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Lung cancer is the leading cause of cancer death worldwide. 85 % of lung cancers are categorized by their histological types as a non-small cell lung cancer (NSCLC) subtype. While the MED23 subunit of the mediator complex has been implicated in lung cancer development, the precise underlying mechanism remains unclear. Our research indicates that elevated MED23 expression is linked to reduced overall survival rates in NSCLC. Depletion of MED23 triggers premature senescence in NSCLC cells. Furthermore, through co-IP and mass spectrometry analyses, we have identified BCLAF1 as a binding partner of MED23, with subsequent confirmation via PLA assays. Subsequently, NUPR1, a transcriptional cofactor known to induce premature senescence in lung cancer cells by disrupting autophagic processes, was validated as a downstream target of the MED23/BCLAF1 complex through RNA-seq and ChIP assays. Thus, the interaction between MED23 and BCLAF1 regulates NUPR1 expression, impacting autophagic flux and leading to premature senescence in NSCLC cells., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Activation domains (ADs) within transcription factors (TFs) induce gene expression by recruiting coactivators such as the Mediator complex. Coactivators lack DNA binding domains (DBDs) and are assumed to passively follow their recruiting TFs. This is supported by direct AD-coactivator interactions seen in vitro but has not yet been tested in living cells. To examine that, we targeted two Med15-recruiting ADs to a range of budding yeast promoters through fusion with different DBDs. The DBD-AD fusions localized to hundreds of genomic sites but recruited Med15 and induced transcription in only a subset of bound promoters, characterized by a fuzzy-nucleosome architecture. Direct DBD-Med15 fusions shifted DBD localization towards fuzzy-nucleosome promoters, including promoters devoid of the endogenous Mediator. We propose that Med15, and perhaps other coactivators, possess inherent promoter preference and thus actively contribute to the selection of TF-induced genes., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Aim: Transcriptional regulation of gene expression plays a crucial role in orchestrating complex morphogenetic and molecular events during heart development and function. Mediator complex is an essential multi-subunit protein complex that governs gene expression in eukaryotic cells. Although Mediator subunits (MEDs) work integrally in the complex, individual MED component displays specialized functions. MED27, categorized as an Upper Tail subunit, possesses an as-yet-uncharacterized function. In this study, we aimed to investigate the physiological role of MED27 in cardiomyocytes., Materials and Methods: we generated a Med27 floxed mouse line, which was further used to generate constitutive (cKO) and inducible (icKO) cardiomyocyte-specific Med27 knockout mouse models. Morphological, histological analysis and cardiac physiological studies were performed in Med27 cKO and icKO mutants. Transcriptional profiles were determined by RNA sequencing (RNAseq) analysis., Key Fundings: Ablation of MED27 in developing mouse cardiomyocytes results in embryonic lethality, while its deletion in adult cardiomyocytes leads to heart failure and mortality. Similar to the ablation of another Upper Tail subunit, MED30 in cardiomyocytes, deletion of MED27 leads to decreased protein levels of most MEDs in cardiomyocytes. Interestingly, overexpression of MED30 fails to restore the protein levels of Mediator subunits in MED27-deficient cardiomyocytes, demonstrating that the role of MED27 in maintaining the integrity and stability of the Mediator complex is independent of MED30., Significance: Our results revealed an essential role of MED27 in cardiac development and function by maintaining the stability of the Mediator core., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: CHRDL1 belongs to a novel class of mRNA molecules. Nonetheless, the specific biological functions and underlying mechanisms of CHRDL1 in oral squamous cell carcinoma (OSCC) remain largely unexplored., Methods: RT-qPCR and immunohistochemical staining were employed to assess the mRNA and protein expression levels of the MED29 gene in clinical samples of OSCC. Additionally, RT-qPCR and Western Blot analyses were conducted to investigate the mRNA and protein expression levels of the MED29 gene specifically in OSCC. The impact of MED29 on epithelial-mesenchymal transition (EMT), invasion, and migration of OSCC was evaluated through scratch assay, transwell assay, and immunofluorescence staining. Furthermore, wound healing assay and Transwell assay were utilized to examine whether CHRDL1 influences the malignant behavior of OSCC by modulating MED29 in vitro. The regulatory role of CHRDL1 on MED29 was further elucidated in vivo through a tail vein lung metastasis model in nude mice., Results: MED29 expression was elevated in tumor tissues of OSCC patients compared with adjacent cancer tissues. Moreover, in CAL27 and SCC25 cell lines, MED29 was upregulated and associated with increased cell migration and invasion abilities. Overexpression of MED29 facilitated EMT in OSCC cell lines, whereas knockdown of MED29 impeded EMT, resulting in diminished cell migration and invasion capacities. CHRDL1 exerted inhibitory effects on the expression of MED29, thereby suppressing EMT progression and consequently restraining the invasion and migration of OSCC cells. Furthermore, CHRDL1 mediated the inhibition of migration of OSCC cell lines to the OSCC through its regulation of MED29., Conclusions: MED29 facilitated the epithelial-mesenchymal transition process in OSCC, thereby promoting migration and invasion. On the other hand, CHRDL1 exerted inhibitory effects on the invasion and metastasis of OSCC by suppressing MED29 through the inhibition of the MAPK signaling pathway., (© 2024. The Author(s).)

Pathogenic variants in the MED13L gene are associated with the autosomal dominant MED13L syndrome, which is characterised by global developmental delay and cardiac malformations. We investigated two heterozygous MED13L variants located at the canonical donor splice site motif of exon 7: c.1009+1G>C and c.1009+5G>C. We report that in silico predictions suggested two possible outcomes: exon 7 skipping, resulting in loss of the phosphodegron motif essential for MED13L regulation, or activation of a cryptic donor site in intron 7, leading to intron retention. RNA analysis confirmed that both variants affected the exon 7 splice donor site, resulting in the retention of 73 bp of intron 7. This retention caused a frameshift and premature translation termination, consistent with haploinsufficiency. Our results highlight the importance of combining predictive and experimental approaches to understand the functional impact of splice site variants. These insights into the molecular consequences of MED13L variants provide a deeper understanding of the genetic basis of MED13L syndrome., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

The eukaryotic transcriptional Mediator comprises a large core (cMED) and a dissociable CDK8 kinase module (CKM). cMED recruits RNA polymerase II (RNA Pol II) and promotes pre-initiation complex formation in a manner repressed by the CKM through mechanisms presently unknown. Herein, we report cryoelectron microscopy structures of the complete human Mediator and its CKM. The CKM binds to multiple regions on cMED through both MED12 and MED13, including a large intrinsically disordered region (IDR) in the latter. MED12 and MED13 together anchor the CKM to the cMED hook, positioning CDK8 downstream and proximal to the transcription start site. Notably, the MED13 IDR obstructs the recruitment of RNA Pol II/MED26 onto cMED by direct occlusion of their respective binding sites, leading to functional repression of cMED-dependent transcription. Combined with biochemical and functional analyses, these structures provide a conserved mechanistic framework to explain the basis for CKM-mediated repression of cMED function., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Mediator, a co-regulator complex required for RNA Polymerase II activity, interacts with tissue-specific transcription factors to regulate development and maintain homeostasis. We observe reduced Mediator subunit MED15 expression in endocrine hormone-producing pancreatic islets isolated from people living with type 2 diabetes and sought to understand how MED15 and Mediator control gene expression programs important for the function of insulin-producing β-cells. Here we show that Med15 is expressed during mouse β-cell development and maturation. Knockout of Med15 in mouse β-cells causes defects in β-cell maturation without affecting β-cell mass or insulin expression. ChIP-seq and co-immunoprecipitation analyses found that Med15 binds β-cell transcription factors Nkx6-1 and NeuroD1 to regulate key β-cell maturation genes. In support of a conserved role during human development, human embryonic stem cell-derived β-like cells, genetically engineered to express high levels of MED15, express increased levels of maturation markers. We provide evidence of a conserved role for Mediator in β-cell maturation and demonstrate an additional layer of control that tunes β-cell transcription factor function., (© 2024. The Author(s).)

