Biology subjects

Ausio, J.

Publications and source records attributed to Ausio, J..

3 recordsLinked to original sources

bioRxiv2018

MeCP2-E1 isoform is a dynamically expressed, weakly DNA-bound protein with different protein and DNA interactions compared to MeCP2-E2

MeCP2 - a chromatin-binding protein associated with Rett syndrome - has two main isoforms, MeCP2-E1 and MeCP2-E2, with 96% amino acid identity differing in a few N-terminal amino acid residues. Previous studies have shown brain region-specific expression of these isoforms which, in addition to their different cellular localization and differential expression during brain development, suggest they may also have non-overlapping molecular mechanisms. However, differential functions of MeCP2-E1 and E2 remain largely unexplored. Here, we show that the N-terminal domains (NTD) of MeCP2-E1 and E2 modulate the ability of the methyl binding domain (MBD) to interact with DNA as well as influencing the turnover rates, binding dynamics, response to nuclear depolarization, and circadian oscillations of the two isoforms. Our proteomics data indicate that both isoforms exhibit unique interacting protein partners. Moreover, genome-wide analysis using ChIP-seq provide evidence for a shared as well as a specific regulation of different sets of genes. Our findings provide insight into the functional complexity of MeCP2 by dissecting differential aspects of its two isoforms.\n\nSignificanceWhether the two E1 and E2 isoforms of MeCP2 have different structural and/or functional implications has been highly controversial and is not well known. Here we show that the relatively short N-terminal sequence variation between the two isoforms impinges them with an important DNA binding difference. Moreover, MeCP2-E1 and E2 exhibit a different cellular dynamic behavior and have some distinctive interacting partners. In addition, while sharing genome occupancy they specifically bind to several distinctive genes.

biochemistry↗

bioRxiv2018

A novel histone H4 variant regulates rDNA transcription in breast cancer

Histone variants, present in various cell types and tissues, are known to exhibit different functions. For example, histone H3.3 and H2A.Z are both involved in gene expression regulation, whereas H2A.X is a specific variant that responds to DNA double-strand breaks. In this study, we characterized H4G, a novel hominidae-specific histone H4 variant. H4G expression was found in a variety of cell lines and was particularly overexpressed in the tissues of breast cancer patients. H4G was found to localize primarily to the nucleoli of the cell nucleus. This localization was controlled by the interaction of the alpha helix 3 of the histone fold motif with the histone chaperone, nucleophosphomin 1. In addition, we found that H4G nucleolar localization increased rRNA levels, protein synthesis rates, and cell cycle progression. Furthermore, micrococcal nuclease digestion of H4G-containing nucleosomes reconstituted in vitro indicated that H4G destabilizes the nucleosome, which may serve to alter nucleolar chromatin in a way that enhances rDNA transcription in breast cancer tissues.

molecular biology↗

bioRxiv2017

The role of N-terminal modification of MeCP2 in the pathophysiology of Rett syndrome

Methyl 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.\n\nSignificance statementThe Rett syndrome protein, MeCP2, undergoes a number of modifications before becoming functionally active in the bodys 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.

genetics↗