AbstractMethyl CpG-binding protein 2 (MeCP2), the mutated protein in Rett syndrome (RTT), is a crucial chromatin-modifying and gene-regulatory protein that has two main isoforms (MeCP2_E1 and MeCP2_ E2) due to the alternative splicing and switching between translation start codons in exons one and two. Functionally, these two isoforms appear to be virtually identical; however, evidence suggests that only MeCP2_E1 is relevant to RTT, including a single RTT missense mutation in exon 1, p.Ala2Val. Here, we show that N-terminal co- and post- translational modifications differ for MeCP2_E1, MeCP2_E1-p.Ala2Val and MeCP2_E2, which result in different protein degradation rates in vitro. We report partial N-methionine excision (NME) for MeCP2_E2, whereas NME for MeCP2_E1 is complete. Surprisingly, we also observed evidence of excision of multiple alanine residues from the N-terminal polyalanine stretch. Regarding MeCP2_E1-Ala2Val, we also observed only partial NME and N-acetylation (NA) of either methionine or valine. The localization of MeCP2_E1 and co-localization with chromatin appear to be unaffected by the p.Ala2Val mutation. However, a higher proteasomal degradation rate was observed for MeCP2_E1-Ala2Val compared with that for wild type (WT) MeCP2_E1. Thus, the etiopathology of p.Ala2Val is likely due to a reduced bio-availability of MeCP2 because of the faster degradation rate of the unmodified defective protein. MeCP2_E1 is thought to have a much higher translational efficiency than MeCP2_E2. Our data suggest that this increased efficiency may be balanced by a higher degradation rate. The higher turnover rate of the MeCP2_E1 protein suggests that it may play a more dynamic role in cells than MeCP2_E2.Significance statementThe Rett syndrome protein, MeCP2, undergoes a number of modifications before becoming functionally active in the body’s cells. Here, we report the presence of N-terminal modifications in both MeCP2 isoforms, MeCP2_E1 and MeCP2_E2, and that the only reported Rett missense mutation in exon 1, p.Ala2Val, disrupts these modifications, decreasing the longevity of the protein. Interestingly, p.Ala2Val mutations have been reported in many other disease genes, such as DKCX, ECHS1, IRF6, SMN1, and TNNI3, and the etiopathological mechanism(s) have never been explained. Thus, this work is important not only for the understanding of the pathophysiology of Rett syndrome but also for a deeper understanding of the effects of genetic mutations at the N-terminal end of genes in general.
A novel DAX1/NR0B1 mutation in a patient with adrenal hypoplasia congenita and hypogonadotropic hypogonadism