Differential somatic CAG repeat instability in variable brain cell lineage in dentatorubral pallidoluysian atrophy (DRPLA): a laser-captured microdissection (LCM)-based analysis

Abstract

Employing a laser-captured microdissection (LCM), we have investigated the somatic instability of CAG repeats in the variable brain cell lineage in three patients with dentatorubral pallidoluysian atrophy (DRPLA). LCM enables the isolation of single lineage brain cells for subsequent molecular analysis. We have found that CAG repeat size and the range of CAG repeats in the cerebellar granular cells is smaller than those in cerebellar glial cells. Similarly, those in the cerebral neuronal cells are significantly shorter than those in cerebral glial cells. These data directly indicate that the CAG repeat is relatively more stable in neuronal cells than in glial cells. Furthermore, cerebellar granular cells show significantly smaller main CAG repeat size and CAG repeat range than either Purkinje cells or cerebral neuronal cells, suggesting that somatic instability in the CAG repeat is markedly variable even among the different types of neuronal populations. The cell-specific CAG repeat instability may thus be more complex than has previously been considered. LCM is a powerful tool for elucidating the mechanism of the triplet repeat instability of each cell type.

A comparison of the lengths of androgen receptor triplet repeats in brain and blood in motor neuron diseases

“…The possibility remains that the variation in AR repeat lengths is restricted to only a subset of cells, such as motor neurons or glia, since cell-type differences in triplet repeat lengths have been described in the brain [15,16]. Such a study would require laser microdissection to obtain a pure sample of motor neurons [4].…”

Isolated short CTG/CAG DNA slip-outs are repaired efficiently by hMutSβ, but clustered slip-outs are poorly repaired

“…The observations that slip-outs of one to three excess repeats require MutSβ for repair and that perturbation of MutSβ levels can reduce repair suggest that the increment of change during a single mutagenic step would be one to three excess repeats. Limited evidence from patient tissues supports incremental changes of 1 to 3 CTG/CAG units per mutation event (48)(49)(50)(51), similar to that occurring at other simple repeats such as (CA)n and (A)n (52,53).…”

Hijacking of the mismatch repair system to cause CAG expansion and cell death in neurodegenerative disease

“…Although there are notable exceptions [69][70], most CAG tracts in proliferating HD patient fibroblasts are also stable or delete trinucleotide repeats even in the presence of an intact MMR system [71][72][73]. Thus, a simple replication slippage model is not entirely consistent with the requirement of MSH2/MSH3 in causing expansion.…”

“…Thus, TNR expansion requires MMR, but does not require enzymes generally needed for homologous recombination or non-homologous end-joining in animals. Since expansion occurs in post-mitotic neurons of HD animals and in DRPLA patients, these results suggest that replication is also not required for expansion and, most often, results in deletion [43,73]. Exclusion of these mechanisms indicates that expansion in animals may arise in the context of single strand break repair, as a consequence of base excision repair [BER], or potentially from nucleotide excision repair [NER][ Figure 7].…”