Mutation of SMARCA4 (BRG1), the ATPase
of BAF (mSWI/SNF) and PBAF complexes, contributes to a range of malignancies and
neurologic disorders. Unfortunately, the effects of SMARCA4 missense mutations
have remained uncertain. Here we show that SMARCA4 cancer missense mutations
target conserved ATPase surfaces and disrupt the mechanochemical cycle of
remodeling. We find that heterozygous expression of mutants alters the open
chromatin landscape at thousands of sites across the genome. Loss of DNA
accessibility does not directly overlap with Polycomb accumulation, but is
enriched in “A compartments” at active enhancers, which lose
H3K27ac but not H3K4me1. Affected positions include hundreds of sites identified
as superenhancers in many tissues. Dominant-negative mutation induced
pro-oncogenic expression changes, including increased expression of
Myc and its target genes. Together, our data suggest that
disruption of enhancer accessibility represents a key source of altered function
in SMARCA4-mutated disorders in a wide variety of tissues.
Stable integration in the host genome renders murine leukemia virus (MLV)-derived vectors attractive tools for gene therapy. Adverse events in otherwise successful clinical trials caused by proto-oncogene activation due to vector integration hamper their application. MLV and MLV-based vectors integrate near strong enhancers, active promoters, and transcription start sites (TSS) through specific interaction of MLV integrase (IN) with the bromodomain and extra-terminal (BET) family of proteins, accounting for insertional mutagenesis. We identified a BET-interaction motif in the C-terminal tail of MLV IN conserved among gammaretroviruses. By deletion of this motif or a single point mutation (INW390A), BET-independent MLV (BinMLV) were engineered. BinMLV vectors carrying INW390A integrate at wild-type efficiency, with an integration profile that no longer correlates with BET chromatin distribution nor with the traditional markers of MLV integration. In particular, BinMLV vector integration associated less with oncogene TSS compared to the MLV vectors currently used in clinical trials. Together, these findings open perspectives to increase the biosafety of gammaretroviral vectors for gene therapy.
Lens epithelium-derived growth factor (LEDGF/p75) is an epigenetic reader and attractive therapeutic target involved in HIV integration and the development of mixed lineage leukaemia (MLL1) fusion-driven leukaemia. Besides HIV integrase and the MLL1-menin complex, LEDGF/p75 interacts with various cellular proteins via its integrase binding domain (IBD). Here we present structural characterization of IBD interactions with transcriptional repressor JPO2 and domesticated transposase PogZ, and show that the PogZ interaction is nearly identical to the interaction of LEDGF/p75 with MLL1. The interaction with the IBD is maintained by an intrinsically disordered IBD-binding motif (IBM) common to all known cellular partners of LEDGF/p75. In addition, based on IBM conservation, we identify and validate IWS1 as a novel LEDGF/p75 interaction partner. Our results also reveal how HIV integrase efficiently displaces cellular binding partners from LEDGF/p75. Finally, the similar binding modes of LEDGF/p75 interaction partners represent a new challenge for the development of selective interaction inhibitors.
Organized by unstructured motifs
The high degree of conservation in protein sequences thought to be unstructured has hinted that these regions may have important biological functions. Although unstructured regions are widely viewed to be crucial for protein signaling, localization, and stability, their roles in many other settings have remained mysterious. Cermakova
et al
. discovered that prominent members of the transcription elongation machinery are linked through a network of interactions involving transcription elongation factor TFIIS N-terminal domains (TNDs) and conserved unstructured sequences called “TND-interacting motifs” (TIMs). The researchers found that mutation of a single TIM in a central organizing protein of this network abolished key protein interactions and induced widespread defects in transcription elongation dynamics. —DJ
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