Functional insights from high resolution structures of mouse protein arginine methyltransferase 6.

PRMT6 is a protein arginine methyltransferase involved in transcriptional regulation, human immunodeficiency virus pathogenesis, DNA base excision repair, and cell cycle progression. Like other PRMTs, PRMT6 is overexpressed in several cancer types and is therefore considered as a potential anti-cancer drug target. In the present study, we described six crystal structures of PRMT6 from Mus musculus, solved and refined at 1.

Structural insight into arginine methylation by the mouse protein arginine methyltransferase 7: a zinc finger freezes the mimic of the dimeric state into a single active site.

Protein arginine methyltransferase 7 (PRMT7) is a type III arginine methyltransferase which has been implicated in several biological processes such as transcriptional regulation, DNA damage repair, RNA splicing, cell differentiation and metastasis. PRMT7 is a unique but less characterized member of the family of PRMTs. The crystal structure of full-length PRMT7 from Mus musculus refined at 1.

Molecular basis for the antiparasitic activity of a mercaptoacetamide derivative that inhibits histone deacetylase 8 (HDAC8) from the human pathogen schistosoma mansoni.

Schistosomiasis, caused by the parasitic flatworm Schistosoma mansoni and related species, is a tropical disease that affects over 200 million people worldwide. A new approach for targeting eukaryotic parasites is to tackle their dynamic epigenetic machinery that is necessary for the extensive phenotypic changes during the life cycle of the parasite. Recently, we identified S.

Cysteine mapping in conformationally distinct kinase nucleotide binding sites: application to the design of selective covalent inhibitors.

Kinases have emerged as one of the most prolific therapeutic targets. An important criterion in the therapeutic success of inhibitors targeting the nucleotide binding pocket of kinases is the inhibitor residence time. Recently, covalent kinase inhibitors have attracted attention since they confer terminal inhibition and should thus be more effective than reversible inhibitors with transient inhibition.

Defining and characterizing protein surface using alpha shapes.

The alpha shape of a molecule is a geometrical representation that provides a unique surface decomposition and a means to filter atomic contacts. We used it to revisit and unify the definition and computation of surface residues, contiguous patches, and curvature. These descriptors are evaluated and compared with former approaches on 85 proteins for which both bound and unbound forms are available.

Histone H3 tails containing dimethylated lysine and adjacent phosphorylated serine modifications adopt a specific conformation during mitosis and meiosis.

Condensation of chromatin, mediated in part by posttranslational modifications of histones, is essential for cell division during mitosis. Histone H3 tails are dimethylated on lysine (Kme2) and become phosphorylated on serine (Sp) residues during mitosis. We have explored the possibility that these double modifications are involved in the establishment of H3 tail conformations during the cell cycle.

Critical role of desolvation in the binding of 20-hydroxyecdysone to the ecdysone receptor.

The insect steroid hormone 20-hydroxyecdysone (20E) binds to its cognate nuclear receptor composed of the ecdysone receptor (EcR) and Ultraspiracle (USP) and triggers the main developmental transitions, in particular molting and metamorphosis. We present the crystal structure of the ligand-binding domains of EcR/USP in complex with 20E at 2.4A resolution and compare it with published structures of EcR/USP bound to ponasterone A (ponA).

The loss of transcriptional inhibition by the photoreceptor-cell specific nuclear receptor (NR2E3) is not a necessary cause of enhanced S-cone syndrome.

Purpose: To investigate functional consequence on photoreceptor-cell specific nuclear receptor (NR2E3) transcriptional activity of enhanced S-cone syndrome (ESCS) mutations localized in ligand binding domain (LBD).

Methods: Point mutations were introduced into the LBD of full length and Gal4 chimeric NR2E3 receptors and transcriptional activity was investigated by using transient co-transfection assay on corresponding luciferase reporters. Expression and DNA binding properties of transfected mutant and wild-type receptors were tested by Western blotting and gel shift assay.

Raloxifene and ICI182,780 increase estrogen receptor-alpha association with a nuclear compartment via overlapping sets of hydrophobic amino acids in activation function 2 helix 12.

The basis for the differential repressive effects of antiestrogens on transactivation by estrogen receptor-alpha (ERalpha) remains incompletely understood. Here, we show that the full antiestrogen ICI182,780 and, to a lesser extent, the selective ER modulator raloxifene (Ral), induce accumulation of exogenous ERalpha in a poorly soluble fraction in transiently transfected HepG2 or stably transfected MDA-MB231 cells and of endogenous receptor in MCF7 cells. ERalpha remained nuclear in HepG2 cells treated with either compound.

Partial defects in transcriptional activity of two novel DAX-1 mutations in childhood-onset adrenal hypoplasia congenita.

Objective: Mutations in DAX-1, an X-linked gene encoding an orphan nuclear receptor, have been associated with adrenal hypoplasia congenita and hypogonadotropic hypogonadism. Here we describe two novel DAX-1 mutations, Y214X and I361T, associated with childhood-onset primary adrenal failure. We aimed at analysing their effects on protein localization, transcriptional activity and propose a structural-function relationship.

The monomeric orphan nuclear receptor Schistosoma mansoni Ftz-F1 dimerizes specifically and functionally with the schistosome RXR homologue, SmRXR1.

In an attempt to understand development and differentiation processes of the parasitic blood fluke Schistosoma mansoni, several members of the nuclear receptor superfamily were cloned, including SmFtz-F1 (S. mansoni Fushi Tarazu-factor 1). The Ftz-F1 nuclear receptor subfamily only contains orphan receptors that bind to their response element as monomers.

Structure-based analysis of the ultraspiracle protein and docking studies of putative ligands.

