Gene Regulation in Neuronal Degeneration - Role of mSin3 and YY1 factors

In neurodegenerative disorders, neural cells can die through two distinctive cell death processes, apoptosis and excitotoxicity. Both of these cell death processes involve changes in mRNA and protein synthesis rates and thus alterations in the level of transcription and translation. Age-related neuronal degeneration involves changes at the transcriptional and protein synthesis level. Transcription factor YY1 and adaptor molecule mSin3 are two proteins which are involved in transcriptional repression. YY1 and mSin3 are units of separate complexes where their repressive function is attributable to several other participants, mostly to histone deacetylases (HDACs). HDACs remove the acetyl groups from the core histone proteins of nucleosomes, and this leads to a conformational change in the nucleosome and further to transcriptional repression. Thus, through HDACs, the YY1 and mSin3 complexes regulate gene expression activity. Our aim was to investigate the changes in the expression of transcripton factor YY1 and adaptor molecules mSin3A/B at the mRNA and protein levels as well as to measure changes in the DNA binding activity of YY1 in neuronal cells and tissues during apoptosis, excitotoxicity and aging. Mouse Neuro-2a neuroblastoma cells, as well as rat primary hippocampal and cerebellar granule cells were used in the apoptosis and excitotoxicity studies. Tissue samples were taken from young and old rats. Several apoptotic inducers increased the mSin3A protein levels, but mSin3B levels remained unchanged. Trichostatin A (TSA), a specific inhib itor of HDACs, induced the strongest increase in the mSin3A protein expression. All apoptotic inducers significantly increased the caspase-3 activities in Neuro-2a cells, whereas etoposide did not induce any caspase-3 activity in primary cerebellar granule cells. Caspase-3 activity is considered to be a reliable marker of apoptosis. A brief exposure of primary neuronal cells to glutamate induced a dramatic change in the size of the binding complex of YY1 to its DNA binding site. The binding of the normal YY1 complex present in control samples disappeared while a new, smaller complex appeared. However, both complexes shifted to the same level with anti-YY1 antibody in EMSA-supershift assays. Interestingly, also okadaic acid -induced apoptosis caused similar changes in the DNA binding complexes of YY1 as did glutamate-induced excitotoxicity. These results indicate that apoptosis and excitotoxic shock in neuronal cells can change the protein levels of mSin3A and YY1. Also the binding complexes of YY1 were disrupted in excitotoxicity and okadaic acid induced apoptosis, highlighting the role of changes in YY1 transcriptional regulation. The changes seen in the DNA binding complexes of YY1 may be due to the glutamate induced Ca influx, which further activates several signalling cascades in the cell, or okadaic acid induced changes in the phosphorylation status of the cellular proteins. Aging in vivo did not have any effect on either the expression of mRNA or protein of mSin3A/B and YY1 or on the complex formation of YY1 in rat brain samples. Thus, aging in vivo does not seem to have a role in the regulation of mSin3 and YY1, although protein synthesis slows down during aging. As a conclusion, apoptosis and excitotoxicity, but not aging, affected the transcriptional repressor factors mSin3A and YY1 indicating that the roles of those protein change during neuronal degeneration. National Library of Medicine Classification: WL 359, WL 102.5 Medical Subject Headings: transcription, genetic; apoptosis; aging; neurons; neuroblastoma/cytology; gene expression; blotting, Northern; blotting, Western; electrophoretic mobility assay; transcription factors; excitatory amino acids; gene expression regulation; down-regulation/genetics; DNA-binding proteins/genetics To Jarmo, Akuliina and Aukusti