ATP-dependent chromatin remodeling by the CHD family of proteins plays an important role in the regulation of gene transcription. Here we report that full-length CHD8 interacts directly with ␤-catenin and that CHD8 is also recruited specifically to the promoter regions of several ␤-catenin-responsive genes. Our results indicate that CHD8 negatively regulates ␤-catenin-targeted gene expression, since short hairpin RNA against CHD8 results in the activation of several ␤-catenin target genes. This regulation is also conserved through evolution; RNA interference against kismet, the apparent Drosophila ortholog of CHD8, results in a similar activation of ␤-catenin target genes. We also report the first demonstration of chromatin remodeling activity for a member of the CHD6-9 family of proteins, suggesting that CHD8 functions in transcription through the ATP-dependent modulation of chromatin structure.The alteration of chromatin structure provides a key regulatory step for all processes that act upon DNA (41). The factors that regulate this structure, commonly referred to as chromatin remodeling enzymes, can be grouped into two broad categories: complexes that alter chromatin structure via the covalent modification of histones (25,42,83) and complexes that use the energy of ATP hydrolysis to alter the structure or position of the nucleosome (6,47,57,67).ATP-dependent chromatin remodeling enzymes modulate the contacts between histones and DNA. In vitro, these enzymes catalyze structural changes that allow factors to access nucleosomal DNA, reposition nucleosomes on a template, transfer histone octamers to donor DNA, and replace histones with histone variants (27,43,80). In vivo, these activities are crucial for transcription, replication, repair, and recombination of the eukaryotic genome (2,21,59,65). These remodeling enzymes can be divided into numerous families based on domain architecture. One such family is the CHD (chromodomain, helicase, DNA binding) group of proteins, which are critical regulators of chromatin structure (23,26,29,49). These enzymes are characterized by tandem chromodomains N-terminal to their catalytic Snf2 helicase domain.The CHD family can be further subdivided into three subfamilies: CHD1-2, CHD3-5, and CHD6-9. While the first two subfamilies have been extensively studied, very little is known about the CHD6-9 family (29, 49). Previous studies have indicated that CHD8 may regulate the Wnt signaling pathway, since an N-terminal fragment of CHD8 was previously identified as a protein in Rattus norvegicus that binds ␤-catenin both in vivo and in vitro (61). This N-terminal fragment, termed Duplin, contains only the chromodomains and lacks the Snf2 helicase domain and C-terminal sequences. Overexpression of this N-terminal fragment results in inhibition of Tcf4-dependent transcription, and studies of Xenopus embryos demonstrated that this fragment inhibited axis formation and ␤-catenin-mediated axis duplication (61).The "canonical" Wnt signaling pathway functions by controlling the soluble pool ...

Mutations in proteins like FUS which cause Amyotrophic Lateral Sclerosis (ALS) result in the aberrant formation of stress granules while ALS-linked mutations in other proteins impede elimination of stress granules. Repeat expansions in C9ORF72, the major cause of ALS, reduce C9ORF72 levels but how this impacts stress granules is uncertain. Here, we demonstrate that C9ORF72 associates with the autophagy receptor p62 and controls elimination of stress granules by autophagy. This requires p62 to associate via the Tudor protein SMN with proteins, including FUS, that are symmetrically methylated on arginines. Mice lacking p62 accumulate arginine-methylated proteins and alterations in FUS-dependent splicing. Patients with C9ORF72 repeat expansions accumulate symmetric arginine dimethylated proteins which co-localize with p62. This suggests that C9ORF72 initiates a cascade of ALS-linked proteins (C9ORF72, p62, SMN, FUS) to recognize stress granules for degradation by autophagy and hallmarks of a defect in this process are observable in ALS patients.

Histone H3 Lys-4 methylation is predominantly catalyzed by a family of methyltransferases whose enzymatic activity depends on their interaction with a three-subunit complex composed of WDR5, RbBP5, and Ash2L. Here, we report that a segment of 50 residues of RbBP5 bridges the Ash2L C-terminal domain to WDR5. The crystal structure of WDR5 in ternary complex with RbBP5 and MLL1 reveals that both proteins binds peptide-binding clefts located on opposite sides of WDR5's β-propeller domain. RbBP5 engages in several hydrogen bonds and van der Waals contacts within a V-shaped cleft formed by the junction of two blades on WDR5. Mutational analyses of both the WDR5 V-shaped cleft and RbBP5 residues reveal that the interactions between RbBP5 and WDR5 are important for the stimulation of MLL1 methyltransferase activity. Overall, this study provides the structural basis underlying the formation of the WDR5-RbBP5 subcomplex and further highlight the crucial role of WDR5 in scaffolding the MLL1 core complex.

T-cell acute lymphoblastic leukemia (T-ALL) is a heterogeneous group of hematological tumors composed of distinct subtypes that vary in their genetic abnormalities, gene expression signatures, and prognoses. However, it remains unclear whether T-ALL subtypes differ at the functional level, and, as such, T-ALL treatments are uniformly applied across subtypes, leading to variable responses between patients. Here we reveal the existence of a subtypespecific epigenetic vulnerability in T-ALL by which a particular subgroup of T-ALL characterized by expression of the oncogenic transcription factor TAL1 is uniquely sensitive to variations in the dosage and activity of the histone 3 Lys27 (H3K27) demethylase UTX/KDM6A. Specifically, we identify UTX as a coactivator of TAL1 and show that it acts as a major regulator of the TAL1 leukemic gene expression program. Furthermore, we demonstrate that UTX, previously described as a tumor suppressor in T-ALL, is in fact a pro-oncogenic cofactor essential for leukemia maintenance in TAL1-positive (but not TAL1-negative) T-ALL. Exploiting this subtype-specific epigenetic vulnerability, we propose a novel therapeutic approach based on UTX inhibition through in vivo administration of an H3K27 demethylase inhibitor that efficiently kills TAL1-positive primary human leukemia. These findings provide the first opportunity to develop personalized epigenetic therapy for T-ALL patients.

The SMYD (SET and MYND domain) family of lysine methyltransferases (KMTs) plays pivotal roles in various cellular processes, including gene expression regulation and DNA damage response. Initially identified as genuine histone methyltransferases, specific members of this family have recently been shown to methylate non-histone proteins such as p53, VEGFR, and the retinoblastoma tumor suppressor (pRb). To gain further functional insights into this family of KMTs, we generated the protein interaction network for three different human SMYD proteins (SMYD2, SMYD3, and SMYD5). Characterization of each SMYD protein network revealed that they associate with both shared and unique sets of proteins. Among those, we found that HSP90 and several of its co-chaperones interact specifically with the tetratrico peptide repeat (TPR)-containing SMYD2 and SMYD3. Moreover, using proteomic and biochemical techniques, we provide evidence that SMYD2 methylates K209 and K615 on HSP90 nucleotide-binding and dimerization domains, respectively. In addition, we found that each methylation site displays unique reactivity in regard to the presence of HSP90 co-chaperones, pH, and demethylation by the lysine amine oxidase LSD1, suggesting that alternative mechanisms control HSP90 methylation by SMYD2. Altogether, this study highlights the ability of SMYD proteins to form unique protein complexes that may underlie their various biological functions and the SMYD2-mediated methylation of the key molecular chaperone HSP90.