Analysis of the chromatin landscape and RNA polymerase II binding at SIN3-regulated genes.

The chromatin environment has a significant impact on gene expression. Chromatin structure is highly regulated by histone modifications and RNA polymerase II binding dynamics. The SIN3 histone modifying complex regulates the chromatin environment leading to changes in gene expression. In Drosophila melanogaster, the Sin3A gene is alternatively spliced to produce different protein isoforms, two of which include SIN3 220 and SIN3 187. Both SIN3 isoforms are scaffolding proteins that interact with several other factors to regulate the chromatin landscape. The mechanism through which the SIN3 isoforms regulate chromatin is not well understood. Here, we analyze publicly available data sets to allow us to ask specific questions on how SIN3 isoforms regulate chromatin and gene activity. We determined that genes repressed by the SIN3 isoforms exhibited enrichment in histone H3K4me2, H3K4me3, H3K14ac and H3K27ac near the transcription start site. We observed an increase in the amount of paused RNA polymerase II on the promoter of genes repressed by the isoforms as compared to genes that require SIN3 for maximum activation. Furthermore, we analyzed a subset of genes regulated by SIN3 187 that suggest a mechanism in which SIN3 187 might exhibit hard regulation as well as soft regulation. Data presented here expand our knowledge of how the SIN3 isoforms regulate the chromatin environment and RNA polymerase II binding dynamics.

SIN-3 acts in distinct complexes to regulate the germline transcriptional program in Caenorhabditis elegans.

The transcriptional co-regulator SIN3 influences gene expression through multiple interactions that include histone deacetylases. Haploinsufficiency and mutations in SIN3 are the underlying cause of Witteveen-Kolk syndrome and related intellectual disability and autism syndromes, emphasizing its key role in development. However, little is known about the diversity of its interactions and functions in developmental processes. Here, we show that loss of SIN-3, the single SIN3 homolog in Caenorhabditis elegans, results in maternal-effect sterility associated with de-regulation of the germline transcriptome, including de-silencing of X-linked genes. We identify at least two distinct SIN3 complexes containing specific histone deacetylases and show that they differentially contribute to fertility. Single-cell, single-molecule fluorescence in situ hybridization reveals that in sin-3 mutants the X chromosome becomes re-expressed prematurely and in a stochastic manner in individual germ cells, suggesting a role for SIN-3 in its silencing. Furthermore, we identify histone residues whose acetylation increases in the absence of SIN-3. Together, this work provides a powerful framework for the in vivo study of SIN3 and associated proteins.

SAP30 promotes breast tumor progression by bridging the transcriptional corepressor SIN3 complex and MLL1.

SAP30 is a core subunit of the transcriptional corepressor SIN3 complex, but little is known about its role in gene regulation and human cancer. Here, we show that SAP30 was a nonmutational oncoprotein upregulated in more than 50% of human breast tumors and correlated with unfavorable outcomes in patients with breast cancer. In various breast cancer mouse models, we found that SAP30 promoted tumor growth and metastasis through its interaction with SIN3A/3B. Surprisingly, the canonical gene silencing role was not essential for SAP30's tumor-promoting actions. SAP30 enhanced chromatin accessibility and RNA polymerase II occupancy at promoters in breast cancer cells, acting as a coactivator for genes involved in cell motility, angiogenesis, and lymphangiogenesis, thereby driving tumor progression. Notably, SAP30 formed a homodimer with 1 subunit binding to SIN3A and another subunit recruiting MLL1 through specific Phe186/200 residues within its transactivation domain. MLL1 was required for SAP30-mediated transcriptional coactivation and breast tumor progression. Collectively, our findings reveal that SAP30 represents a transcriptional dependency in breast cancer.

The additional PRC2 subunit and Sin3 histone deacetylase complex are required for the normal distribution of H3K27me3 occupancy and transcriptional silencing in Magnaporthe oryzae.

