Systematic characterization of BAF mutations explains intra-complex synthetic lethalities

Aberrations in genes coding for subunits of the BAF chromatin remodeling complex occur in human cancers with high frequency. Currently, it is not understood how these loss-of-function mutations contribute to cancer initiation or progression and how they can be targeted therapeutically. Here, we apply an isogenic cell line model of knock-out clones for 22 targetable BAF subunits to investigate the effects of individual subunit loss on complex composition, chromatin accessibility and gene expression. Dependent on the lost subunit, we observe specific alterations in complex composition causing pronounced widespread changes in chromatin accessibility and gene expression. Using this validated cell line system, we evaluated all intra-complex synthetic interactions and identified the sensitivity of SMARCA4 mutant cells to loss of ARID2. These data provide novel insights in the importance of intact BAF complexes for genome-wide chromatin organization and suggest novel approaches to therapeutically target BAF mutant cancers.

Study overview

Arrayed knockout of BAF complex subunits

CRISPR-Cas9 knockout of BAF complex subunits in an arrayed format in isogenic haploid cell lines is throughly validated at protein and transcriptional level.

Mapping of BAF complex composition

Immunoprecipitation of BAF complex subunits followed by mass spectrometry (IP-MS) reveals complex composition changes upon loss of individual subunits.

Synthetic interactions between BAF complex subunits

siRNA screen reveals intra-complex synthetic lethality interactions.

Regulatory interrogation of BAF complex subunit function

Chromatin accessibility changes of cells knockout for each subunit of the BAF complex reveal major functional subunits.

Extent and context of BAF complex chromatin binding revealed by ChIP-seq

Immunoprecipitation of key BAF complex subunits followed by sequencing (ChIP-seq) reveals the breadth of complex binding to chromatin. ChIP-seq allows discovery of BAF-specific and pBAF-specific binding to chromatin. Comprehensive ChIP-seq for histone modifications reveals the chromatin context and likely functions of both complexes.

Transcriptional interrogation of BAF complex subunit function

Transcriptome profiling in all knockout cells reveals downstream effects upon individual subunit loss.

Genome browser tracks

We performed chromatin accessibility mapping by ATAC-seq, chromatin binding mapping by ChIP-seq and transcriptome quantification by RNA-seq on cell lines with single knockout of a BAF complex member.
The following genome browser tracks provide genome-wide maps of the effect of BAF complex mutation:

Click here to add the track hub to UCSC Genome Browser.
Click here to add the tracks to a local IGV Genome Browser session (please launch IGV first, e.g. via this URL).

Data

This section provides access to raw data and analysis results underlying the presented analysis of the BAF complex.

Name	Description and link
Raw and processed NGS data	GEO SuperSeries accession: GSE108390 GEO Series on ATAC-seq data: GSE108386 GEO Series on ChIP-seq data: GSE108387 GEO Series on RNA-seq data: GSE108398
Protein-protein interaction of BAF complex subunits	Comparison of IP-MS abundances between WT and knockout backgrounds: SMARCA4 IP, replicate 1, SMARCA4 IP, replicate 2; ARID1A IP, replicate 1, ARID1A IP, replicate 2
Structural analysis of BAF complex	Input files for structural analysis of BAF complex: SMARCA4 IP data, SMARCA4 IP p-values; ARID1A IP data, ARID1A IP p-values
Chromatin accessiblity of BAF complex subunit knockout cells	Read count quantification of chromatin accessiblity signal in all samples: download here Normalized quantification of chromatin accessiblity signal in all samples: download here Coefficients of change in chromatin accessibility in knockout cells compared to WT: download here
Binding of BAF complex subunits to chromatin	Read count quantification of ChIP-seq signal in all samples: download here Normalized quantification of ChIP-seq signal in all samples: download here BAF- and pBAF-specific bound regions: BAF-specific; pBAF-specific
Transcriptome of BAF complex subunits	Read count quantification of RNA-seq signal in all samples at transcript level: download here Read count quantification of RNA-seq signal in all samples at gene level: download here Normalized quantification of RNA-seq signal in all samples: at gene level download here Coefficients of change in RNA-seq signal in knockout cells compared to WT: download here
Synthetic interaction of BAF complex subunits	Viability measurements of BAF complex knock-out cell lines upon knock-down of additional subunits: download here

Software

To foster reproducibility and facilitate reuse, the source code underlying the analysis is contained in a Git repository at Github.

Citation

If you use these data in your research, please cite:

Sandra Schick, André F. Rendeiro, Kathrin Runggatscher, Anna Ringler, Bernd Boidol, Melanie Hinkel, Peter Májek, Loan Vulliard, Thomas Penz, Katja Parapatics, Christian Schmidl, Jörg Menche, Guido Boehmelt, Mark Petronczki, André Mueller, Christoph Bock, Stefan Kubicek (2019). Systematic characterization of BAF mutations explains intra-complex synthetic lethalities. Nature Genetics. DOI: 10.1038/s41588-019-0477-9