SUMOylated non-canonical polycomb PRC1.6 complex as a prerequisite for recruitment of transcription factor RBPJ

Małgorzata Sotomska,Francesca Ferrante,Heiko Schwederski,Robert Liefke,Tilman Borggrefe,Franz Oswald

doi:10.1186/s13072-021-00412-9

Abstract

BackgroundNotch signaling controls cell fate decisions in many contexts during development and adult stem cell homeostasis and, when dysregulated, leads to carcinogenesis. The central transcription factor RBPJ assembles the Notch coactivator complex in the presence of Notch signaling, and represses Notch target gene expression in its absence.ResultsWe identified L3MBTL2 and additional members of the non-canonical polycomb repressive PRC1.6 complex in DNA-bound RBPJ associated complexes and demonstrate that L3MBTL2 directly interacts with RBPJ. Depletion of RBPJ does not affect occupancy of PRC1.6 components at Notch target genes. Conversely, absence of L3MBTL2 reduces RBPJ occupancy at enhancers of Notch target genes. Since L3MBTL2 and additional members of the PRC1.6 are known to be SUMOylated, we investigated whether RBPJ uses SUMO-moieties as contact points. Indeed, we found that RBPJ binds to SUMO2/3 and that this interaction depends on a defined SUMO-interaction motif. Furthermore, we show that pharmacological inhibition of SUMOylation reduces RBPJ occupancy at Notch target genes.ConclusionsWe propose that the PRC1.6 complex and its conjugated SUMO-modifications provide a favorable environment for binding of RBPJ to Notch target genes.

Highlights

Notch signaling controls cell fate decisions in many contexts during development and adult stem cell homeostasis and, when dysregulated, leads to carcinogenesis
We reveal that RBPJ utilizes its Small ubiquitin-like modifier (SUMO) interaction motif as additional docking site and provide evidence that high local SUMO levels and occupancy of the PRC1.6 complex are favorable for RBPJ binding
DNA‐bound RBPJ is associated with several PRC1.6 components To biochemically isolate RBPJ-containing complexes, we took advantage of a DNA double-stranded oligonucleotide containing two RBPJ binding sites 5′-GTGGGAA-3’ (Fig. 1a), which allows the formation of DNA-bound dimeric RBPJ complexes [27]

Summary

Introduction

Notch signaling controls cell fate decisions in many contexts during development and adult stem cell homeostasis and, when dysregulated, leads to carcinogenesis. The central transcription factor RBPJ assembles the Notch coactivator complex in the presence of Notch signaling, and represses Notch target gene expression in its absence. Notch signal transduction is an evolutionary conserved pathway that regulates stem cell maintenance and differentiation decisions throughout development. Dysregulation of either NOTCH receptors or their modifiers are linked to carcinogenesis [1,2,3]. Ligand binding leads to the proteolytic processing of the Notch receptor. The central player controlling the expression of Notch target genes is the transcription factor RBPJ, known as CBF1 (C promoter binding factor) or CSL (Homo sapiens CBF1, Drosophila melanogaster Suppressor of Hairless, and Caenorhabditis elegans Lag-1). In the absence of a Notch signal, RBPJ assembles a corepressor complex containing NCoR/HDACs [13] and histone demethylases, such as KDM1A/LSD1 [14]

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Conclusion