Backgroud: Mediator complex subunit 19 (MED19), a member of the mediator complex, has been demonstrated to involve in tumorigenesis of hepatocellular carcinoma (HCC). However, the regulation mechanisms of MED19, the immune landscape linking MED19 to HCC and its predictive value of immunotherapy treatment in HCC are so far unknown., Methods: Here, we analyzed data from The Cancer Genome Atlas and other databases to assess the expression of MED19 and its prognosis and therapeutical-targets impact in HCC., Results: MED19 expression was upregulated in HCC tissues compared to non-tumorous liver tissues and that its upregulation was positively associated with advanced clinicopathology features. The multivariate analysis showed that MED19 was an independent predictor of outcome in HCC. In vitro experiments revealed that MED19 knockdown suppressed hepG2 cells proliferation, colony forming and invasion and induced apoptosis. Furthermore, MED19 inhibition resulted in G0/G1 phase arrest in hepG2 cells. We screened differentially expressed genes between low and high MED19 expression groups. Enrichment analyses showed that these genes were mainly linked to nuclear division and cell cycle. The pattern of tumor-infiltrating immune was demonstrated to be related with MED19 expression in HCC. TIDE analyses showed that patients in the low-expression group presented significantly better immunotherapy. Moreover, we developed a predicted model for HCC patient's prognosis. Receiver operating characteristic analyses revealed that this model processed a favorable performance in predicting the prognosis of HCC patients. Finally, a nomogram was built for predicting survival probability of individual HCC patient., Conclusion: These findings suggest that MED19 as a novel biomarker that has significant association with immune landscape and immunotherapy response in HCC. The proposed prediction model composed of MED19 and pathological stage has a better role in determining prognosis and stratifying of HCC., (© 2024. The Author(s).)

The members of the evolutionary conserved actin-binding Ezrin, Radixin and Moesin (ERM) protein family are involved in numerous key cellular processes in the cytoplasm. In the last decades, ERM proteins, like actin and other cytoskeletal components, have also been shown to be functional components of the nucleus; however, the molecular mechanism behind their nuclear activities remained unclear. Therefore, our primary aim was to identify the nuclear protein interactome of the single Drosophila ERM protein, Moesin. We demonstrate that Moesin directly interacts with the Mediator complex through direct binding to its Med15 subunit, and the presence of Moesin at the regulatory regions of the Hsp70Ab heat shock gene was found to be Med15-dependent. Both Moesin and Med15 bind to heat shock factor (Hsf), and they are required for proper Hsp gene expression under physiological conditions. Moreover, we confirmed that Moesin, Med15 and Hsf are able to bind the monomeric form of actin and together they form a complex in the nucleus. These results elucidate a mechanism by which ERMs function within the nucleus. Finally, we present the direct interaction of the human orthologues of Drosophila Moesin and Med15, which highlights the evolutionary significance of our finding.

Flooding impairs plant growth through oxygen deprivation, which activates plant survival and acclimation responses. Transcriptional responses to low oxygen are generally associated with the activation of group VII ETHYLENE-RESPONSE FACTOR (ERFVII) transcription factors. However, the exact mechanisms and molecular components by which ERFVII factors initiate gene expression are not fully elucidated. Here, we show that the ERFVII factors RELATED TO APETALA 2.2 (RAP2.2) and RAP2.12 cooperate with the Mediator complex subunit AtMED25 to coordinate gene expression under hypoxia in Arabidopsis thaliana. Respective med25 knock-out mutants display reduced low-oxygen stress tolerance. AtMED25 physically associates with a distinct set of hypoxia core genes and its loss partially impairs transcription under hypoxia due to decreased RNA polymerase II recruitment. Association of AtMED25 with target genes requires the presence of ERFVII transcription factors. Next to ERFVII protein stabilisation, also the composition of the Mediator complex including AtMED25 is potentially affected by hypoxia stress as shown by protein-complex pulldown assays. The dynamic response of the Mediator complex to hypoxia is furthermore supported by the fact that two subunits, AtMED8 and AtMED16, are not involved in the establishment of hypoxia tolerance, whilst both act in coordination with AtMED25 under other environmental conditions. We furthermore show that AtMED25 function under hypoxia is independent of ethylene signalling. Finally, functional conservation at the molecular level was found for the MED25-ERFVII module between A. thaliana and the monocot species Oryza sativa, pointing to a potentially universal role of MED25 in coordinating ERFVII-dependent transcript responses to hypoxia in plants., (© 2024 The Author(s). The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Objective: Molecular status of uterine leiomyomas has been shown to affect both tumor characteristics and treatment response. Mutations in mediator complex subunit 12 (MED12), the most prevalent alterations in leiomyomas, are associated with tumor size and number of leiomyomas. Myomectomy can be performed by laparoscopy or by open abdominal surgery, depending on the size and number of leiomyomas removed. The aim of this study was to examine the association between MED12 mutation status and surgical approach of myomectomy. We also evaluated myomectomy patients' quality of life after laparoscopic or abdominal surgery and according to the MED12 mutation status., Study Design: The prospective cohort study included 104 women who underwent laparoscopic or abdominal myomectomy at the Helsinki University Hospital during 2015-2019. Patients filled in the validated Uterine Fibroid Symptom and Quality of Life (UFS-QOL) questionnaire before the operation and 6 and 12 months after the operation. Medical records were reviewed to collect clinical data. Leiomyoma tissue samples were collected and screened for MED12 mutations., Results: Patients undergoing abdominal myomectomy had larger and more numerous leiomyomas compared to patients with laparoscopic myomectomy (10 cm vs 7.4 cm, p < 0.001 and 3 vs 1 leiomyomas, p < 0.001, respectively). A mean change of over 20 points was seen in UFS-QOL scores at 6 months after both laparoscopic and abdominal myomectomy (p < 0.001). MED12 mutations were detected in 178/242 (74 %) of leiomyomas. Of the patients, 45/97 (46 %) had only MED12 positive leiomyomas, while 39/97 (40 %) had only MED12 wild type leiomyomas. The number of leiomyomas removed was higher among patients with MED12 positive leiomyomas than in patients with MED12 wild type tumors (p < 0.001). Laparoscopic approach was equally common in both groups (62 % and 64 %), and there was no statistically significant difference in the UFS-QOL scores., Conclusion: Both laparoscopic and abdominal myomectomy significantly improved the quality of life. While MED12 mutations were related with multiple leiomyomas and therefore potentially generated a greater leiomyoma burden, they were not associated with the surgical approach. Pre- and postoperative quality of life was comparable between patients regardless of MED12 status., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose: Hardikar syndrome (HS, MIM #301068) is a female-specific multiple congenital anomaly syndrome characterized by retinopathy, orofacial clefting, aortic coarctation, biliary dysgenesis, genitourinary malformations, and intestinal malrotation. We previously showed that heterozygous nonsense and frameshift variants in MED12 cause HS. The phenotypic spectrum of disease and the mechanism by which MED12 variants cause disease is unknown. We aim to expand the phenotypic and molecular landscape of HS and elucidate the mechanism by which MED12 variants cause disease., Methods: We clinically assembled and molecularly characterized a cohort of 11 previously unreported individuals with HS. Additionally, we studied the effect of MED12 deficiency on ciliary biology, hedgehog, and yes-associated protein (YAP) signaling; pathways implicated in diseases with phenotypic overlap with HS., Results: We report novel phenotypes associated with HS, including cardiomyopathy, arrhythmia, and vascular anomalies, and expand the molecular landscape of HS to include splice site variants. We additionally demonstrate that MED12 deficiency causes decreased cell ciliation, and impairs hedgehog and YAP signaling., Conclusion: Our data support updating HS standard-of-care to include regular cardiac imaging, arrhythmia screening, and vascular imaging. We further propose that dysregulation of ciliogenesis and YAP and hedgehog signaling contributes to the pathogenesis of HS., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2024 American College of Medical Genetics and Genomics. Published by Elsevier Inc. All rights reserved.)