Development in higher organisms requires proper gene silencing, partially achieved through trimethylation of lysine 27 on histone H3 (H3K27me3). However, how the normal distribution of this modification is established and maintained and how it affects gene expression remains unclear, especially in fungi. Polycomb repressive complex 2 (PRC2) catalyses H3K27me3 to assemble transcriptionally repressed facultative heterochromatin and is crucial in animals, plants, and fungi. Here, we report on the critical role of an additional PRC2 subunit in the normal distribution of H3K27me3 occupancy and the stable maintenance of gene repression in the rice fungal pathogen Magnaporthe oryzae. P55, identified as an additional PRC2 subunit, is physically associated with core subunits of PRC2 and is required for a complete level of H3K27me3 modification. Loss of P55 caused severe global defects in the normal distribution of H3K27me3 and transcriptional reprogramming on the H3K27me3-occupied genes. Furthermore, we found that the Sin3 histone deacetylase complex was required to sustain H3K27me3 occupancy and stably maintain gene repression by directly interacting with P55. Our results revealed a novel mechanism by which P55 and Sin3 participate in the normal distribution of facultative heterochromatic modifications and the stable maintenance of gene repression in eukaryotes.

The Sin3A/MAD1 Complex, through Its PAH2 Domain, Acts as a Second Repressor of Retinoic Acid Receptor Beta Expression in Breast Cancer Cells.

Retinoids are essential in balancing proliferation, differentiation and apoptosis, and they exert their effects through retinoic acid receptors (RARs) and retinoid X receptors (RXRs). RARß is a tumor-suppressor gene silenced by epigenetic mechanisms such as DNA methylation in breast, cervical and non-small cell lung cancers. An increased expression of RARß has been associated with improved breast cancer-specific survival. The PAH2 domain of the scaffold protein SIN3A interacts with the specific Sin3 Interaction Domain (SID) of several transcription factors, such as MAD1, bringing chromatin-modifying proteins such as histone deacetylases, and it targets chromatin for specific modifications. Previously, we have established that blocking the PAH2-mediated Sin3A interaction with SID-containing proteins using SID peptides or small molecule inhibitors (SMI) increased RARß expression and induced retinoic acid metabolism in breast cancer cells, both in in vitro and in vivo models. Here, we report studies designed to understand the mechanistic basis of RARß induction and function. Using human breast cancer cells transfected with MAD1 SID or treated with the MAD SID peptide, we observed a dissociation of MAD1, RARα and RARß from Sin3A in a coimmunoprecipitation assay. This was associated with increased RARα and RARß expression and function by a luciferase assay, which was enhanced by the addition of AM580, a specific RARα agonist; EMSA showed that MAD1 binds to E-Box, similar to MYC, on the RARß promoter, which showed a reduced enrichment of Sin3A and HDAC1 by ChIP and was required for the AM580-enhanced RARß activation in MAD1/SID cells. These data suggest that the Sin3A/HDAC1/2 complex co-operates with the classical repressors in regulating RARß expression. These data suggest that SIN3A/MAD1 acts as a second RARß repressor and may be involved in fine-tuning retinoid sensitivity.

Structural Insight into the Binding of TGIF1 to SIN3A PAH2 Domain through a C-Terminal Amphipathic Helix.

TGIF1 is a transcriptional repressor playing crucial roles in human development and function and is associated with holoprosencephaly and various cancers. TGIF1-directed transcriptional repression of specific genes depends on the recruitment of corepressor SIN3A. However, to date, the exact region of TGIF1 binding to SIN3A was not clear, and the structural basis for the binding was unknown. Here, we demonstrate that TGIF1 utilizes a C-terminal domain (termed as SIN3A-interacting domain, SID) to bind with SIN3A PAH2. The TGIF1 SID adopts a disordered structure at the apo state but forms an amphipathic helix binding into the hydrophobic cleft of SIN3A PAH2 through the nonpolar side at the holo state. Residues F379, L382 and V383 of TGIF1 buried in the hydrophobic core of the complex are critical for the binding. Moreover, homodimerization of TGIF1 through the SID and key residues of F379, L382 and V383 was evidenced, which suggests a dual role of TGIF1 SID and a correlation between dimerization and SIN3A-PAH2 binding. This study provides a structural insight into the binding of TGIF1 with SIN3A, improves the knowledge of the structure-function relationship of TGIF1 and its homologs and will help in recognizing an undiscovered SIN3A-PAH2 binder and developing a peptide inhibitor for cancer treatment.

