Abstract
The HPC-1/syntaxin 1A (Stx1a) gene, which is involved in synaptic transmission and neurodevelopmental disorders, is a TATA-less gene with several transcription start sites. It is activated by the binding of Sp1 and acetylated histone H3 to the −204 to +2 core promoter region (CPR) in neuronal cell/tissue. Furthermore, it is depressed by the association of class 1 histone deacetylases (HDACs) to Stx1a–CPR in non-neuronal cell/tissue. To further clarify the factors characterizing Stx1a gene silencing in non-neuronal cell/tissue not expressing Stx1a, we attempted to identify the promoter region forming DNA–protein complex only in non-neuronal cells. Electrophoresis mobility shift assays (EMSA) demonstrated that the −183 to −137 OL2 promoter region forms DNA–protein complex only in non-neuronal fetal rat skin keratinocyte (FRSK) cells which do not express Stx1a. Furthermore, the Yin-Yang 1 (YY1) transcription factor binds to the −183 to −137 promoter region of Stx1a in FRSK cells, as shown by competitive EMSA and supershift assay. Chromatin immunoprecipitation assay revealed that YY1 in vivo associates to Stx1a–CPR in cell/tissue not expressing Stx1a and that trichostatin A treatment in FRSK cells decreases the high-level association of YY1 to Stx1a-CPR in default. Reporter assay indicated that YY1 negatively regulates Stx1a transcription. Finally, mass spectrometry analysis showed that gene silencing factors, including HDAC1, associate onto the −183 to −137 promoter region together with YY1. The current study is the first to report that Stx1a transcription is negatively regulated in a cell/tissue-specific manner by YY1 transcription factor, which binds to the −183 to −137 promoter region together with gene silencing factors, including HDAC.
Highlights
HPC-1/syntaxin 1A (Stx1a) was first discovered as a gene exhibiting neuron-specific expression and is a key acceptor molecule of a complex called soluble N-ethylmaleimidesensitive fusion protein attachment protein receptor (SNARE), which is involved in presynaptic vesicle exocytosis in the central nervous system (CNS) [1,2,3,4]
We demonstrated that Stx1a gene transcription is negatively regulated in a cell-/tissue-specific manner by the Yin-Yang 1 (YY1) transcription factor and that YY1 binds to the −183 to −137 promoter region together with factors participating in gene silencing, such as histone deacetylases (HDACs)
D10392) promotes Stx1a transcription in neuronal cell/tissue; we showed that the association of class 1 HDACs, such as HDAC1, HDAC2 and HDAC8, with Stx1a–core promoter region (CPR) negatively regulates Stx1a transcription in non-neuronal cell/tissue [8]
Summary
HPC-1/syntaxin 1A (Stx1a) was first discovered as a gene exhibiting neuron-specific expression and is a key acceptor molecule of a complex called soluble N-ethylmaleimidesensitive fusion protein attachment protein receptor (SNARE), which is involved in presynaptic vesicle exocytosis in the central nervous system (CNS) [1,2,3,4]. Sp1 and acetylated histone H3 (AcH3) to the −204 to +2 core promoter region (CPR) in neuronal cell and tissue [8]. Biomolecules 2021, 11, 146 is determined by gene silencing via the binding of class 1 histone deacetylases (HDACs), such as HDAC1, HDAC2 and HDAC8, to Stx1a–CPR in non-neuronal cell and tissue [8,9]. The detailed transcription mechanism involved in Stx1a gene silencing is unclear. To further clarify the factors characterizing Stx1a gene silencing in non-neuronal cell and tissue, in the present study, we first attempted to identify the promoter region forming DNA–protein complex only in non-neuronal cells by electrophoresis mobility shift assays (EMSA). EMSA demonstrated that DNA–protein complex is formed on the −183 to
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