Mediator, a multiprotein complex, is an important component of the transcription machinery. In plants, the latest studies have established that it functions as a signal processor that conveys transcriptional signals from transcription factors to RNA polymerase II. Mediator has been found to be involved in different developmental and stress-adaptation conditions, ranging from embryo, root, and shoot development to flowering and senescence, and also in responses to different biotic and abiotic stresses. In the last decade, significant progress has been made in understanding the role of Mediator subunits in root development. They have been shown to transcriptionally regulate development of almost all the components of the root system architecture-primary root, lateral roots, and root hairs. They also have a role in nutrient acquisition by the root. In this review, we discuss all the known functions of Mediator subunits during root development. We also highlight the role of Mediator as a nodal point for processing different hormone signals that regulate root morphogenesis and growth., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Objective: Mediator complex subunit 12 (MED12) is among the most frequently mutated genes in various types of human cancers. However, there is still a lack of understanding regarding the role of MED12 in breast cancer patient. Therefore, the aim of this study is to explore the roles of MED12 in breast cancer. Materials and Methods: We utilized the UALCAN platform (http://ualcan.path.uab.edu/) for analyzing the transcriptional expression, protein expression, and protein phosphorylation data of MED12. Our study involved 35 breast cancer patients. From these samples, we extracted proteins and RNA. To obtain the sequence of MED12 3'-UTR, we performed reverse transcription-polymerase chain reaction and sequencing. We then used TargetScan to predict the miRNA targets of MED12 3'-UTR and confirmed the interactions between miRNAs and MED12 3'-UTR through dual luciferase assay. Results: The protein level of MED12 was upregulated in breast cancer, while the mRNA level did not show significant changes. Interestingly, higher levels of MED12 mRNA were associated with better prognosis, whereas patients with increased MED12 protein levels tended to have a poorer prognosis. Furthermore, through our analysis of the MED12 3'-UTR sequence, we identified a specific C->T variation that was unique to breast tumors. We also identified four miRNAs (miR-204, -211, -450 b, and -518a) that directly target MED12 3'-UTR. Most important, this C->T variation disrupts the interaction between MED12 3'-UTR and miR-450b, ultimately leading to the upregulation of MED12 in breast cancer. Conclusion: Our study revealed a significant finding regarding a mutation site in the MED12 3'-UTR that contributes to the upregulation of MED12 in breast cancer. This mutation disrupts the interactions between specific miRNAs and MED12 mRNA, leading to increased expression of MED12. These findings have important implications for breast cancer diagnosis, as this mutation site can serve as a potent biomarker.

The Mediator complex is an essential coregulator of RNA polymerase II transcription. More recent developments suggest Mediator functions as a link between transcription regulation, genome organisation and DNA repair mechanisms including nucleotide excision repair, base excision repair, and homologous recombination. Dysfunctions of these processes are frequently associated with human pathologies, and growing evidence shows Mediator involvement in cancers, neurological, metabolic and infectious diseases. The detailed deciphering of molecular mechanisms of Mediator functions, using interdisciplinary approaches in different biological models and considering all functions of this complex, will contribute to our understanding of relevant human diseases., Competing Interests: Declaration of Competing Interest We hereby declare that we do not have any conflicts of interests regarding this manuscript., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Abstract: This is the first report of pediatric catatonia syndrome in MED13L haploinsufficiency syndrome. This report describes unique challenges in diagnosis and management of catatonia in rare genetic conditions. The case also illustrates the use of electroconvulsive therapy in patients with epilepsy, epileptic encephalopathy, or other epileptic diathesis and the clinical conundrum in determining the course of maintenance electroconvulsive therapy., Competing Interests: Conflict of Interest: Dr. Neera Ghaziuddin declares following conflict of interests: 1. royalties from Oxford University Press for book titled, "Use of Electroconvulsive Therapy in Children and Adolescents"; 2. honorarium for invited presentation at Brown University, Rhode Island; and 3. honorarium (travel support only) for invited presentation, "The Maudsley Forum, Scandinavia" at Lund University, Helsingborg, Sweden., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

The protein-protein interaction (PPI) network of the Mediator complex is very tightly regulated and depends on different developmental and environmental cues. Here, we present an interactive platform for comparative analysis of the Mediator subunits from humans, baker's yeast Saccharomyces cerevisiae, and model plant Arabidopsis thaliana in a user-friendly web-interface database called MediatorWeb. MediatorWeb provides an interface to visualize and analyze the PPI network of Mediator subunits. The database facilitates downloading the untargeted and unweighted network of Mediator complex, its submodules, and individual Mediator subunits to better visualize the importance of individual Mediator subunits or their submodules. Further, MediatorWeb offers network visualization of the Mediator complex and interacting proteins that are functionally annotated. This feature provides clues to understand functions of Mediator subunits in different processes. In an additional tab, MediatorWeb provides quick access to secondary and tertiary structures, as well as residue-level contact information for Mediator subunits in each of the three model organisms. Another useful feature of MediatorWeb is detection of interologs based on orthologous analyses, which can provide clues to understand the functions of Mediator complex in less explored kingdoms. Thus, MediatorWeb and its features can help the user to understand the role of Mediator complex and its subunits in the transcription regulation of gene expression., (© 2024 Federation of European Biochemical Societies.)

