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Key messageSCL3 inhibits transcriptional activity of IDD-DELLA complex by acting as a co-repressor and repression activity is enhanced in the presence of GAF1 in a TOPLESS-independent manner. GRAS [GIBBERELLIN-INSENSITIVE (GAI), REPRESSOR OF ga1-3 (RGA) and SCARECROW (SCR)] proteins are a family of plant-specific transcriptional regulators that play diverse roles in development and signaling. GRAS family DELLA proteins act as growth repressors by inhibiting gibberellin (GA) signaling in response to developmental and environmental cues. DELLAs also act as co-activators of transcription factor GAI-ASSOCIATED FACTOR1 (GAF1)/INDETERMINATE DOMAIN2 (IDD2), the GAF1-DELLA complex activating transcription of GAF1 target genes. GAF1 also interacts with TOPLESS (TPL), a transcriptional co-repressor, in the absence of DELLA, the GAF1-TPL complex repressing transcription of the target genes. SCARECROW-LIKE3 (SCL3), another member of the GRAS family, is thought to inhibit transcriptional activity of the IDD-DELLA complex through competitive interaction with IDD. Here, we also revealed that SCL3 inhibits transcriptional activation by the GAF1-DELLA complex via repression activity rather than via competitive inhibition of the GAF1-DELLA interaction. Moreover, the repression activity of SCL3 was enhanced by GAF1 in a TPL-independent manner. While the GRAS domain of DELLA has transcriptional activation activity, that of SCL3 has repression activity. SCL3 also inhibited transcriptional activity of GAF1-RGA fusion proteins. Results from the co-immunoprecipitation assays and the yeast three-hybrid assay suggested the possibility that SCL3 forms a ternary complex with GAF1 and DELLA. These findings provide important information on DELLA-regulated GA signaling and new insight into the transcriptional repression mechanism.

Bone remodeling consists of resorption by osteoclasts followed by formation by osteoblasts, and osteoclasts are a source of bone formation-stimulating factors. Here we utilize osteoclast ablation by denosumab (DMAb) and RNA-sequencing of bone biopsies from postmenopausal women to identify osteoclast-secreted factors suppressed by DMAb. Based on these analyses, LIF, CREG2, CST3, CCBE1 , and DPP4 are likely osteoclast-derived coupling factors in humans. Given the role of Dipeptidyl Peptidase-4 (DPP4) in glucose homeostasis, we further demonstrate that DMAb-treated participants have a significant reduction in circulating DPP4 and increase in Glucagon-like peptide (GLP)-1 levels as compared to the placebo-treated group, and also that type 2 diabetic patients treated with DMAb show significant reductions in HbA1c as compared to patients treated either with bisphosphonates or calcium and vitamin D. Thus, our results identify several coupling factors in humans and uncover osteoclast-derived DPP4 as a potential link between bone remodeling and energy metabolism. Anti-resorptive bone therapies also inhibit bone formation, as osteoclasts secrete factors that stimulate bone formation by osteoblasts. Here, the authors identify osteoclast-secreted factors that couple bone resorption to bone formation in healthy subjects, and show that osteoclast-derived DPP4 may be a factor coupling bone resorption to energy metabolism.

The deacetylase enzyme HDAC8 is identified as a crucial regulator of cohesin in humans, and loss-of-function mutations in the HDAC8 gene are found in patients with Cornelia de Lange syndrome. HDAC defects in Cornelia de Lange syndrome The cohesin complex is important for sister-chromatid cohesion and chromosome segregation, as well as for other chromosomal processes such as gene expression and DNA repair. Cornelia de Lange syndrome (CdLS) is a human developmental disorder associated with significant cognitive deficits and structural birth defects. It is caused by mutations in genes that encode subunits of the cohesin complex or the cohesin regulator NIPL. Here, a deacetylase enzyme, HDAC8, is shown to be a critical regulator of cohesin in human cells, and loss-of-function HDAC8 mutations are found in six patients with CdLS from different families. Cornelia de Lange syndrome (CdLS) is a dominantly inherited congenital malformation disorder, caused by mutations in the cohesin-loading protein NIPBL 1 , 2 for nearly 60% of individuals with classical CdLS 3 , 4 , 5 , and by mutations in the core cohesin components SMC1A (∼5%) and SMC3 (<1%) for a smaller fraction of probands 6 , 7 . In humans, the multisubunit complex cohesin is made up of SMC1, SMC3, RAD21 and a STAG protein. These form a ring structure that is proposed to encircle sister chromatids to mediate sister chromatid cohesion 8 and also has key roles in gene regulation 9 . SMC3 is acetylated during S-phase to establish cohesiveness of chromatin-loaded cohesin 10 , 11 , 12 , 13 , and in yeast, the class I histone deacetylase Hos1 deacetylates SMC3 during anaphase 14 , 15 , 16 . Here we identify HDAC8 as the vertebrate SMC3 deacetylase, as well as loss-of-function HDAC8 mutations in six CdLS probands. Loss of HDAC8 activity results in increased SMC3 acetylation and inefficient dissolution of the ‘used’ cohesin complex released from chromatin in both prophase and anaphase. SMC3 with retained acetylation is loaded onto chromatin, and chromatin immunoprecipitation sequencing analysis demonstrates decreased occupancy of cohesin localization sites that results in a consistent pattern of altered transcription seen in CdLS cell lines with either NIPBL or HDAC8 mutations.

