Recurrent SMARCB1 Mutations Reveal a Nucleosome Acidic Patch Interaction Site That Potentiates mSWI/SNF Complex Chromatin Remodeling.

Mammalian switch/sucrose non-fermentable (mSWI/SNF) complexes are multi-component machines that remodel chromatin architecture. Dissection of the subunit- and domain-specific contributions to complex activities is needed to advance mechanistic understanding. Here, we examine the molecular, structural, and genome-wide regulatory consequences of recurrent, single-residue mutations in the putative coiled-coil C-terminal domain (CTD) of the SMARCB1 (BAF47) subunit, which cause the intellectual disability disorder Coffin-Siris syndrome (CSS), and are recurrently found in cancers. We find that the SMARCB1 CTD contains a basic α helix that binds directly to the nucleosome acidic patch and that all CSS-associated mutations disrupt this binding. Furthermore, these mutations abrogate mSWI/SNF-mediated nucleosome remodeling activity and enhancer DNA accessibility without changes in genome-wide complex localization. Finally, heterozygous CSS-associated SMARCB1 mutations result in dominant gene regulatory and morphologic changes during iPSC-neuronal differentiation. These studies unmask an evolutionarily conserved structural role for the SMARCB1 CTD that is perturbed in human disease.

Synthesis of Oriented Hexasomes and Asymmetric Nucleosomes Using a Template Editing Process.

Nucleosomes, the structural building blocks of chromatin, possess 2-fold pseudo symmetry which can be broken through differential modification or removal of one copy of a pair of sister histones. The resultant asymmetric nucleosomes and hexasomes have been implicated in gene regulation, yet the use of these noncanonical substrates in chromatin biochemistry is limited, owing to the lack of efficient methods for their preparation. Here, we report a strategy that allows the orientation of these asymmetric species to be tightly controlled relative to the underlying DNA sequence. Our approach is based on the use of truncated DNA templates to assemble oriented hexasomes followed by DNA ligation and, in the case of asymmetric nucleosomes, addition of the missing heterotypic histones. We show that this approach is compatible with multiple nucleosome positioning sequences, allowing the generation of desymmetrized mononucleosomes and oligonucleosomes with varied DNA overhangs and heterotypic histone H2A/H2B dimer compositions. Using this technology, we examine the functional consequences of asymmetry on BRG1/BRM associated factor (BAF) complex-mediated chromatin remodeling. Our results indicate that cancer-associated histone mutations can reprogram the inherent activity of BAF chromatin remodeling to induce aberrant chromatin structure.

Acidic patch histone mutations and their effects on nucleosome remodeling.

Structural and biochemical studies have identified a histone surface on each side of the nucleosome disk termed 'the nucleosome acidic patch' that acts as a regulatory hub for the function of numerous nuclear proteins, including ATP-dependent chromatin complexes (remodelers). Four major remodeler subfamilies, SWI/SNF, ISWI, CHD, and INO80, have distinct modes of interaction with one or both nucleosome acidic patches, contributing to their specific remodeling outcomes. Genome-wide sequencing analyses of various human cancers have uncovered high-frequency mutations in histone coding genes, including some that map to the acidic patch. How cancer-related acidic patch histone mutations affect nucleosome remodeling is mainly unknown. Recent advances in in vitro chromatin reconstitution have enabled access to physiologically relevant nucleosomes, including asymmetric nucleosomes that possess both wild-type and acidic patch mutant histone copies. Biochemical investigation of these substrates revealed unexpected remodeling outcomes with far-reaching implications for alteration of chromatin structure. This review summarizes recent findings of how different remodeler families interpret wild-type and mutant acidic patches for their remodeling functions and discusses models for remodeler-mediated changes in chromatin landscapes as a consequence of acidic patch mutations.

A basic motif anchoring ISWI to nucleosome acidic patch regulates nucleosome spacing.