Background: Black women are at an increased risk of developing uterine leiomyomas and experiencing worse disease prognosis than White women. Epidemiologic and molecular factors have been identified as underlying these disparities, but there remains a paucity of deep, multiomic analysis investigating molecular differences in uterine leiomyomas from Black and White patients., Objective: To identify molecular alterations within uterine leiomyoma tissues correlating with patient race by multiomic analyses of uterine leiomyomas collected from cohorts of Black and White women., Study Design: We performed multiomic analysis of uterine leiomyomas from Black (42) and White (47) women undergoing hysterectomy for symptomatic uterine leiomyomata. In addition, our analysis included the application of orthogonal methods to evaluate fibroid biomechanical properties, such as second harmonic generation microscopy, uniaxial compression testing, and shear-wave ultrasonography analyses., Results: We found a greater proportion of MED12 mutant uterine leiomyomas from Black women (>35% increase; Mann-Whitney U, P<.001). MED12 mutant tumors exhibited an elevated abundance of extracellular matrix proteins, including several collagen isoforms, involved in the regulation of the core matrisome. Histologic analysis of tissue fibrosis using trichrome staining and secondary harmonic generation microscopy confirmed that MED12 mutant tumors are more fibrotic than MED12 wild-type tumors. Using shear-wave ultrasonography in a prospectively collected cohort, Black patients had fibroids that were firmer than White patients, even when similar in size. In addition, these analyses uncovered ancestry-linked expression quantitative trait loci with altered allele frequencies in African and European populations correlating with differential abundance of several proteins in uterine leiomyomas independently of MED12 mutation status, including tetratricopeptide repeat protein 38., Conclusion: Our study shows that Black women have a higher prevalence of uterine leiomyomas harboring mutations in MED12 and that this mutational status correlates with increased tissue fibrosis compared with wild-type uterine leiomyomas. Our study provides insights into molecular alterations correlating with racial disparities in uterine leiomyomas and improves our understanding of the molecular etiology underlying uterine leiomyoma development within these populations., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The Mediator complex is essential for eukaryotic transcription, yet its role and the function of its individual subunits in plants, especially in rice, remain poorly understood. Here, we investigate the function of OsMED14&#95;2, a subunit of the Mediator tail module, in rice development. Overexpression and knockout of OsMED14&#95;2 resulted in notable changes in panicle morphology and grain size. Microscopic analysis revealed impact of overexpression on pollen maturation, reflected by reduced viability, irregular shapes, and aberrant intine development. OsMED14&#95;2 was found to interact with proteins involved in pollen development, namely, OsMADS62, OsMADS63 and OsMADS68, and its overexpression negatively affected the expression of OsMADS68 and the expression of other genes involved in intine development, including OsCAP1, OsGCD1, OsRIP1, and OsCPK29. Additionally, we found that OsMED14&#95;2 overexpression influences jasmonic acid (JA) homeostasis, affecting bioactive JA levels, and expression of OsJAZ genes. Our data suggest OsMED14&#95;2 may act as a regulator of JA-responsive genes through its interactions with OsHDAC6 and OsJAZ repressors. These findings contribute to better understanding of the Mediator complex's role in plant traits regulation., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The phenotypes associated with MED12 pathogenic variants are diverse. Male patients usually have missense variants, but the effects of base substitutions on mRNA splicing have not been investigated. Here, we report a Japanese brother with intellectual disability, characteristic facial appearance with blepharophimosis, cleft palate, Fallot tetralogy, vesicoureteral reflux, and deafness. A known missense pathogenic variant was detected in MED12, NM&#95;005120.3:c.887G>A p.(Arg296Gln), and X-linked Ohdo syndrome was diagnosed in combination with their phenotype. mRNA splicing of MED12 was evaluated qualitatively and quantitatively using long-range PCR-based targeted RNA sequencing (reverse transcribed long amplicon sequencing), and it was shown that this missense variant simultaneously causes aberrant splicing of the 42-bp in-frame deletion in exon 7, r.847&#95;888del, which accounts for approximately 30% of the mRNAs in both siblings. The X chromosome inactivation study showed that the X chromosome carrying the mutant allele was 100% inactivated in the carrier mothers. mRNA level analysis is essential for the accurate interpretation of the effects of variants. In this case, the MED12 protein function may be reduced by more than just an amino acid substitution, resulting in the patients with the most severe phenotype of MED12-related syndrome in males., (© 2024 Wiley Periodicals LLC.)

Over 70% of leiomyoma (LM) harbor MED12 mutations, primarily in exon 2 at c.130-131 (GG). Myometrial cells are the cell origin of leiomyoma, but the MED12 mutation status in non-neoplastic myometrial cells is unknown. In this study, we investigated the mutation burden of MED12 in myometrium. As traditional Sanger or even NGS sequencing may not be able to detect MED12 mutations that are lower than 0.1% in the testing sample, we used duplex deep sequencing analysis (DDS) to overcome this limitation. Tumor-free myometria (confirmed by pathology evaluation) were dissected, and genomic DNA from MED12 exon 2 (test) and TP53 exon 5 (control) were captured by customer-designed probe sets, followed by DDS. Notably, DDS demonstrated that myometrial cells harbored a high frequency of mutations in MED12 exon 2 and predominantly in code c.130-131. In contrast, the baseline mutations in other coding sequences of MED12 exon 2 as well as in the TP53 mutation hotspot, c.477-488 were comparably low in myometrial cells. This is the first report demonstrating a non-random accumulation of MED12 mutations at c.130-131 sites in non-neoplastic myometrial cells which provide molecular evidence of early somatic mutation events in myometrial cells. This early mutation may contribute to the cell origin for uterine LM development in women of reproductive age.

Prostate cancer progression is driven by androgen receptor (AR) activity, which is a target for therapeutic approaches. Enzalutamide is an AR inhibitor that prolongs the survival of patients with advanced prostate cancer. However, resistance mechanisms arise and impair its efficacy. One of these mechanisms is the expression of AR-V7, a constitutively active AR splice variant. The Mediator complex is a multisubunit protein that modulates gene expression on a genome-wide scale. MED12 and cyclin-dependent kinase (CDK)8, or its paralog CDK19, are components of the kinase module that regulates the proliferation of prostate cancer cells. In this study, we investigated how MED12 and CDK8/19 influence cancer-driven processes in prostate cancer cell lines, focusing on AR activity and the enzalutamide response. We inhibited MED12 expression and CDK8/19 activity in LNCaP (AR+, enzalutamide-sensitive), 22Rv1 (AR-V7+, enzalutamide-resistant), and PC3 (AR-, enzalutamide-insensitive) cells. Both MED12 and CDK8/19 inhibition reduced cell proliferation in all cell lines, and MED12 inhibition reduced proliferation in the respective 3D spheroids. MED12 knockdown significantly inhibited c-Myc protein expression and signaling pathways. In 22Rv1 cells, it consistently inhibited the AR response, prostate-specific antigen (PSA) secretion, AR target genes, and AR-V7 expression. Combined with enzalutamide, MED12 inhibition additively decreased the AR activity in both LNCaP and 22Rv1 cells. CDK8/19 inhibition significantly decreased PSA secretion in LNCaP and 22Rv1 cells and, when combined with enzalutamide, additively reduced proliferation in 22Rv1 cells. Our study revealed that MED12 and CDK8/19 regulate AR activity and that their inhibition may modulate response to enzalutamide in prostate cancer., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