Analysis of multiple gene co-expression networks to discover interactions favoring CFTR biogenesis and ΔF508-CFTR rescue.

BACKGROUND: We previously reported that expression of a miR-138 mimic or knockdown of SIN3A in primary cultures of cystic fibrosis (CF) airway epithelia increased ΔF508-CFTR mRNA and protein levels, and partially restored CFTR-dependent chloride transport. Global mRNA transcript profiling in ΔF508-CFBE cells treated with miR-138 mimic or SIN3A siRNA identified two genes, SYVN1 and NEDD8, whose inhibition significantly increased ΔF508-CFTR trafficking, maturation, and function. Little is known regarding the dynamic changes in the CFTR gene network during such rescue events. We hypothesized that analysis of condition-specific gene networks from transcriptomic data characterizing ΔF508-CFTR rescue could help identify dynamic gene modules associated with CFTR biogenesis. METHODS: We applied a computational method, termed M-module, to analyze multiple gene networks, each of which exhibited differential activity compared to a baseline condition. In doing so, we identified both unique and shared gene pathways across multiple differential networks. To construct differential networks, gene expression data from CFBE cells were divided into three groups: (1) siRNA inhibition of NEDD8 and SYVN1; (2) miR-138 mimic and SIN3A siRNA; and (3) temperature (27 °C for 24 h, 40 °C for 24 h, and 27 °C for 24 h followed by 40 °C for 24 h). RESULTS: Interrogation of individual networks (e.g., NEDD8/SYVN1 network), combinations of two networks (e.g., NEDD8/SYVN1 + temperature networks), and all three networks yielded sets of 1-modules, 2-modules, and 3-modules, respectively. Gene ontology analysis revealed significant enrichment of dynamic modules in pathways including translation, protein metabolic/catabolic processes, protein complex assembly, and endocytosis. Candidate CFTR effectors identified in the analysis included CHURC1, GZF1, and RPL15, and siRNA-mediated knockdown of these genes partially restored CFTR-dependent transepithelial chloride current to ΔF508-CFBE cells. CONCLUSIONS: The ability of the M-module to identify dynamic modules involved in ΔF508 rescue provides a novel approach for studying CFTR biogenesis and identifying candidate suppressors of ΔF508.

miR-138-5p inhibits proliferation and invasion in kidney renal clear cell carcinoma by targeting SINA3 and regulation of the Notch signaling pathway.

BACKGROUND: The function of miR-138-5p as an oncogenic factor has been reported in certain cancers. This study was performed to analyze the potential involvement of miR-138-5p in kidney renal clear cell carcinoma (KIRC). METHODS: The Cancer Genome Atlas (TCGA) database was used to explain the expression of miR-138-5p in cancer and paired non-cancer tissues of KIRC patients. Subsequently, miR-138-5p expression in KIRC tissues and cell lines, as well as that in normal tissues and normal renal tubular epithelial cell line, was detected. Artificial overexpressing of miR-138-5p was applied to observe its effect on the biological behaviors of KIRC cells. The target mRNA of miR-138-5p, SIN3A, was predicted and validated. Altered expression of miR-138-5p and SIN3A was introduced to confirm their functions in KIRC proliferation and invasion. RESULTS: We showed that miR-138-5p was down-regulated in tumor tissues of KIRC patients comparing to adjacent healthy tissues and linked to dismal prognosis in patients. miR-138-5p could hinder KIRC proliferation and invasion, while artificial overexpression of SIN3A led to reversed trends. SIN3A was a target mRNA of miR-138-5p. miR-138-5p and SIN3A together affect the activation of the Notch signaling pathway. CONCLUSION: This study evidenced that up-regulated miR-138-5p inhibits proliferation and invasion of KIRC cells involving the transcription of SIN3A and the following regulation of the Notch signaling pathway.

