We recently demonstrated the utility of induced pluripotent stem cells to investigate the functional consequences of mutations in the gene encoding the Methyl CpG binding protein-2 (MeCP2) on neurons from RTT patients, a syndromic form of ASD [11
]. RTT patients appear to develop normally for up to 6–18 months, after which they enter a period of regression characterized by deceleration of head growth and loss of acquired motor and language skills. Patients often develop autistic behaviors, stereotypic hand wringing, abnormal breathing and seizures [22
]. Postmortem analyses of affected individuals have revealed phenotypes at the cellular level, including decreased soma size, reduced dendritic branching, and altered dendritic spine density and morphology [23
MeCP2 is an abundant nuclear protein implicated in a number of molecular functions, but was first identified as an epigenetic regulator of target genes by binding to methylated CpG dinucleotides [25
]. Long established as a repressor of transcription, MeCP2 has recently been shown to bind active genes as well as to influence RNA splicing [22
]. MeCP2 is found in a wide variety of tissues but appears to be most abundant in the brain [27
]. Accordingly, MeCP2 appears to be critical for normal CNS development and function, and its dysfunction results in abnormal neurological phenotypes ().
Figure 1 Phenotypes demonstrated by human iPSC-derived RTT neurons. We generated a human cellular model for RTT by reprogramming the fibroblasts of human patients to iPSCs. Neurons differentiated from these iPSCs exhibited cellular phenotypes such as (a) smaller (more ...)
Neurons derived from RTT-iPSCs carrying four different MeCP2 mutations showed several alterations compared to five healthy non-affected individuals, such as decreased soma size, altered dendritic spine density and reduced excitatory synapses. These phenotypes were validated using wild-type MeCP2 cDNA and specific shRNAs against MeCP2 in gain-and-loss of function experiments to demonstrate causality. Importantly, some of these cellular defects were immediately validated by independent groups, revealing the robustness and reproducibility of the system [12
]. We were able to rescue the defects in the number of glutamatergic synapses using two candidate drugs, insulin-like growth factor 1 (IGF1) and gentamicin. IGF1 is considered to be a candidate for pharmacological treatment of RTT and potentially other CNS disorders in ongoing clinical trials [28
]. Gentamicin, a read-through drug, was also used to rescue neurons carrying a nonsense MeCP2 mutation, by elevating the amount of MeCP2 protein.
Moreover, we took advantage of the RTT-iPSCs to demonstrate that neural progenitor cells carrying MeCP2 mutations have increased susceptibility for L1 retrotransposition. Long interspersed nuclear elements-1 (LINE-1 or L1s) are abundant retrotransposons that comprise approximately 20% of mammalian genomes [29
] and are highly active in the nervous system [32•
]. Our data demonstrate that L1 retrotransposition can be controlled in a tissue-specific manner and that disease-related genetic mutations can influence the frequency of neuronal L1 retrotransposition [34
]. The work revealed an unexpected and novel phenomenon, adding a new layer of complexity to the understanding of genomic plasticity. These observations bring valued information for RTT and, potentially, other ASD patients, since they suggest that pre-symptomatic defects may represent novel biomarkers to be exploited as diagnostic tools. The data also suggest that early intervention may be beneficial.