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Protein Sci. 2017 May;26(5):997-1011. doi: 10.1002/pro.3142. Epub 2017 Mar 12.

Stepwise assembly of functional C-terminal REST/NRSF transcriptional repressor complexes as a drug target.

Author information

1
Department of Biomedicine, Aarhus University, DK-8000 Aarhus C, Denmark.
2
Centre for Stochastic Geometry and Advanced Bioimaging, Aarhus University, DK-8000 Aarhus C, Denmark.
3
Institute of Human Genetics, Hannover Medical School, D-30625 Hannover, Germany.

Abstract

In human cells, thousands of predominantly neuronal genes are regulated by the repressor element 1 (RE1)-silencing transcription factor/neuron-restrictive silencer factor (REST/NRSF). REST/NRSF represses transcription of these genes in stem cells and non-neuronal cells by tethering corepressor complexes. Aberrant REST/NRSF expression and intracellular localization are associated with cancer and neurodegeneration in humans. To date, detailed molecular analyses of REST/NRSF and its C-terminal repressor complex have been hampered largely by the lack of sufficient amounts of purified REST/NRSF and its complexes. Therefore, the aim of this study was to express and purify human REST/NRSF and its C-terminal interactors in a baculovirus multiprotein expression system as individual proteins and coexpressed complexes. All proteins were enriched in the nucleus, and REST/NRSF was isolated as a slower migrating form, characteristic of nuclear REST/NRSF in mammalian cells. Both REST/NRSF alone and its C-terminal repressor complex were functionally active in histone deacetylation and histone demethylation and bound to RE1/neuron-restrictive silencer element (NRSE) sites. Additionally, the mechanisms of inhibition of the small-molecule drugs 4SC-202 and SP2509 were analyzed. These drugs interfered with the viability of medulloblastoma cells, where REST/NRSF has been implicated in cancer pathogenesis. Thus, a resource for molecular REST/NRSF studies and drug development has been established.

KEYWORDS:

CoREST; DNA transcription; HDAC1; LSD1; REST/NRSF; drug mechanism; histone deacetylation; histone demethylation; medulloblastoma; transcriptional repression

PMID:
28218430
PMCID:
PMC5405421
DOI:
10.1002/pro.3142
[Indexed for MEDLINE]
Free PMC Article

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