Fusion transcripts FYN-TRAF3IP2 and KHDRBS1-LCK hijack T cell receptor signaling in peripheral T-cell lymphoma, not otherwise specified.

Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of non-Hodgkin lymphomas with poor prognosis. Up to 30% of PTCL lack distinctive features and are classified as PTCL, not otherwise specified (PTCL-NOS). To further improve our understanding of the genetic landscape and biology of PTCL-NOS, we perform RNA-sequencing of 18 cases and validate results in an independent cohort of 37 PTCL cases. We identify FYN-TRAF3IP2, KHDRBS1-LCK and SIN3A-FOXO1 as new in-frame fusion transcripts, with FYN-TRAF3IP2 as a recurrent fusion detected in 8 of 55 cases. Using ex vivo and in vivo experiments, we demonstrate that FYN-TRAF3IP2 and KHDRBS1-LCK activate signaling pathways downstream of the T cell receptor (TCR) complex and confer therapeutic vulnerability to clinically available drugs.

Overlapping and non-overlapping roles of the class-I histone deacetylase-1 corepressors LET-418, SIN-3, and SPR-1 in Caenorhabditis elegans embryonic development.

BACKGROUND: Histone deacetylase (HDAC)-1, a Class-I HDAC family member, forms three types of complexes, the nucleosome remodeling deacetylase, Sin3, and CoREST complexes with the specific corepressor components chromodomain-helicase-DNA-binding protein 3 (Mi2/CHD-3), Sin3, and REST corepressor 1 (RCOR1), respectively, in humans. OBJECTIVE: To elucidate the functional relationships among the three transcriptional corepressors during embryogenesis. METHODS: The activities of HDA-1, LET-418, SIN-3, and SPR-1, the homologs of HDAC-1, Mi2, Sin3, and RCOR1 in Caenorhabditis elegans during embryogenesis were investigated through measurement of relative mRNA expression levels and embryonic lethality given either gene knockdown or deletion. Additionally, the terminal phenotypes of each knockdown and mutant embryo were observed using a differential-interference contrast microscope. Finally, the functional relationships among the three corepressors were examined through genetic interactions and transcriptome analyses. RESULTS: Here, we report that each of the corepressors LET-418, SIN-3, and SPR-1 are expressed and have essential roles in C. elegans embryonic development. Our terminal phenotype observations of single mutants further implied that LET-418, SIN-3, and SPR-1 play similar roles in promoting advancement to the middle and late embryonic stages. Combined analysis of genetic interactions and gene ontology of these corepressors indicate a prominent overlapping role among SIN-3, SPR-1, and LET-418 and between SIN-3 and SPR-1. CONCLUSION: Our findings suggest that the class-I HDAC-1 corepressors LET-418, SIN-3, and SPR-1 may cooperatively regulate the expression levels of some genes during C. elegans embryogenesis or may have some similar roles but functioning independently within a specific cell.

Forkhead transcription factor Fkh1: insights into functional regulatory domains crucial for recruitment of Sin3 histone deacetylase complex.