We describe a time-resolved nascent single-cell RNA sequencing (RNA-seq) approach that measures gene-specific transcriptional noise and the fraction of active genes in S. cerevisiae. Most genes are expressed with near-constitutive behavior, while a subset of genes show high mRNA variance suggestive of transcription bursting. Transcriptional noise is highest in the cofactor/coactivator-redundant (CR) gene class (dependent on both SAGA and TFIID) and strongest in TATA-containing CR genes. Using this approach, we also find that histone gene transcription switches from a low-level, low-noise constitutive mode during M and M/G1 to an activated state in S phase that shows both an increase in the fraction of active promoters and a switch to a noisy and bursty transcription mode. Rapid depletion of cofactors SAGA and MED Tail indicates that both factors play an important role in stimulating the fraction of active promoters at CR genes, with a more modest role in transcriptional noise., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Complementary studies by Zhao et al. 1 and Chen et al. 2 reveal how an intrinsically disordered region in MED13 controls mutually exclusive binding of RNA Polymerase II and CDK8 kinase module to Mediator, switching Mediator and transcription activation on and off., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Hyperosmotic stress tolerance is crucial for Saccharomyces cerevisiae in producing value-added products from renewable feedstock. The limited understanding of its tolerance mechanism has impeded the application of these microbial cell factories. Previous studies have shown that Med3 plays a role in hyperosmotic stress in S. cerevisiae . However, the specific function of Med3 in hyperosmotic stress tolerance remains unclear. In this study, we showed that the deletion of the mediator Med3 impairs S. cerevisiae growth under hyperosmotic stress. Phenotypic analyses and yeast two-hybrid assays revealed that Med3 interacts with the transcription factor Stb5 to regulate the expression of the genes gnd1 and ald6 , which are involved in NADPH production under hyperosmotic stress conditions. The deletion of med3 resulted in a decrease in intracellular NADPH content, leading to increased oxidative stress and elevated levels of intracellular reactive oxygen species under hyperosmotic stress, thereby impacting bud formation. These findings highlight the significant role of Med3 as a regulator in maintaining NADPH generation and redox homeostasis in S. cerevisiae during hyperosmotic stress.IMPORTANCEHyperosmotic stress tolerance in the host strain is a significant challenge for fermentation performance in industrial production. In this study, we showed that the S. cerevisiae mediator Med3 is essential for yeast growth under hyperosmotic conditions. Med3 interacts with the transcription factor Stb5 to regulate the expression of genes involved in the NADPH-generation system during hyperosmotic stress. Adequate NADPH ensures the timely removal of excess reactive oxygen species and supports bud formation under these conditions. This work highlights the crucial role of Med3 as a regulator in maintaining NADPH generation and redox homeostasis in S. cerevisiae during hyperosmotic stress., Competing Interests: The authors declare no conflict of interest.

Discontinuous transcription is evolutionarily conserved and a fundamental feature of gene regulation; yet, the exact mechanisms underlying transcriptional bursting are unresolved. Analyses of bursting transcriptome-wide have focused on the role of cis-regulatory elements, but other factors that regulate this process remain elusive. We applied mathematical modeling to single-cell RNA sequencing data to infer bursting dynamics transcriptome-wide under multiple conditions to identify possible molecular mechanisms. We found that Mediator complex subunit 26 (MED26) primarily regulates frequency, MYC regulates burst size, while cohesin and Bromodomain-containing protein 4 (BRD4) can modulate both. Despite comparable effects on RNA levels among these perturbations, acute depletion of MED26 had the most profound impact on the entire gene regulatory network, acting downstream of chromatin spatial architecture and without affecting TATA box-binding protein (TBP) recruitment. These results indicate that later steps in the initiation of transcriptional bursts are primary nodes for integrating gene networks in single cells.

The mediator complex subunit 13 (MED13) gene is implicated in neurodevelopmental disorders including autism spectrum disorder (ASD), intellectual disability, and speech delay with varying severity and course. Additional, extra central nervous system, features include eye or vision problems, hypotonia, congenital heart abnormalities, and dysmorphisms. We describe a 7-year- and 4-month-old girl evaluated for ASD whose brain magnetic resonance imaging was suggestive of multiple cortical tubers. The exome sequencing (ES - trio analysis) uncovered a unique, de novo, frameshift variant in the MED13 gene (c.4880del, D1627Vfs*17), with a truncating effect on the protein. This case report thus expands the phenotypic spectrum of MED13-related disorders to include brain abnormalities., (© 2024 Wiley Periodicals LLC.)

The essential Mediator (MED) coactivator complex plays a well-understood role in regulation of basal transcription in all eukaryotes, but the mechanism underlying its role in activator-dependent transcription remains unknown. We investigated modulation of metazoan MED interaction with RNA polymerase II (RNA Pol II) by antagonistic effects of the MED26 subunit and the CDK8 kinase module (CKM). Biochemical analysis of CKM-MED showed that the CKM blocks binding of the RNA Pol II carboxy-terminal domain (CTD), preventing RNA Pol II interaction. This restriction is eliminated by nuclear receptor (NR) binding to CKM-MED, which enables CTD binding in a MED26-dependent manner. Cryoelectron microscopy (cryo-EM) and crosslinking-mass spectrometry (XL-MS) revealed that the structural basis for modulation of CTD interaction with MED relates to a large intrinsically disordered region (IDR) in CKM subunit MED13 that blocks MED26 and CTD interaction with MED but is repositioned upon NR binding. Hence, NRs can control transcription initiation by priming CKM-MED for MED26-dependent RNA Pol II interaction., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objectives: TFE3-rearranged renal cell carcinomas (RCCs) harbor gene fusions between TFE3 and 1 of many partner genes. MED15::TFE3 fusion RCC is rare, often cystic, and easily misdiagnosed., Methods: This study aimed to characterize 2 cases of MED15::TFE3 fusion RCC with extensive cystic change using fluorescence in situ hybridization and targeted RNA sequencing., Results: Both patients were young adult women aged 29 and 35 years. Radiologically, both presented with a cystic Bosniak category II renal lesion. The cysts measured 9.3 cm and 4.8 cm in greatest dimension. Both patients underwent cyst enucleation, and neither had tumor recurrence or metastasis at 26 and 6 months of follow-up, respectively. Microscopically, both tumors were entirely cystic, with thick, fibrous cystic walls lined by small clusters of cells with clear to eosinophilic cytoplasm and uniform, round nuclei with inconspicuous nucleoli. There were also small aggregations of similar clear cells within the cystic walls. Foci of basement membrane-like material depositions were noted in 1 case; calcifications were observed in both cases. Both cases demonstrated nuclear positivity for PAX8 and TFE3 and cytoplasmic staining for Melan-A; HMB45, CAIX, and CK7 were negative. Fluorescence in situ hybridization revealed that both tumors were positive for TFE3 rearrangements. RNA sequencing identified MED15::TFE3 gene fusions in both cases., Conclusions: The main differential diagnosis of MED15::TFE3 fusion RCC includes multilocular cystic renal neoplasm of low malignant potential and atypical renal cysts. Molecular confirmation of TFE3 fusion is essential for establishing the correct diagnosis., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society for Clinical Pathology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

The presence of basal lineage characteristics signifies hyperaggressive human adenocarcinomas of the breast, bladder and pancreas. However, the biochemical mechanisms that maintain this aberrant cell state are poorly understood. Here we performed marker-based genetic screens in search of factors needed to maintain basal identity in pancreatic ductal adenocarcinoma (PDAC). This approach revealed MED12 as a powerful regulator of the basal cell state in this disease. Using biochemical reconstitution and epigenomics, we show that MED12 carries out this function by bridging the transcription factor ΔNp63, a known master regulator of the basal lineage, with the Mediator complex to activate lineage-specific enhancer elements. Consistent with this finding, the growth of basal-like PDAC is hypersensitive to MED12 loss when compared to PDAC cells lacking basal characteristics. Taken together, our genetic screens have revealed a biochemical interaction that sustains basal identity in human cancer, which could serve as a target for tumor lineage-directed therapeutics., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Trio exome sequencing was performed on a female fetus with an increased nuchal translucency, along with nasal bone hypoplasia, suspected cleft palate and abnormal outflow tract of the heart. A de novo heterozygous variant c.5500&#95;5507del, p.(Tyr1834Argfs × 58) in the MED12 gene was detected. Loss-of-function variants in MED12 in females are associated with Hardikar syndrome (HS). A follow-up ultrasound at 15 +5  weeks of gestation identified multiple fetal anomalies including bilateral cleft lip and palate, diaphragmatic hernia, atrioventricular septal defect, persistent truncus arteriosus, and bilateral renal pelvis dilation. Fetal autopsy confirmed the prenatal sonographic findings, and the MED12 variant was discussed by our multidisciplinary team to be the cause of fetal anomalies. Our case is the first prenatal one in which HS was diagnosed due to first trimester structural malformations. This case report presents another example of early identification of a major anomaly which allows earlier genetic diagnosis and more time for clinical management., (© 2024 John Wiley & Sons Ltd.)

