Interplay between FACT subunit SPT16 and TRIM33 can remodel chromatin at macrophage distal regulatory elements

Cell type-specific use of cis-acting regulatory elements is mediated by the combinatorial activity of transcription factors involved in lineage determination and maintenance of cell identity. In macrophages, specific transcriptional programs are dictated by the transcription factor PU.1 that primes distal regulatory elements for macrophage identities and makes chromatin competent for activity of stimuli-dependent transcription factors. Although the advances in genome-wide approaches have elucidated the functions of these macrophage-specific distal regulatory elements in transcriptional responses, chromatin structures associated with PU.1 priming and the underlying mechanisms of action of these cis-acting sequences are not characterized.

Here, we show that, in macrophages, FACT subunit SPT16 can bind to positioned nucleosomes directly flanking PU.1-bound sites at previously uncharacterized distal regulatory elements located near genes essential for macrophage development and functions. SPT16 can interact with the transcriptional co-regulator TRIM33 and binds to half of these sites in a TRIM33-dependent manner. Using the Atp1b3 locus as a model, we show that FACT binds to two positioned nucleosomes surrounding a TRIM33/PU.1-bound site in a region, located 35 kb upstream the Atp1b3 TSS, that interact with the Atp1b3 promoter. At this - 35 kb region, TRIM33 deficiency leads to FACT release, loss of the two positioned nucleosomes, RNA Pol II recruitment and bidirectional transcription. These modifications are associated with higher levels of FACT binding at the Atp1b3 promoter, an increase of RNA Pol II recruitment and an increased expression of Atp1b3 in Trim33^-/- macrophages.

Thus, sequestering of SPT16/FACT by TRIM33 at PU.1-bound distal regions might represent a new regulatory mechanism for RNA Pol II recruitment and transcription output in macrophages.