Despite preventive vaccines for oncogenic human papillomaviruses (HPVs), cervical intraepithelial neoplasia (CIN) is common, and current treatments are ablative and can lead to long-term reproductive morbidity. We assessed whether VGX-3100, synthetic plasmids targeting HPV-16 and HPV-18 E6 and E7 proteins, delivered by electroporation, would cause histopathological regression in women with CIN2/3. Efficacy, safety, and immunogenicity of VGX-3100 were assessed in CIN2/3 associated with HPV-16 and HPV-18, in a randomised, double-blind, placebo-controlled phase 2b study. Patients from 36 academic and private gynaecology practices in seven countries were randomised (3:1) to receive 6 mg VGX-3100 or placebo (1 mL), given intramuscularly at 0, 4, and 12 weeks. Randomisation was stratified by age (<25 vs ≥25 years) and CIN2 versus CIN3 by computer-generated allocation sequence (block size 4). Funder and site personnel, participants, and pathologists were masked to treatment. The primary efficacy endpoint was regression to CIN1 or normal pathology 36 weeks after the first dose. Per-protocol and modified intention-to-treat analyses were based on patients receiving three doses without protocol violations, and on patients receiving at least one dose, respectively. The safety population included all patients who received at least one dose. The trial is registered at ClinicalTrials.gov (number NCT01304524) and EudraCT (number 2012-001334-33). Between Oct 19, 2011, and July 30, 2013, 167 patients received either VGX-3100 (n=125) or placebo (n=42). In the per-protocol analysis 53 (49·5%) of 107 VGX-3100 recipients and 11 (30·6%) of 36 placebo recipients had histopathological regression (percentage point difference 19·0 [95% CI 1·4–36·6]; p=0·034). In the modified intention-to-treat analysis 55 (48·2%) of 114 VGX-3100 recipients and 12 (30·0%) of 40 placebo recipients had histopathological regression (percentage point difference 18·2 [95% CI 1·3–34·4]; p=0·034). Injection-site reactions occurred in most patients, but only erythema was significantly more common in the VGX-3100 group (98/125, 78·4%) than in the placebo group (24/42, 57·1%; percentage point difference 21·3 [95% CI 5·3–37·8]; p=0·007). VGX-3100 is the first therapeutic vaccine to show efficacy against CIN2/3 associated with HPV-16 and HPV-18. VGX-3100 could present a non-surgical therapeutic option for CIN2/3, changing the treatment outlook for this common disease. Inovio Pharmaceuticals.

Dysregulation of the transcriptional repressor element-1 silencing transcription factor (REST)/neuron-restrictive silencer factor is important in a broad range of diseases, including cancer, diabetes, and heart disease. The role of REST-dependent epigenetic modifications in neurodegeneration is less clear. Here, we show that neuronal insults trigger activation of REST and CoREST in a clinically relevant model of ischemic stroke and that REST binds a subset of \"transcriptionally responsive\" genes (gria2, grin1, chrnb2, nefh, nfκb2, trpv1, chrm4, and syt6), of which the AMPA receptor subunit GluA2 is a top hit. Genes with enriched REST exhibited decreased mRNA and protein. We further show that REST assembles with CoREST, mSin3A, histone deacetylases 1 and 2, histone methyl-transferase G9a, and methyl CpG binding protein 2 at the promoters of target genes, where it orchestrates epigenetic remodeling and gene silencing. RNAi-mediated depletion of REST or administration of dominant-negative REST delivered directly into the hippocampus in vivo prevents epigenetic modifications, restores gene expression, and rescues hippocampal neurons. These findings document a causal role for REST-dependent epigenetic remodeling in the neurodegeneration associated with ischemic stroke and identify unique therapeutic targets for the amelioration of hippocampal injury and cognitive deficits.

In this study, the authors show that MeCP2 interacts with the NCoR/SMRT co-repressor complex and that a discrete cluster of Rett syndrome–causing mutations in the C-terminal domain of MeCP2 disrupts this interaction, impairing transcriptional repression. Knock-in mice expressing one of these MeCP2 missense mutations exhibit severe motor phenotypes. Rett syndrome (RTT) is a severe neurological disorder that is caused by mutations in the MECP2 gene. Many missense mutations causing RTT are clustered in the DNA-binding domain of MeCP2, suggesting that association with chromatin is critical for its function. We identified a second mutational cluster in a previously uncharacterized region of MeCP2. We found that RTT mutations in this region abolished the interaction between MeCP2 and the NCoR/SMRT co-repressor complexes. Mice bearing a common missense RTT mutation in this domain exhibited severe RTT-like phenotypes. Our data are compatible with the hypothesis that brain dysfunction in RTT is caused by a loss of the MeCP2 'bridge' between the NCoR/SMRT co-repressors and chromatin.