Thirty years have passed since the discovery of the Mediator complex in yeast. We are witnessing breakthroughs and advances that have led to high-resolution structural models of yeast and mammalian Mediators in the preinitiation complex, showing how it is assembled and how it positions the RNA polymerase II and its C-terminal domain (CTD) to facilitate the CTD phosphorylation that initiates transcription. This information may be also used to guide future plant research on the mechanisms of Mediator transcriptional control. Here, we review what we know about the subunit composition and structure of plant Mediators, the roles of the individual subunits and the genetic analyses that pioneered Mediator research, and how transcription factors recruit Mediators to regulatory regions adjoining promoters. What emerges from the research is a Mediator that regulates transcription activity and recruits hormonal signaling modules and histone-modifying activities to set up an off or on transcriptional state that recruits general transcription factors for preinitiation complex assembly.

The ability to sense and respond to proteotoxic insults declines with age, leaving cells vulnerable to chronic and acute stressors. Reproductive cues modulate this decline in cellular proteostasis to influence organismal stress resilience in Caenorhabditis elegans We previously uncovered a pathway that links the integrity of developing embryos to somatic health in reproductive adults. Here, we show that the nuclear receptor NHR-49, an ortholog of mammalian peroxisome proliferator-activated receptor α (PPARα), regulates stress resilience and proteostasis downstream from embryo integrity and other pathways that influence lipid homeostasis and upstream of HSF-1. Disruption of the vitelline layer of the embryo envelope, which activates a proteostasis-enhancing intertissue pathway in somatic cells, triggers changes in lipid catabolism gene expression that are accompanied by an increase in fat stores. NHR-49, together with its coactivator, MDT-15, contributes to this remodeling of lipid metabolism and is also important for the elevated stress resilience mediated by inhibition of the embryonic vitelline layer. Our findings indicate that NHR-49 also contributes to stress resilience in other pathways known to change lipid homeostasis, including reduced insulin-like signaling and fasting, and that increased NHR-49 activity is sufficient to improve proteostasis and stress resilience in an HSF-1-dependent manner. Together, our results establish NHR-49 as a key regulator that links lipid homeostasis and cellular resilience to proteotoxic stress., (© 2024 Sala et al.; Published by Cold Spring Harbor Laboratory Press.)

Mediator is a well-known transcriptional co-regulator and serves as an adaptor between gene-specific regulatory proteins and RNA polymerase II. Studies on the chromatin-bound form of Mediator revealed interactions with additional protein complexes involved in various transcription-related processes, such as the Lsm2-8 complex that is part of the spliceosomal U6 small nuclear ribonucleoprotein complex. Here, we employ Chromatin Immunoprecipitation sequencing (ChIP-seq) of chromatin associated with the Lsm3 protein and the Med1 or Med15 Mediator subunits. We identify 86 genes co-occupied by both Lsm3 and Mediator, of which 73 were intron-containing ribosomal protein genes. In logarithmically growing cells, Mediator primarily binds to their promoter regions but also shows a second, less pronounced occupancy at their 3'-exons. During the late exponential phase, we observe a near-complete transition of Mediator from these promoters to a position in their 3'-ends, overlapping the Lsm3 binding sites ∼250 bp downstream of their last intron-exon boundaries. Using an unbiased RNA sequencing approach, we show that transition of Mediator from promoters to the last exon of these genes correlates to reduction of both their messenger RNA levels and splicing ratios, indicating that the Mediator and Lsm complexes cooperate to control growth-regulated expression of intron-containing ribosomal protein genes at the levels of transcription and splicing., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Genetic sequencing has identified high-confidence ASD risk genes with loss-of-function mutations. How the haploinsufficiency of distinct ASD risk genes causes ASD remains to be elucidated. In this study, we examined the role of four top-ranking ASD risk genes, ADNP, KDM6B, CHD2, and MED13, in gene expression regulation. ChIP-seq analysis reveals that gene targets with the binding of these ASD risk genes at promoters are enriched in RNA processing and DNA repair. Many of these targets are found in ASD gene database (SFARI), and are involved in transcription regulation and chromatin remodeling. Common gene targets of these ASD risk genes include a network of high confidence ASD genes associated with gene expression regulation, such as CTNNB1 and SMARCA4. We further directly examined the transcriptional impact of the deficiency of these ASD risk genes. Our mRNA profiling with qPCR assays in cells with the knockdown of Adnp, Kdm6b, Chd2 or Med13 has revealed an intricate pattern of their cross-regulation, as well as their influence on the expression of other ASD genes. In addition, some synaptic genes, such as Snap25 and Nrxn1, are strongly regulated by deficiency of the four ASD risk genes, which could be through the direct binding at promoters or indirectly through the targets like Ctnnb1 or Smarca4. The identification of convergent and divergent gene targets that are regulated by multiple ASD risk genes will help to understand the molecular mechanisms underlying common and unique phenotypes associated with haploinsufficiency of ASD-associated genes., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Cyclin-dependent kinase 7 (CDK7), part of the general transcription factor TFIIH, promotes gene transcription by phosphorylating the C-terminal domain of RNA polymerase II (RNA Pol II). Here, we combine rapid CDK7 kinase inhibition with multi-omics analysis to unravel the direct functions of CDK7 in human cells. CDK7 inhibition causes RNA Pol II retention at promoters, leading to decreased RNA Pol II initiation and immediate global downregulation of transcript synthesis. Elongation, termination, and recruitment of co-transcriptional factors are not directly affected. Although RNA Pol II, initiation factors, and Mediator accumulate at promoters, RNA Pol II complexes can also proceed into gene bodies without promoter-proximal pausing while retaining initiation factors and Mediator. Further downstream, RNA Pol II phosphorylation increases and initiation factors and Mediator are released, allowing recruitment of elongation factors and an increase in RNA Pol II elongation velocity. Collectively, CDK7 kinase activity promotes the release of initiation factors and Mediator from RNA Pol II, facilitating RNA Pol II escape from the promoter., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

