pathomechanisms of this disorder.…”

Differences in splicing defects between the grey and white matter in myotonic dystrophy type 1

“…We previously examined the mis-splicing in DM1 patients for each area of the brain and found that mis-splicing in the cerebellum is less apparent than in any other area [3]. We also demonstrated the different degrees of mis-splicing that occurs among cell layers of the cerebellum [7]. These results clearly showed the heterogenicity of the brain and show that detailed splicing analysis for each cell or region is essential to clarify the pathomechanisms of this disorder.…”

“…Using NCBI Primer-BLAST, we designed two sets of primers for each spliced gene; exon 4 forward and exon 6 reverse primers for exon 5 splicing, and exon 6 forward and exon 9 reverse primers for exons 7 and 8 splicing. In addition, we examined splicing patterns of other genes controlled by MBNL2: ADD1 exon 15, CACNA1D exon 11, CLASP2 exon 23a and 23b, CSCNK1D exon 9, KCNMA1 exon 27a, TANC2 exon 22a, GRIN1 exon 4, MAPT exon 2 and exon 12, using the primers used in our previous report [3, 7] (Tables 1 and 2).…”

A comprehensive atlas of fetal splicing patterns in the brain of adult myotonic dystrophy type 1 patients

“…DM1-related aberrant splicing has been associated with the expression of fetal transcript variants (17,26,(44)(45)(46)(47). We performed a comprehensive evaluation of the developmental regulation of DM1related exon skipping events by comparing exon inclusion in frontal cortex samples from the human BrainSpan dataset (Figure 1A) and from a recent dataset including 21 DM1 patients and 8 controls (Figure 1B) (23,31).…”