Rett syndrome (RTT) is an X-linked neurological disorder caused by mutations in the methyl-CpG–binding protein 2 (MeCP2) gene. The majority of RTT missense mutations disrupt the interaction of the MeCP2 with DNA or the nuclear receptor corepressor (NCoR)/silencing mediator of retinoic acid and thyroid receptors (SMRT) corepressor complex. Here, we show that the “NCoR/SMRT interaction domain” (NID) of MeCP2 directly contacts transducin beta-like 1 (TBL1) and TBL1 related (TBLR1), two paralogs that are core components of NCoR/SMRT. We determine the cocrystal structure of the MeCP2 NID in complex with the WD40 domain of TBLR1 and confirm by in vitro and ex vivo assays that mutation of interacting residues of TBLR1 and TBL1 disrupts binding to MeCP2. Strikingly, the four MeCP2-NID residues mutated in RTT are those residues that make the most extensive contacts with TBLR1. Moreover, missense mutations in the gene for TBLR1 that are associated with intellectual disability also prevent MeCP2 binding. Our study therefore reveals the molecular basis of an interaction that is crucial for optimal brain function.

• Anthocyanin and proanthocyanidin (PA) accumulation is regulated by both myeloblastosis (MYB) activators and repressors, but little information is available on hierarchical interactions between the positive and negative regulators. Here, we report on a R2R3-MYB repressor in peach, designated PpMYB18, which acts as a negative regulator of anthocyanin and PA accumulation. • PpMYB18 can be activated by both anthocyanin- and PA-related MYB activators, and is expressed both at fruit ripening and juvenile stages when anthocyanins or PAs, respectively, are being synthesized. • The PpMYB18 protein competes with MYB activators for binding to basic Helix Loop Helixes (bHLHs), which develops a fine-tuning regulatory loop to balance PA and anthocyanin accumulation. In addition, the bHLH binding motif in the R3 domain and the C1 and C2 repression motifs in the C-terminus of PpMYB18 both confer repressive activity of PpMYB18. • Our study also demonstrates a modifying negative feedback loop, which prevents cells from excess accumulation of anthocyanin and PAs, and serves as a model for balancing secondary metabolite accumulation at the transcriptional level.

• As excess iron (Fe) is toxic, uptake of this essential micronutrient must be tightly controlled. Previous studies have shown that Oryza sativa (rice) POSITIVE REGULATOR OF IRON HOMEOSTASIS1 (OsPRI1) acts upstream of the iron-related transcription factor 2 (OsIRO2) and OsIRO3 to positively regulate root-to-shoot Fe translocation. However, as expression of OsPRI1 is constitutive it is unclear how the Fe-deficiency response is turned off to prevent toxicity when Fe is sufficient. • The bHLH transcription factor OsbHLH061 interacts with OsPRI1, and this study used molecular, genetics, biochemical and physiological approaches to functionally characterise OsbHLH061 and how it affects Fe homeostasis. • OsbHLH061 knockout or overexpression lines increase or decrease Fe accumulation in shoots respectively. Mechanistically, OsbHLH061 expression is upregulated by high Fe, and physically interacts with OsPRI1, the OsbHLH061–OsPRI1 complex recruits TOPLESS/TOPLESS-RELATED (OsTPL/TPR) co-repressors to repress OsIRO2 and OsIRO3 expression. The OsbHLH061 ethylene-responsive element-binding factor-associated amphiphilic repression (EAR) motif is required for this transcriptional repression activity. • These results define a functional OsTPL/TPR–OsbHLH061–OsPRI1–OsIRO2/3 module that negatively controls long-distance transport of Fe in plants for adaptation to changing Fe environments and maintain Fe homeostasis in rice.

Here we report corin, a synthetic hybrid agent derived from the class I HDAC inhibitor (entinostat) and an LSD1 inhibitor (tranylcypromine analog). Enzymologic analysis reveals that corin potently targets the CoREST complex and shows more sustained inhibition of CoREST complex HDAC activity compared with entinostat. Cell-based experiments demonstrate that corin exhibits a superior anti-proliferative profile against several melanoma lines and cutaneous squamous cell carcinoma lines compared to its parent monofunctional inhibitors but is less toxic to melanocytes and keratinocytes. CoREST knockdown, gene expression, and ChIP studies suggest that corin’s favorable pharmacologic effects may rely on an intact CoREST complex. Corin was also effective in slowing tumor growth in a melanoma mouse xenograft model. These studies highlight the promise of a new class of two-pronged hybrid agents that may show preferential targeting of particular epigenetic regulatory complexes and offer unique therapeutic opportunities. Alteration of the epigenetic landscape has been implicated in several disease processes, where targeting histone modifiers may have therapeutic applications. Here the authors report a bifunctional small molecule inhibitor that simultaneously targets the deacetylase (HDAC1) and demethylase (LSD1) activities of the CoREST complex.