Transcription factors are inextricably linked with histone deacetylases leading to compact chromatin. The Forkhead transcription factor Fkh1 is mainly a negative transcriptional regulator which affects cell cycle control, silencing of mating-type cassettes and induction of pseudohyphal growth in the yeast Saccharomyces cerevisiae. Markedly, Fkh1 impinges chromatin architecture by recruiting large regulatory complexes. Implication of Fkh1 with transcriptional corepressor complexes remains largely unexplored. In this work we show that Fkh1 directly recruits corepressors Sin3 and Tup1 (but not Cyc8), providing evidence for its influence on epigenetic regulation. We also identified the specific domain of Fkh1 mediating Sin3 recruitment and substantiated that amino acids 51-125 of Fkh1 bind PAH2 of Sin3. Importantly, this part of Fkh1 overlaps with its Forkhead-associated domain (FHA). To analyse this domain in more detail, selected amino acids were replaced by alanine, revealing that hydrophobic amino acids L74 and I78 are important for Fkh1-Sin3 binding. In addition, we could prove Fkh1 recruitment to promoters of cell cycle genes CLB2 and SWI5. Notably, Sin3 is also recruited to these promoters but only in the presence of functional Fkh1. Our results disclose that recruitment of Sin3 to Fkh1 requires precisely positioned Fkh1/Sin3 binding sites which provide an extended view on the genetic control of cell cycle genes CLB2 and SWI5 and the mechanism of transcriptional repression by modulation of chromatin architecture at the G2/M transition.

Transcription Repressor Protein ZBTB25 Associates with HDAC1-Sin3a Complex in Mycobacterium tuberculosis-Infected Macrophages, and Its Inhibition Clears Pathogen by Autophagy.

Downregulation of host gene expression is a key strategy employed by intracellular pathogens for their survival in macrophages and subsequent pathogenesis. In a previous study, we have shown that histone deacetylase 1 (HDAC1) levels go up in macrophages infected with Mycobacterium tuberculosis, and it hypoacetylates histone H3 at the promoter of IL-12B gene, leading to its downregulation. We now show that after infection with M. tuberculosis, HDAC1 is phosphorylated, and the levels of phosphorylated HDAC1 (pHDAC1) increase significantly in macrophages. We found that transcriptional repressor protein zinc finger and BTB domain 25 (ZBTB25) and transcriptional corepressor Sin3a associate with the HDAC1 silencing complex, which is recruited to the promoter of IL-12B to downregulate its expression in infected macrophages. Knocking down of ZBTB25 enhanced release of IL-12p40 from infected macrophages. Inhibition of HDAC1 and ZBTB25 promoted colocalization of M. tuberculosis and LC3 (microtubule-associated protein 1A/1B-light chain 3) in autophagosomes. Induction of autophagy resulted in the killing of intracellular M. tuberculosis Enhanced phosphorylation of JAK2 and STAT4 was observed in macrophages upon treatment with HDAC1 and ZBTB inhibitors, and inhibition of JAK2/STAT4 negated the killing of the intracellular pathogen, suggesting their role in the autophagy-mediated killing of intracellular M. tuberculosis In view of the emergence of drug resistance in M. tuberculosis, host-directed therapy is an attractive alternative strategy to combat tuberculosis (TB). HDACs have been proposed to be host targets for TB treatment. Our study indicates that ZBTB25, a functional subunit of the HDAC1/Sin3a repressor complex involved in IL-12B suppression, could be an alternative target for host-directed anti-TB therapy.IMPORTANCE Following infection with M. tuberculosis, levels of HDAC1 go up in macrophages, and it is recruited to the promoter of IL-12B where it hypoacetylates histone H3, leading to the downregulation of the gene. Here, we show that host transcriptional repressor protein ZBTB25 and transcriptional corepressor Sin3a associate with HDAC1 in the silencing complex. Knocking down of ZBTB25 prevented the recruitment of the complex to the promoter and consequently enhanced the gene expression and the release of IL-12p40 from infected macrophages. Pharmacological inhibition of ZBTB25 in infected macrophages resulted in the induction of autophagy and killing of intracellular M. tuberculosis Drug-resistant TB is a serious challenge to TB control programs all over the world which calls for finding alternative therapeutic methods. Host-directed therapy is gaining significant momentum in treating infectious diseases. We propose that ZBTB25 is a potential target for host-directed treatment of TB.

