Expanded CUG repeats Dysregulate RNA splicing by altering the stoichiometry of the muscleblind 1 complex

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Myotonic dystrophy type 1 (DM1) is a multisystem disorder characterized by RNA splice defects.
• Muscleblind 1 (MBNL1) is a key splice-regulating protein implicated in DM1 pathogenesis.
• Toxic CUG repeat RNA foci in DM1 nuclei sequester MBNL1 and other proteins.

Purpose of the Study:

• To investigate the role of MBNL1 complexes in DM1-associated RNA splicing abnormalities.
• To characterize the protein composition of MBNL1 complexes in normal and DM1 myoblasts.
• To determine how expanded CUG repeat RNA affects MBNL1 complex stoichiometry and function.

Main Methods:

• Purification of RNA-independent MBNL1 complexes from normal human myoblasts.
• Immunoprecipitation using antibodies targeting MBNL1 variants (MBNL1(CUG)).
• Analysis of MBNL1 complex composition and protein localization in normal and DM1 myoblasts, and in Cos7 cells expressing expanded CUG repeat RNA.

Main Results:

• MBNL1(CUG) complexes in normal myoblasts contain 10 associated proteins involved in ribonucleoprotein remodeling.
• In DM1 myoblasts, the stoichiometry of MBNL1(CUG) complexes is altered, with increased levels of nine partner proteins.
• Expanded CUG repeat RNA expression in Cos7 cells recapitulates these stoichiometric changes.
• Increased levels of specific hnRNPs and DDX5 independently dysregulate splicing in overlapping RNA subsets.

Conclusions:

• Expanded CUG repeat RNA expression in DM1 alters MBNL1(CUG) complex stoichiometry.
• These stoichiometric changes contribute to the reinforcement and expansion of RNA processing defects in DM1.
• Aberrant protein synthesis or stability, rather than PKCα function, underlies the altered MBNL1 complex composition.