Defective myelination in an RNA polymerase III mutant leukodystrophic mouse

Abstract

RNA polymerase (Pol) III synthesizes abundant short non-coding RNAs that have essential functions in protein synthesis, secretion and other processes. Despite the ubiquitous functions of these RNAs, mutations in Pol III subunits cause Pol III-related leukodystrophy, an early-onset neurodegenerative disease. The basis of this neural sensitivity and the mechanisms of disease pathogenesis are unknown. Here we show that mice expressing pathogenic mutations in the largest Pol III subunit, Polr3a, specifically in Olig2-expressing cells, have impaired growth, neurobehavioral deficits and hypomyelination in multiple regions of the brain and spinal cord. We identify two mechanisms of disease pathogenesis within the oligodendrocyte (OL) lineage; a differentiation defect of oligodendrocyte precursor cells reduces the number of mature myelinating OLs and an intrinsic myelination defect in mature OLs impairs myelin deposition. Thus, we reveal cell-specific roles for Pol III in the normal development and function of oligodendrocytes.

POLR3-Related Leukodystrophy: Exploring Potential Therapeutic Approaches

“…Third, attempts to produce a representative animal model of POLR3-HLD in which to perform pre-clinical testing have proven difficult (Choquet et al, 2017(Choquet et al, , 2019b, and this barrier will need to be overcome to properly test any novel therapeutic candidate. Recently, progress in generating an animal model has been made using an Olig2-Cre conditional double Polr3a mutant knock-in strategy, in a recent pre-print which has yet to be peer-reviewed at the time of writing (Merheb et al, 2020). Overcoming these challenges would elucidate the potential for POLR3-HLD gene transfer, and will also inform the future development of more advanced and/or personalized (e.g., gene editing) therapeutic strategies for POLR3-HLD.…”

“…As the genes associated with POLR3-HLD have been discovered relatively recently and attempts at generating an animal model were predominantly unsuccessful (Choquet et al, 2017(Choquet et al, , 2019b, the cellular and molecular mechanisms underlying the white matter pathology of this disease are largely unknown. Research is ongoing regarding the investigation of the pathophysiology of POLR3-HLD; recent modeling of the disease has been accomplished in yeast (Moir et al, 2020), as well as in a conditional mouse model (pre-print data, not yet peer-reviewed; Merheb et al, 2020). Moreover, a variety of different types of pathogenic variants are known to cause POLR3-HLD, including nonsense, missense, intronic, synonymous, and splice site variants, as well as large exonic deletions, and small insertions or deletions (Bernard et al, 2011;Tétreault et al, 2011;Potic et al, 2012;Terao et al, 2012;Daoud et al, 2013;Takanashi et al, 2014;Wolf et al, 2014b;Gutierrez et al, 2015;Thiffault et al, 2015;La Piana et al, 2016;Jurkiewicz et al, 2017;Richards et al, 2017;Al Yazidi et al, 2019;Gauquelin et al, 2019;Harting et al, 2020;Hiraide et al, 2020b;Perrier et al, 2020).…”

RNA Polymerase III Subunit Mutations in Genetic Diseases

“…The second approach was more successful and generated the first mouse model demonstrating hypomyelination as seen in POLR3-HLD, but with a very mild phenotype and absent motor features (pre-print on https://www.biorxiv.org/content/10.1101/ 2020.12.09.418657v2, currently under peer review at the time this manuscript is written) (Merheb et al, 2021). To achieve this, the Willis laboratory first screened a panel of POLR3A HLD mutations by introducing them in the S. cerevisiae orthologous gene, Rpc160, focusing on a cluster of mutations in the pore region of Pol III, which included Gly672Glu (Moir et al, 2021).…”

“…In vitro transcription experiments demonstrated a defect in both factor-independent and factor-dependent transcription for genes representative of the yeast Pol III transcriptome in this double mutant (Moir et al, 2021). Next, the authors generated mice with the corresponding human double allele Trp671Arg/Gly672Glu (Merheb et al, 2021). Since homozygosity for the whole-body KI of this allele was embryonic lethal, a conditional KI mouse was engineered using an Olig2-Cre driver, directing expression of the mutant allele throughout the oligodendrocyte lineage and in a subset of other CNS cells (Merheb et al, 2021).…”

“…Next, the authors generated mice with the corresponding human double allele Trp671Arg/Gly672Glu (Merheb et al, 2021). Since homozygosity for the whole-body KI of this allele was embryonic lethal, a conditional KI mouse was engineered using an Olig2-Cre driver, directing expression of the mutant allele throughout the oligodendrocyte lineage and in a subset of other CNS cells (Merheb et al, 2021). Homozygous conditional KI mice displayed growth defects, neurobehavioral deficits and impaired myelination, myelin integrity and oligodendrogliogenesis.…”