Single-cell analysis of nonhuman primate preimplantation development in comparison to humans and mice.

BACKGROUND: Genetic programs underlying preimplantation development and early lineage segregation are highly conserved across mammals. It has been suggested that nonhuman primates would be better model organisms for human embryogenesis, but a limited number of studies have investigated the monkey preimplantation development. In this study, we collect single cells from cynomolgus monkey preimplantation embryos for transcriptome profiling and compare with single-cell RNA-seq data derived from human and mouse embryos. RESULTS: By weighted gene-coexpression network analysis, we found that cynomolgus gene networks have greater conservation with human embryos including a greater number of conserved hub genes than that of mouse embryos. Consistently, we found that early ICM/TE lineage-segregating genes in monkeys exhibit greater similarity with human when compared to mouse, so are the genes in signaling pathways such as LRP1 and TCF7 involving in WNT pathway. Last, we tested the role of one conserved pre-EGA hub gene, SIN3A, using a morpholino knockdown of maternal RNA transcripts in monkey embryos followed by single-cell RNA-seq. We found that SIN3A knockdown disrupts the gene-silencing program during the embryonic genome activation transition and results in developmental delay of cynomolgus embryos. CONCLUSION: Taken together, our study provided new insight into evolutionarily conserved and divergent transcriptome dynamics during mammalian preimplantation development.

Comprehensive study of 28 individuals with SIN3A-related disorder underscoring the associated mild cognitive and distinctive facial phenotype.

Witteveen-Kolk syndrome (OMIM 613406) is a recently defined neurodevelopmental syndrome caused by heterozygous loss-of-function variants in SIN3A. We define the clinical and neurodevelopmental phenotypes related to SIN3A-haploinsufficiency in 28 unreported patients. Patients with SIN3A variants adversely affecting protein function have mild intellectual disability, growth and feeding difficulties. Involvement of a multidisciplinary team including a geneticist, paediatrician and neurologist should be considered in managing these patients. Patients described here were identified through a combination of clinical evaluation and gene matching strategies (GeneMatcher and Decipher). All patients consented to participate in this study. Mean age of this cohort was 8.2 years (17 males, 11 females). Out of 16 patients ≥ 8 years old assessed, eight (50%) had mild intellectual disability (ID), four had moderate ID (22%), and one had severe ID (6%). Four (25%) did not have any cognitive impairment. Other neurological symptoms such as seizures (4/28) and hypotonia (12/28) were common. Behaviour problems were reported in a minority. In patients ≥2 years, three were diagnosed with Autism Spectrum Disorder (ASD) and four with Attention Deficit Hyperactivity Disorder (ADHD). We report 27 novel variants and one previously reported variant. 24 were truncating variants; three were missense variants and one large in-frame gain including exons 10-12.

Expression of the Neural REST/NRSF-SIN3 Transcriptional Corepressor Complex as a Target for Small-Molecule Inhibitors.

The repressor element 1 (RE1) silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF) modulates the expression of genes with RE1/neuron-restrictive silencing element (RE1/NRSE) sites by recruiting the switch independent 3 (SIN3) factor and the REST corepressor (COREST) to its N and C-terminal repressor domain, respectively. Both, SIN3 and COREST assemble into protein complexes that are composed of multiple subunits including a druggable histone deacetylase (HDAC) enzyme. The SIN3 core complex comprises the eponymous proteins SIN3A or SIN3B, the catalytically active proteins HDAC1 or HDAC2, the histone chaperone retinoblastoma-associated protein 46/retinoblastoma-binding protein 7 (RBAP46/RBBP7) or RBAP48/RBBP4, the SIN3-associated protein 30 (SAP30), and the suppressor of defective silencing 3 (SDS3). Here, we overcome a bottleneck limiting the molecular characterization of the REST/NRSF-SIN3 transcriptional corepressor complex. To this end, SIN3 genes were amplified from the complementary DNA of neural stem/progenitor cells, and expressed in a baculovirus/insect cell expression system. We show that the isolates bind to DNA harboring RE1/NRSE sites and demonstrate that the histone deacetylase activity is blocked by small-molecule inhibitors. Thus, our isolates open up for future biomedical research on this critical transcriptional repressor complex and are envisioned as tool for drug testing.