X-linkded Ohdo syndrome is characterized mainly by intellectual disability, delays in reaching development, feeding difficulties, thyroid dysfunction, and dysmorphic appearance with blepharophimosis, immobile mask-like face and bulbous nose. The X-linked Ohdo syndrome is caused by loss of function mutation in MED12 gene on X chromosome. The peripheral blood mononuclear cells from a patient carrying missense mutation of the MED12 gene were reprogrammed using the CytoTune-iPS2.0 Sendai Reprogramming Kit. The missense mutation in MED12 gene causes the abnormal protein variant. The established human induced pluripotent cell line will enable proper in vitro disease modelling of X-linked Ohdo syndrome., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Genetic alterations are well characterized as contributors to the pathogenesis of cancers. Epigenetic abnormalities can lead to perturbations of the expression of genes in cancer cells without structural defects. Deregulation of proteins of the transcription machinery may result in perturbations of target genes. Mediator, a multiprotein component of the transcription machinery facilitates the function of RNA polymerase II, which transcribes most human genes. A part of the mediator with kinase activity, called the Mediator kinase module shows genetic alterations in a sub-set of colorectal cancers., Methods: Data from publicly available genomic series of colorectal cancer patients were examined to determine alterations of Mediator kinase module component genes, including MED12, MED12L, MED13, MED13L, CDK8, CDK19, and CCNC. The prevalence of alterations in genomically defined colorectal cancer sub-sets was also interrogated. The effect of Mediator kinase module member gene expression on colorectal cancer relapse-free survival was investigated., Results: Mutations in genes of the Mediator kinase module were present in a small percentage of colorectal cancers, ranging between 2 to 10% for MED12 and MED13 and alternative units MED12L and MED13L and below 2% for kinases CDK8 and CDK19 and cyclin C. Amplifications of the CDK8 gene were observed in 3% to 5% of colorectal cancers. The highest prevalence of mutations was observed in MSI cancers and the equivalent CMS1 group, with other genomic groups showing much lower frequency. An association of higher expression of MED12 with inferior relapse-free survival was observed. In contrast, higher expression of cyclin C was associated with improved survival. Colorectal cancer cell lines with CDK8 amplifications displayed sensitivity to several small molecule inhibitors of the KRAS/PI3K pathway but not to BET inhibitors., Conclusion: The Mediator kinase module is deregulated in a sub-set of colorectal cancers with differences observed in genomically defined groups. These variations may result in differences in sensitivity to targeted therapies and may have to be taken into consideration as such therapies are developed., (© 2024. The Author(s) under exclusive licence to Maj Institute of Pharmacology Polish Academy of Sciences.)

Muscle stem cells (MuSCs) contribute to a robust muscle regeneration process after injury, which is highly orchestrated by the sequential expression of multiple key transcription factors. However, it remains unclear how key transcription factors and cofactors such as the Mediator complex cooperate to regulate myogenesis. Here, we show that the Mediator Med23 is critically important for MuSC-mediated muscle regeneration. Med23 is increasingly expressed in activated/proliferating MuSCs on isolated myofibers or in response to muscle injury. Med23 deficiency reduced MuSC proliferation and enhanced its precocious differentiation, ultimately compromising muscle regeneration. Integrative analysis revealed that Med23 oppositely impacts Ternary complex factor (TCF)-targeted MuSC proliferation genes and myocardin-related transcription factor (MRTF)-targeted myogenic differentiation genes. Consistently, Med23 deficiency decreases the ETS-like transcription factor 1 (Elk1)/serum response factor (SRF) binding at proliferation gene promoters but promotes MRTF-A/SRF binding at myogenic gene promoters. Overall, our study reveals the important transcriptional control mechanism of Med23 in balancing MuSC proliferation and differentiation in muscle regeneration., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Whole exome sequencing allows rapid identification of causative single nucleotide variants and short insertions/deletions in children with congenital anomalies and/or intellectual disability, which aids in accurate diagnosis, prognosis, appropriate therapeutic interventions, and family counselling. Recently, de novo variants in the MED13 gene were described in patients with an intellectual developmental disorder that included global developmental delay, mild congenital heart anomalies, and hearing and vision problems in some patients., Results: Here we describe an infant who carried a de novo p.Pro835Ser missense variant in the MED13 gene, according to whole exome trio sequencing. He presented with congenital heart anomalies, dysmorphic features, hydrocephalic changes, hypoplastic corpus callosum, bilateral optic nerve atrophy, optic chiasm atrophy, brain stem atrophy, and overall a more severe condition compared to previously described patients., Conclusions: Therefore, we propose to expand the MED13-associated phenotype to include severe complications that could end up with multiple organ failure and neonatal death., (© 2024. The Author(s).)

DELLA proteins are negative regulators of the gibberellin response pathway in angiosperms, acting as central hubs that interact with hundreds of transcription factors (TFs) and regulators to modulate their activities. While the mechanism of TF sequestration by DELLAs to prevent DNA binding to downstream targets has been extensively documented, the mechanism that allows them to act as coactivators remains to be understood. Here, we demonstrate that DELLAs directly recruit the Mediator complex to specific loci in Arabidopsis, facilitating transcription. This recruitment involves DELLA amino-terminal domain and the conserved MED15 KIX domain. Accordingly, partial loss of MED15 function mainly disrupted processes known to rely on DELLA coactivation capacity, including cytokinin-dependent regulation of meristem function and skotomorphogenic response, gibberellin metabolism feedback, and flavonol production. We have also found that the single DELLA protein in the liverwort Marchantia polymorpha is capable of recruiting MpMED15 subunits, contributing to transcriptional coactivation. The conservation of Mediator-dependent transcriptional coactivation by DELLA between Arabidopsis and Marchantia implies that this mechanism is intrinsic to the emergence of DELLA in the last common ancestor of land plants., Competing Interests: Competing interests statement:The authors declare no competing interest.

The mediator complex subunit 23 ( MED23 ) gene encodes a protein that acts as a tail module mediator complex, a multi-subunit co-activator involved in several cellular activities. MED23 has been shown to have substantial roles in myogenesis and other molecular mechanisms. The functions of MED23 in the neurological system remain unclear and the clinical phenotype is not thoroughly described. Whole exome sequencing was used to identify a novel mutation in the MED23 gene. DNA capture probes using next-generation sequencing-based copy number variation analysis with Illumina array were performed. The clinical, demographic, neuroimaging, and electrophysiological data of the patients were collected, and similarly, the data of all reported cases in the literature were extracted to compare findings. Screening a total of 9,662 articles, we identified 22 main regulatory processes for the MED23 gene, including suppressive activity for carcinogenic processes. MED23 is also involved in the brain's neurogenesis and functions. The identified cases mainly presented with intellectual disability (87.5%) and developmental delay (50%). Seizures were present in only 18.75% of the patients. Slow backgrounds and spike and sharp-wave complexes were reported on the electroencephalogram (EEG) of a few patients and delayed myelination, thin corpus callosum, and pontine hypoplasia on magnetic resonance imaging (MRI). The MED23 gene regulates several processes in which its understanding promotes considerable therapeutic potential for patients. It is crucial to consider genetic and laboratory testing, particularly when encountering potential carriers. Intellectual disability and developmental delay are the most notable clinical signs with heterogeneous features on EEG and MRI., Competing Interests: The authors declare no conflict of interest., (© 2024 by the authors.)

