Leukodystrophy-associated POLR3A mutations down-regulate the RNA polymerase III transcript and important regulatory RNA BC200

J Biol Chem. 2019 May 3;294(18):7445-7459. doi: 10.1074/jbc.RA118.006271. Epub 2019 Mar 21.


RNA polymerase III (Pol III) is an essential enzyme responsible for the synthesis of several small noncoding RNAs, a number of which are involved in mRNA translation. Recessive mutations in POLR3A, encoding the largest subunit of Pol III, cause POLR3-related hypomyelinating leukodystrophy (POLR3-HLD), characterized by deficient central nervous system myelination. Identification of the downstream effectors of pathogenic POLR3A mutations has so far been elusive. Here, we used CRISPR-Cas9 to introduce the POLR3A mutation c.2554A→G (p.M852V) into human cell lines and assessed its impact on Pol III biogenesis, nuclear import, DNA occupancy, transcription, and protein levels. Transcriptomic profiling uncovered a subset of transcripts vulnerable to Pol III hypofunction, including a global reduction in tRNA levels. The brain cytoplasmic BC200 RNA (BCYRN1), involved in translation regulation, was consistently affected in all our cellular models, including patient-derived fibroblasts. Genomic BC200 deletion in an oligodendroglial cell line led to major transcriptomic and proteomic changes, having a larger impact than those of POLR3A mutations. Upon differentiation, mRNA levels of the MBP gene, encoding myelin basic protein, were significantly decreased in POLR3A-mutant cells. Our findings provide the first evidence for impaired Pol III transcription in cellular models of POLR3-HLD and identify several candidate effectors, including BC200 RNA, having a potential role in oligodendrocyte biology and involvement in the disease.

Keywords: CRISPR/Cas; RNA polymerase III; RNA-seq; brain cytoplasmic 200 RNA (BCYRN1); leukodystrophy; myelin; oligodendrocyte; proteomics; transcription; transfer RNA (tRNA).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Down-Regulation / genetics*
  • Genes, Recessive
  • HeLa Cells
  • Hereditary Central Nervous System Demyelinating Diseases / genetics*
  • Humans
  • Mutation*
  • RNA Polymerase III / genetics*
  • RNA, Long Noncoding / genetics*
  • RNA, Messenger / genetics*


  • RNA, Long Noncoding
  • RNA, Messenger
  • long non-coding RNA BC200, human
  • POLR3A protein, human
  • RNA Polymerase III