αα-Hub domains and intrinsically disordered proteins: A decisive combo.

Hub proteins are central nodes in protein-protein interaction networks with critical importance to all living organisms. Recently, a new group of folded hub domains, the αα-hubs, was defined based on a shared αα-hairpin supersecondary structural foundation. The members PAH, RST, TAFH, NCBD, and HHD are found in large proteins such as Sin3, RCD1, TAF4, CBP, and harmonin, which organize disordered transcriptional regulators and membrane scaffolds in interactomes of importance to human diseases and plant quality. In this review, studies of structures, functions, and complexes across the αα-hubs are described and compared to provide a unified description of the group. This analysis expands the associated molecular concepts of "one domain-one binding site", motif-based ligand binding, and coupled folding and binding of intrinsically disordered ligands to additional concepts of importance to signal fidelity. These include context, motif reversibility, multivalency, complex heterogeneity, synergistic αα-hub:ligand folding, accessory binding sites, and supramodules. We propose that these multifaceted protein-protein interaction properties are made possible by the characteristics of the αα-hub fold, including supersite properties, dynamics, variable topologies, accessory helices, and malleability and abetted by adaptability of the disordered ligands. Critically, these features provide additional filters for specificity. With the presentations of new concepts, this review opens for new research questions addressing properties across the group, which are driven from concepts discovered in studies of the individual members. Combined, the members of the αα-hubs are ideal models for deconvoluting signal fidelity maintained by folded hubs and their interactions with intrinsically disordered ligands.

HDAC1-Smad3-mSin3A complex is required for Smad3-induced transcriptional inhibition of hepatocyte growth factor receptor in human lung cancers.

c-Met hyperactivity has been observed in numerous neoplasms. Several researchers have shown that the abnormal activation of c-Met is mainly caused by transcriptional activation. However, the molecular mechanism behind this transcriptional regulation is poorly understood. Here, we suggest that Smad3 negatively regulates the expression and activation of c-Met via a transcriptional mechanism. We explore the molecular mechanisms that underlie Smad3-induced c-Met transcription inhibition. We found in contrast to the high expression of c-Met, Smad3 showed low protein and mRNA levels. Smad3 and c-Met expressions were inconsistent between lung cancer tissues and cell lines. We also found that Smad3 overexpression suppresses whereas Smad3 knockdown significantly promotes Epithelial-Mesenchymal Transition and production of the angiogenic factors VEGF, CTGF and COX-2 through the ERK1/2 pathway. In addition, Smad3 overexpression decreases whereas Smad3 knockdown significantly increases protein and mRNA levels of invasion-related ß-catenin and FAK through the PI3K/Akt pathway. Furthermore, using the chromatin immunoprecipitation analysis method, we demonstrate that a transcriptional regulatory complex consisting of HDAC1, Smad3 and mSin3A binds to the promoter of the c-Met gene. By either silencing endogenous mSin3A expression with siRNA or by pretreating cells with a specific HDAC1 inhibitor (MS-275), Smad3-induced transcriptional suppression of c-Met could be effectively attenuated. These results demonstrate that Smad3-induced inhibition of c-Met transcription depends on of a functional transcriptional regulatory complex that includes Smad3, mSin3A and HDAC1 at the c-Met promoter. Collectively, our findings reveal a new regulatory mechanism of c-Met signaling, and suggest a potential molecular target for the development of anticancer drugs.

Witteveen-Kolk syndrome: The first patient from Turkey.