Recent studies have implicated the nucleosome acidic patch in the activity of ATP-dependent chromatin remodeling machines. We used a photocrosslinking-based nucleosome profiling technology (photoscanning) to identify a conserved basic motif within the catalytic subunit of ISWI remodelers, SNF2h, which engages this nucleosomal epitope. This region of SNF2h is essential for chromatin remodeling activity in a reconstituted biochemical system and in cells. Our studies suggest that the basic motif in SNF2h plays a critical role in anchoring the remodeler to the nucleosomal surface. We also examine the functional consequences of several cancer-associated histone mutations that map to the nucleosome acidic patch. Kinetic studies using physiologically relevant heterotypic nucleosomal substrates ('Janus' nucleosomes) indicate that these cancer-associated mutations can disrupt regularly spaced chromatin structure by inducing ISWI-mediated unidirectional nucleosome sliding. These results indicate a potential mechanistic link between oncogenic histones and alterations to the chromatin landscape.

Hydromethylation of Unactivated Olefins.

A solution to the classic unsolved problem of olefin hydromethylation is presented. This highly chemoselective method can tolerate labile and reactive chemical functionalities and uses a simple set of reagents. An array of olefins, including mono-, di-, and trisubstituted olefins, are all smoothly hydromethylated. This mild protocol can be used to simplify the synthesis of a specific target or to directly "edit" complex natural products and other advanced materials. The method is also amenable to the simple installation of radioactive and stable labeled methyl groups.

Quinone diazides for olefin functionalization.

The utility of quinone diazides in materials science is vast and well-documented, yet this potentially useful motif has languished in the annals of organic synthesis. Herein we show that modern tools of catalysis can be employed with free or suitably masked quinone diazides to unleash the power of these classic diazo compounds in the context of both inter- and intramolecular olefin cyclopropanation.

Chromatin landscape signals differentially dictate the activities of mSWI/SNF family complexes.

Mammalian SWI/SNF (mSWI/SNF) adenosine triphosphate-dependent chromatin remodelers modulate genomic architecture and gene expression and are frequently mutated in disease. However, the specific chromatin features that govern their nucleosome binding and remodeling activities remain unknown. We subjected endogenously purified mSWI/SNF complexes and their constituent assembly modules to a diverse library of DNA-barcoded mononucleosomes, performing more than 25,000 binding and remodeling measurements. Here, we define histone modification-, variant-, and mutation-specific effects, alone and in combination, on mSWI/SNF activities and chromatin interactions. Further, we identify the combinatorial contributions of complex module components, reader domains, and nucleosome engagement properties to the localization of complexes to selectively permissive chromatin states. These findings uncover principles that shape the genomic binding and activity of a major chromatin remodeler complex family.

The nucleosome acidic patch and H2A ubiquitination underlie mSWI/SNF recruitment in synovial sarcoma.

Interactions between chromatin-associated proteins and the histone landscape play major roles in dictating genome topology and gene expression. Cancer-specific fusion oncoproteins, which display unique chromatin localization patterns, often lack classical DNA-binding domains, presenting challenges in identifying mechanisms governing their site-specific chromatin targeting and function. Here we identify a minimal region of the human SS18-SSX fusion oncoprotein (the hallmark driver of synovial sarcoma) that mediates a direct interaction between the mSWI/SNF complex and the nucleosome acidic patch. This binding results in altered mSWI/SNF composition and nucleosome engagement, driving cancer-specific mSWI/SNF complex targeting and gene expression. Furthermore, the C-terminal region of SSX confers preferential affinity to repressed, H2AK119Ub-marked nucleosomes, underlying the selective targeting to polycomb-marked genomic regions and synovial sarcoma-specific dependency on PRC1 function. Together, our results describe a functional interplay between a key nucleosome binding hub and a histone modification that underlies the disease-specific recruitment of a major chromatin remodeling complex.

Unusual carbon-carbon bond formations between allylboronates and acetals or ketals catalyzed by a peculiar indium(I) Lewis acid.

In(I)OTf has been uncovered as an effective Lewis acid catalyst for unprecedented nucleophilic substitution of acetals or ketals with allylboronates. A transmetalative S(N)1 mechanism is proposed in which a single In(I) center acts as a dual catalyst to activate both reagents sequentially. Contrary to the classic gamma-selectivity of allylsilanes (Hosomi-Sakurai reaction), this In(I)-catalyzed borono variant displays distinct alpha-selectivity. Substrate scope and functional group tolerance proved to be excellent.