The ultraspiracle protein (USP) is the insect ortholog of the mammalian retinoid X receptor (RXR). Fundamental questions concern the functional role of USP as the heterodimerization partner of insect nuclear receptors such as the ecdysone receptor. The crystallographic structures of the ligand binding domain of USPs of Heliothis virescens and Drosophila melanogaster solved recently show that helix 12 is locked in an antagonist conformation raising the question whether USPs could adopt an agonist conformation as observed in RXRalpha.

Identification of an alternative ligand-binding pocket in the nuclear vitamin D receptor and its functional importance in 1alpha,25(OH)2-vitamin D3 signaling.

Structural and molecular studies have shown that the vitamin D receptor (VDR) mediates 1alpha,25(OH)2-vitamin D3 gene transactivation. Recent evidence indicates that both VDR and the estrogen receptor are localized to plasma membrane caveolae and are required for initiation of nongenomic (NG) responses. Computer docking of the NG-specific 1alpha,25(OH)2-lumisterol to the VDR resulted in identification of an alternative ligand-binding pocket that partially overlaps the genomic pocket described in the experimentally determined x-ray structure.

Ataxin-7 is a subunit of GCN5 histone acetyltransferase-containing complexes.

Spinocerebellar ataxia type 7 (SCA7) is a neurodegenerative disorder caused by a CAG repeat expansion in the SCA7 gene leading to elongation of a polyglutamine tract in ataxin-7, a protein of unknown function. A putative ataxin-7 yeast orthologue (SGF73) has been identified recently as a new component of the SAGA (Spt/Ada/Gcn5 acetylase) multisubunit complex, a coactivator required for transcription of a subset of RNA polymerase II-dependent genes. We show here that ataxin-7 is an integral component of the mammalian SAGA-like complexes, the TATA-binding protein-free TAF-containing complex (TFTC) and the SPT3/TAF9/GCN5 acetyltransferase complex (STAGA).

Unique functional properties of a member of the Fushi Tarazu-Factor 1 family from Schistosoma mansoni.

SmFtz-F1 (Schistosoma mansoni Fushi Tarazu-Factor 1) belongs to the Ftz-F1 subfamily of nuclear receptors, but displays marked structural differences compared with its mammalian homologues SF-1 (steroidogenic factor-1) or liver receptor homologue-1. These include a long F domain (104 amino acids), an unusually large hinge region (133 amino acids) and a poorly conserved E-domain. Here, using Gal4 constructs and a mammalian two-hybrid assay, we have characterized the roles of these specific regions both in the transcriptional activity of the receptor and in its interactions with cofactors.

Crystal structures of the vitamin D nuclear receptor liganded with the vitamin D side chain analogues calcipotriol and seocalcitol, receptor agonists of clinical importance. Insights into a structural basis for the switching of calcipotriol to a receptor antagonist by further side chain modification.

The plethora of actions of 1alpha,25(OH)(2)D(3) in various systems suggested wide clinical applications of vitamin D nuclear receptor (VDR) ligands in treatments of inflammation, dermatological indication, osteoporosis, cancers, and autoimmune diseases. More than 3000 vitamin D analogues have been synthesized in order to reduce the calcemic side effects while maintaining the transactivation potency of these ligands. Here, we report the crystal structures of VDR ligand binding domain bound to two vitamin D agonists of therapeutical interest, calcipotriol and seocalcitol, which are characterized by their side chain modifications.

Structure-function analysis reveals the molecular determinants of the impaired biological function of DAX-1 mutants in AHC patients.

Mutations in the DAX-1 (NR0B1) gene cause the X-linked form of adrenal hypoplasia congenita (AHC), which is constantly found associated with hypogonadotropic hypogonadism (HHG). DAX-1 encodes an atypical orphan member of the nuclear hormone receptor superfamily. DAX-1 acts at multiple levels to repress the expression of genes involved in steroid hormone metabolism through a potent transcriptional repression domain present in its C-terminus, which is similar to the nuclear receptors' ligand binding domain.

Common characteristics of the human and rhesus macaque CD1e molecules: conservation of biochemical and biological properties during primate evolution.

In humans, a family of five genes encodes the CD1 molecules. Four of these proteins, CD1a, b, c, and d, are expressed on the plasma membrane and traffic between the cell surface and endocytic compartments, where they are loaded with antigenic glycolipids. The existence of human CD1e was demonstrated recently.

A new class of transcription initiation factors, intermediate between TATA box-binding proteins (TBPs) and TBP-like factors (TLFs), is present in the marine unicellular organism, the dinoflagellate Crypthecodinium cohnii.

Dinoflagellates are marine unicellular eukaryotes that exhibit unique features including a very low level of basic proteins bound to the chromatin and the complete absence of histones and nucleosomal structure. A cDNA encoding a protein with a strong homology to the TATA box-binding proteins (TBP) has been isolated from an expressed sequence tag library of the dinoflagellate Crypthecodinium cohnii. The typical TBP repeat signature and the amino acid motives involved in TFIIA and TFIIB interactions were conserved in this new TBP-like protein.

Structural and functional evidence for ligand-independent transcriptional activation by the estrogen-related receptor 3.

The crystal structure of the ligand binding domain (LBD) of the estrogen-related receptor 3 (ERR3) complexed with a steroid receptor coactivator-1 (SRC-1) peptide reveals a transcriptionally active conformation in absence of any ligand. The structure explains why estradiol does not bind ERRs with significant affinity. Docking of the previously reported ERR antagonists, diethylstilbestrol and 4-hydroxytamoxifen, requires structural rearrangements enlarging the ligand binding pocket that can only be accommodated with an antagonist LBD conformation.