Witteveen-Kolk syndrome is a rare genetic disorder characterized by intellectual disability, developmental delay and dysmorphic facial features including a long face with prominent forehead, depressed nasal bridge, long-smooth philtrum and malformed ears. Skeletal abnormalities, microcephaly and malformation of the brain are other findings. This syndrome is caused by mutations in the SIN3A gene or microdeletions encompassing this gene. The protein encoded by SIN3A gene plays a regulatory role in the control of various developmental processes, especially cortical expansion and maturation. To date, 17 patients have been reported in the medical literature. In this article, we reported a patient with Witteveen-Kolk syndrome who had a retrognathia as an unusually finding. To the best of our knowledge, this is the first patient of Witteveen-Kolk syndrome reported from Turkey.

Electroacupuncture reduces scopolamine-induced amnesia via mediating the miR-210/SIN3A and miR-183/SIN3A signaling pathway.

BACKGROUND: The expression of SIN3A is closely correlated with electroacupuncture (EA) treatment efficacy of scopolamine-induced amnesia (SIA), but its underlying mechanisms remain to be further explored. METHODS: Quantitative real-time PCR was performed to analyze the expression of candidate microRNAs (miRNAs) and SIN3A mRNA in a rat model of SIA. Western blot was carried out to evaluate the differential expression of SIN3A proteins under different circumstances. Luciferase assay was used to explore the inhibitory role of certain miRNAs in SIN3A expression. A novel object recognition (NOR) test was performed to assess the memory function of SIA rats undergoing EA treatment. Immunohistochemistry was carried out to evaluate the expression of SIN3A in the hippocampus of SIA rats. RESULTS: Rno-miR-183-5p, rno-miR-34c-3p and rno-miR-210-3p were significantly up-regulated in SIA rats treated with EA. In addition, rno-miR-183-5p and rno-miR-210-3p exerted an inhibitory effect on SIN3A expression. EA treatment of SIA rats effectively restored the dysregulated expression of rno-miR-183-5p, rno-miR-210-3p and SIN3A. EA treatment also promoted the inhibited expression of neuronal IEGs including Arc, Egr1, Homer1 and Narp in the hippocampus of SIA rats. Accordingly, the NOR test also confirmed the effect of EA treatment on the improvement of memory in SIA rats. CONCLUSION: In summary, the findings of this study demonstrated that scopolamine-induced amnesia was associated with downregulated expression of miR-210/miR-183 and upregulated expression of SIN3A. Furthermore, treatment with EA alleviated scopolamine-induced amnesia in rats and was associated with upregulated expression of miR-210/miR-183 and downregulated expression of SIN3A.

Haploinsufficiency of RREB1 causes a Noonan-like RASopathy via epigenetic reprogramming of RAS-MAPK pathway genes.

RAS-MAPK signaling mediates processes critical to normal development including cell proliferation, survival, and differentiation. Germline mutation of RAS-MAPK genes lead to the Noonan-spectrum of syndromes. Here, we present a patient affected by a 6p-interstitial microdeletion with unknown underlying molecular etiology. Examination of 6p-interstitial microdeletion cases reveals shared clinical features consistent with Noonan-spectrum disorders including short stature, facial dysmorphia and cardiovascular abnormalities. We find the RAS-responsive element binding protein-1 (RREB1) is the common deleted gene in multiple 6p-interstitial microdeletion cases. Rreb1 hemizygous mice display orbital hypertelorism and cardiac hypertrophy phenocopying the human syndrome. Rreb1 haploinsufficiency leads to sensitization of MAPK signaling. Rreb1 recruits Sin3a and Kdm1a to control H3K4 methylation at MAPK pathway gene promoters. Haploinsufficiency of SIN3A and mutations in KDM1A cause syndromes similar to RREB1 haploinsufficiency suggesting genetic perturbation of the RREB1-SIN3A-KDM1A complex represents a new category of RASopathy-like syndromes arising through epigenetic reprogramming of MAPK pathway genes.