Breast cancer is often treated with chemotherapy. However, the development of chemoresistance results in treatment failure. Long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) has been shown to contribute to chemoresistance in breast cancer cells. In studying the transcriptional regulation of NEAT1 using multi-omics approaches, we showed that NEAT1 is up-regulated by 5-fluorouracil in breast cancer cells with wild-type cellular tumor antigen p53 but not in mutant-p53-expressing breast cancer cells. The regulation of NEAT1 involves mediator complex subunit 12 (MED12)-mediated repression of histone acetylation marks at the promoter region of NEAT1. Knockdown of MED12 but not coactivator-associated arginine methyltransferase 1 (CARM1) induced histone acetylation at the NEAT1 promoter, leading to elevated NEAT1 mRNAs, resulting in a chemoresistant phenotype. The MED12-dependent regulation of NEAT1 differs between wild-type and mutant p53-expressing cells. MED12 depletion led to increased expression of NEAT1 in a wild-type p53 cell line, but decreased expression in a mutant p53 cell line. Chemoresistance caused by MED12 depletion can be partially rescued by NEAT1 knockdown in p53 wild-type cells. Collectively, our study reveals a novel mechanism of chemoresistance dependent on MED12 transcriptional regulation of NEAT1 in p53 wild-type breast cancer cells., (© 2024 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Lipid droplets (LDs) are organelles that store excess lipids and provide fatty acids for energy production during starvation. LDs are also essential for cellular maintenance, but excessive accumulation of LDs triggers various cancers in addition to metabolic diseases such as diabetes. In this study, we aimed to develop a strategy to identify new genes that reduces accumulation of LDs in cancer cells using an RNA interference (RNAi) screening system employing artificial sequence-enriched shRNA libraries. Monitoring LDs by fluorescent activated cell sorting, the subsequently collected cumulative LDs cells, and shRNA sequence analysis identified a clone that potentially functioned to accumulate LDs. The clone showed no identical sequence to human Refseq. It showed very similar sequence to seven genes by allowing three mismatches. Among these genes, we identified the mediator complex subunit 6 (MED6) gene as a target of this shRNA. Silencing of MED6 led to an increase in LD accumulation and expression of the marker genes, PLIN2 and DGAT1, in fatty cells. MED6 is a member of the mediator complex that regulates RNA polymerase II transcription through transcription factor II. Some mediator complexes play important roles in both normal and pathophysiological transcription processes. These results suggest that MED6 transcriptionally regulates the genes involved in lipid metabolism and suppresses LD accumulation., (© 2023 International Union of Biochemistry and Molecular Biology.)

The human Mediator complex subunit MED25 binds transactivation domains (TADs) present in various cellular and viral proteins using two binding interfaces, named H1 and H2, which are found on opposite sides of its ACID domain. Here, we use and compare deep learning methods to characterize human MED25-TAD interfaces and assess the predicted models to published experimental data. For the H1 interface, AlphaFold produces predictions with high-reliability scores that agree well with experimental data, while the H2 interface predictions appear inconsistent, preventing reliable binding modes. Despite these limitations, we experimentally assess the validity of MED25 interface predictions with the viral transcriptional activators Lana-1 and IE62. AlphaFold predictions also suggest the existence of a unique hydrophobic pocket for the Arabidopsis MED25 ACID domain., (© 2024 Federation of European Biochemical Societies.)

Familial dilated cardiomyopathy (DCM) is among the leading indications for heart transplantation. DCM alters the transcriptomic profile. The alteration or activation/silencing of physiologically operating transcripts may explain the onset and progression of this pathological state. The mediator complex (MED) plays a fundamental role in the transcription process. The aim of this study is to investigate the MED subunits, which are altered in DCM, to identify target crossroads genes. RNA sequencing allowed us to identify specific MED subunits that are altered during familial DCM, transforming into human myocardial samples. N = 13 MED subunits were upregulated and n = 7 downregulated. MED9 alone was significantly reduced in patients compared to healthy subjects (HS) (FC = -1.257; p < 0.05). Interestingly, we found a short MED9 isoform (MED9s) (ENSG00000141026.6), which was upregulated when compared to the full-transcript isoform (MED9f). Motif identification analysis yielded several significant matches ( p < 0.05), such as GATA4, which is downregulated in CHD. Moreover, although the protein-protein interaction network showed FOG2/ZFPM2, FOS and ID2 proteins to be the key interacting partners of GATA4, only FOG2/ZFPM2 overexpression showed an interaction score of "high confidence" ≥ 0.84. A significant change in the MED was observed during HF. For the first time, the MED9 subunit was significantly reduced between familial DCM and HS ( p < 0.05), showing an increased MED9s isoform in DCM patients with respect to its full-length transcript. MED9 and GATA4 shared the same sequence motif and were involved in a network with FOG2/ZFPM2, FOS, and ID2, proteins already implicated in cardiac development.

Background: Lung cancer is the leading cause of cancer-related death worldwide. We previously found that Mediator complex subunit 23 (MED23) is important for the tumourigenicity of lung cancer cells with hyperactive Ras activity in vitro, although the in vivo function of MED23 in lung tumourigenesis remains to be explored., Methods: In this study, we utilized well-characterized Kras G12D -driven non-small cell lung cancer mouse model to investigate the role of MED23 in lung cancer. The lung tumour progression was evaluated by H&E and IHC analysis. Western blotting and qRT-PCR assays were performed to detect changes in gene expression. Immune cells were analyzed by FACS technology. RNA-seq and reporter assays were conducted to explore the mechanism., Results: We observed that lung epithelial Med23 deletion by adeno-Cre resulted in a significant increase in Kras G12D tumour number and size, which was further verified with another mouse model with Med23 specifically deleted in alveolar type II cells. Mice with lung-specific Med23 deficiency also exhibited accelerated tumourigenesis, and a higher proliferation rate for tumour cells, along with increased ERK phosphorylation. Notably, the numbers of infiltrating CD4 + T cells and CD8 + T cells were significantly reduced in the lungs of Med23-deficient mice, while the numbers of myeloid-derived suppressor cells (MDSCs) and Treg cells were significantly increased, suggesting the enhanced immune escape capability of the Med23-deficient lung tumours. Transcriptomic analysis revealed that the downregulated genes in Med23-deficient lung tumour tissues were associated with the immune response. Specifically, Med23 deficiency may compromise the MHC-I complex formation, partially through down-regulating B2m expression., Conclusions: Collectively, these findings revealed that MED23 may negatively regulate Kras-induced lung tumourigenesis in vivo, which would improve the precise classification of KRAS-mutant lung cancer patients and provide new insights for clinical interventions., (© 2024. The Author(s).)

Objectives: The MED12 gene encodes mediator complex subunit 12, which is a component of the mediator complex involved in the transcriptional regulation of nearly all RNA polymerase II-dependent genes. MED12 variants have previously been associated with developmental disorders with or without nonspecific intellectual disability. This study aims to explore the association between MED12 variants and epilepsy., Materials and Methods: Trios-based whole-exome sequencing was performed in a cohort of 349 unrelated cases with partial (focal) epilepsy without acquired causes. The genotype-phenotype correlations of MED12 variants were analyzed., Results: Five hemizygous missense MED12 variants, including c.958A>G/p.Ile320Val, c.1757G>A/p.Ser586Asn, c.2138C>T/p.Pro713Leu, c.3379T>C/p.Ser1127Pro, and c.4219A>C/p.Met1407Leu were identified in five unrelated males with partial epilepsy. All patients showed infrequent focal seizures and achieved seizure free without developmental abnormalities or intellectual disability. All the hemizygous variants were inherited from asymptomatic mothers (consistent with the X-linked recessive inheritance pattern) and were absent in the general population. The two variants with damaging hydrogen bonds were associated with early-onset seizures. Further genotype-phenotype analysis revealed that congenital anomaly disorder (Hardikar syndrome) was associated with (de novo) destructive variants in an X-linked dominant inheritance pattern, whereas epilepsy was associated with missense variants in an X-linked recessive inheritance pattern. Phenotypic features of intellectual disability appeared as the intermediate phenotype in terms of both genotype and inheritance. Epilepsy-related variants were located at the MED12-LCEWAV domain and the regions between MED12-LCEWAV and MED12-POL., Conclusion: MED12 is a potentially causative gene for X-linked recessive partial epilepsy without developmental or intellectual abnormalities. The genotype-phenotype correlation of MED12 variants explains the phenotypic variations and can help the genetic diagnosis., Competing Interests: Declaration of Competing Interest The authors have no conflicts of interest to declare., (Copyright © 2023 British Epilepsy Association. Published by Elsevier Ltd. All rights reserved.)