Adenovirus E1A Activation Domain Regulates H3 Acetylation Affecting Varied Steps in Transcription at Different Viral Promoters.

How histone acetylation promotes transcription is not clearly understood. Here, we confirm an interaction between p300 and the adenovirus 2 large E1A activation domain (AD) and map the interacting regions in E1A by observing colocalization at an integrated array of fusions of LacI-mCherry to E1A fragments with YFP-p300. Viruses with mutations in E1A subdomains were constructed and analyzed for kinetics of early viral RNA expression and association of acetylated H3K9, K18, K27, TBP, and RNA polymerase II (Pol II) across the viral genome.

Tandem SAM domain structure of human Caskin1: a presynaptic, self-assembling scaffold for CASK.

The synaptic scaffolding proteins CASK and Caskin1 are part of the fibrous mesh of proteins that organize the active zones of neural synapses. CASK binds to a region of Caskin1 called the CASK interaction domain (CID). Adjacent to the CID, Caskin1 contains two tandem sterile α motif (SAM) domains.

Adenovirus E1B 55-kilodalton protein is a p53-SUMO1 E3 ligase that represses p53 and stimulates its nuclear export through interactions with promyelocytic leukemia nuclear bodies.

Oncogenic transformation by adenovirus E1A and E1B-55K requires E1B-55K inhibition of p53 activity to prevent E1A-induced apoptosis. During viral infection, E1B-55K and E4orf6 substitute for the substrate-binding subunits of the host cell cullin 5 class of ubiquitin ligases, resulting in p53 polyubiquitinylation and proteasomal degradation. Here we show that E1B-55K alone also functions as an E3 SUMO1-p53 ligase.

Complexity in transcription control at the activation domain-mediator interface.

Transcript elongation by polymerase II paused at the Egr1 promoter is activated by mitogen-activated protein kinase phosphorylation of the ternary complex factor (TCF) ELK1 bound at multiple upstream sites and subsequent phospho-ELK1 interaction with mediator through the MED23 subunit. Consequently, Med23 knockout (KO) nearly eliminates Egr1 (early growth response factor 1) transcription in embryonic stem (ES) cells, leaving a paused polymerase at the promoter. Med23 KO did not, however, eliminate Egr1 transcription in fibroblasts.

Resonance assignments of the metal sensor CzrA in the apo-, Zn2- and DNA-bound (42 kDa) states.

Zn/Co binding to homodimeric CzrA de-represses transcription of the czr (cobalt zinc resistance) operon in Staphylococcus aureus. Essentially complete backbone and stereospecific methyl group resonance assignments for apo-, Zn2 and a 42 kDa apo-CzrA-DNA complex are presented.

Individual metal ligands play distinct functional roles in the zinc sensor Staphylococcus aureus CzrA.

Recent studies on metalloregulatory proteins suggest that coordination number/geometry and metal ion availability in a host cytosol are key determinants for biological specificity. Here, we investigate the contribution that individual metal ligands of the alpha5 sensing site of Staphylococcus aureus CzrA (Asp84, His86, His97', and His100') make to in vitro metal ion binding affinity, coordination geometry, and allosteric negative regulation of DNA operator/promoter region binding. All ligand substitution mutants exhibit significantly reduced metal ion binding affinity (K(Me)) by > or =10(3) M(-1).

Structural determinants of metal selectivity in prokaryotic metal-responsive transcriptional regulators.

Metal ion homeostasis in prokaryotes is maintained by metal-responsive transcriptional regulatory proteins that regulate the transcription of genes encoding proteins responsible for metal detoxification, sequestration, efflux and uptake. These metalloregulatory, or metal sensor proteins, bind a wide range of specific metal ions directly; this in turn, allosterically regulates (enhances or decreases) operator/promoter binding affinity or promoter structure. Recent structural studies reveal five distinct families of metal sensor proteins.

A metal-ligand-mediated intersubunit allosteric switch in related SmtB/ArsR zinc sensor proteins.

The origin of metal ion selectivity by members of the SmtB/ArsR family of bacterial metal-sensing transcriptional repressors and the mechanism of negative allosteric regulation of DNA binding is poorly understood. Here, we report that two homologous zinc sensors, Staphylococcus aureus CzrA and cyanobacterial SmtB, are "winged" helix homodimeric DNA-binding proteins that bind Zn(II) to a pair of tetrahedral, interhelical binding sites, with two ligands derived from the alpha5 helix of one subunit, Asp84 O(delta1) (Asp104 in SmtB), His86 N(delta1) (His106), and two derived from the alpha5 helix of the other, His97' N(delta1) (His117') and His100' N(epsilon2) (Glu120'). Formation of the metal chelate drives a quaternary structural switch mediated by an intersubunit hydrogen-binding network that originates with the non-liganding N(epsilon2) face of His97 in CzrA (His117 in SmtB) that stabilizes a low-affinity, DNA-binding conformation.

The SmtB/ArsR family of metalloregulatory transcriptional repressors: Structural insights into prokaryotic metal resistance.

The SmtB/ArsR family of prokaryotic metalloregulatory transcriptional repressors represses the expression of operons linked to stress-inducing concentrations of di- and multivalent heavy metal ions. Derepression results from direct binding of metal ions by these homodimeric "metal sensor" proteins. An evolutionary analysis, coupled with comparative structural and spectroscopic studies of six SmtB/ArsR family members, suggests a unifying "theme and variations" model, in which individual members have evolved distinct metal selectivity profiles by alteration of one or both of two structurally distinct metal coordination sites.

Structural elements of metal selectivity in metal sensor proteins.

Staphylococcus aureus CzrA and Mycobacterium tuberculosis NmtR are homologous zinccobalt-responsive and nickelcobalt-responsive transcriptional repressors in vivo, respectively, and members of the ArsRSmtB superfamily of prokaryotic metal sensor proteins. We show here that Zn(II) is the most potent negative allosteric regulator of czr operatorpromoter binding in vitro with the trend Zn(II)>Co(II)Ni(II), whereas the opposite holds for the binding of NmtR to the nmt operatorpromoter, Ni(II)>Co(II)>Zn(II). Characterization of the metal coordination complexes of CzrA and NmtR by UVvisible and x-ray absorption spectroscopies reveals that metals that form four-coordinate tetrahedral complexes with CzrA [Zn(II) and Co(II)] are potent regulators of DNA binding, whereas metals that form five- or six-coordinate complexes with NmtR [Ni(II) and Co(II)] are the strongest allosteric regulators in this system.

A nickel-cobalt-sensing ArsR-SmtB family repressor. Contributions of cytosol and effector binding sites to metal selectivity.

NmtR from Mycobacterium tuberculosis is a new member of the ArsR-SmtB family of metal sensor transcriptional repressors. NmtR binds to the operator-promoter of a gene encoding a P(1) type ATPase (NmtA), repressing transcription in vivo except in medium supplemented with nickel or, to some extent, cobalt. In a cyanobacterial host, Synechococcus PCC 7942 strain R2-PIM8(smt), NmtR-mediated repression is alleviated by cobalt but not nickel or zinc addition, while the related sensor SmtB responds exclusively to zinc.