Social Media and Financial Market (<Special Issue>Theory and Application of Econometrics)

textabstractAll members of the steroid hormone receptor family are phosphoproteins. Additional phosphorylation occurs in the presence of hormone. This hormone-induced phosphorylation, which is 2- to 7-fold more than the basal phosphorylation, is a rapid process. All steroid receptors are phosphorylated at more than one single site. Most phosphorylation sites are located in the N-terminal domain, and phosphorylation occurs mainly on serine residues. Phosphorylation on threonine residues occurs in only a few cases. Phosphorylation on tyrosine residues has been found only for the estrogen receptor. Six different protein kinases are possibly involved in steroid receptor phosphorylation (estrogen receptor kinase; protein kinase A; protein kinase C; casein kinase II; DNA-dependent kinase; Ser-Pro kinases). Steroid receptor phosphorylation has been directly implicated in: activation of hormone binding, nuclear import of steroid receptors, modulation of binding to hormone response elements, and consequently in transcription activation

Het nieuwe betavak Natuur, Leven en Techniek: waarom, hoe en wanneer?

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Substitution of cysteine for selenocysteine in the catalytic center of type III iodothyronine deiodinase reduces catalytic efficiency and alters substrate preference

Human type III iodothyronine deiodinase (D3) catalyzes the conversion of T(4) to rT(3) and of T(3) to 3, 3'-diiodothyronine (T2) by inner-ring deiodination. Like types I and II iodothyronine deiodinases, D3 protein contains selenocysteine (SeC) in the highly conserved core catalytic center at amino acid position 144. To evaluate the contribution of SeC144 to the catalytic properties of D3 enzyme, we generated mutants in which cysteine (D3Cys) or alanine (D3Ala) replaces SeC144 (D3wt). COS cells were transfected with expression vectors encoding D3wt, D3Cys, or D3Ala protein. Kinetic analysis was performed on homogenates with dithiothreitol as reducing cofactor. The Michaelis constant of T(3) was 5-fold higher for D3Cys than for D3wt protein. In contrast, the Michaelis constant of T(4) increased 100-fold. The D3Ala protein was enzymatically inactive. Semiquantitative immunoblotting of homogenates with a D3 antiserum revealed that about 50-fold higher amounts of D3Cys and D3Ala protein are expressed relative to D3wt protein. The relative substrate turnover number of D3Cys is 2-fold reduced for T(3) and 6-fold reduced for T(4) deiodination, compared with D3wt enzyme. Studies in intact COS cells expressing D3wt or D3Cys showed that the D3Cys enzyme is also active under in situ conditions. In conclusion, the SeC residue in the catalytic center of D3 is essential for efficient inner-ring deiodination of T(3) and in particular T(4) at physiological substrate concentrations

Androgen receptor phosphorylation

Many physiological processes in organisms are regulated by a relatively small number of steroid honnones. Androgens are the so-called male sex steroid hormones which control growth, differentiation and functions of male reproductive and accessory sex tissues. Androgens are mainly produced in the testis and circulate in the blood. They diffuse in and out of all cells, but are retained with high affinity and specificity in target cells by an intranuclear binding protein, termed the androgen receptor. Once bound by androgens, the androgen receptor undergoes a conformational change allowing the receptor to bind with high affinity to DNA and to modulate transcription of certain genes. The androgen receptor appears to be a transcription factor, regulated by androgenic steroids. Phosphorylation is the predominant cellular mechanism for reversible modification of proteins, and the fact that many transcription factors are phosphoproteins suggests a regulatory role of phosphorylation. In this thesis, studies on phosphorylation of the androgen receptor in human prostate tumor cells (LNCaP) are described. In LNCaP cells, the androgen receptor protein is present as two isofonlls with apparent molecular masses of 110 and 112 kDa during SDS-PAGE. The 112 kDa isoform reflects the phosphorylated receptor, whereas the 110 kDa isoform is the non-phosphorylated receptor. Both isoforms are able to bind androgens with high affinity and can subsequently be transformed to the DNA binding form. It appears to be unlikely that phosphorylation is involved in the regulation of steroid- or DNA binding affinity. Upon incubation of the prostate tumor cells with androgens, the phosphOlylation degree of the androgen receptor was rapidly increased. Multiple phosphorylation sites on serine residues are located in the N-terminal f/'al/~activation domain and not in the DNA- and ligand binding domains. Tryptic phosphopeptide maps of the androgen receptor show induction of phosphorylation at a novel site(s) by hormone treatment. It is proposed that this extra phosphorylation in the N-terminal domain causes a conformational change, enabling protein-protein contacts of the trails-activation domain with other transcription factors or co-activators on a target gene promoter

Substitution of cysteine for a conserved alanine residue in the catalytic center of type II iodothyronine deiodinase alters interaction with reducing cofactor

Human type II iodothyronine deiodinase (D2) catalyzes the activation of T(4) to T(3). The D2 enzyme, like the type I (D1) and type III (D3) deiodinases, contains a selenocysteine (SeC) residue (residue 133 in D2) in the highly conserved catalytic center. Remarkably, all of the D2 proteins cloned so far have an alanine two residue-amino terminal to the SeC, whereas all D1 and D3 proteins contain a cysteine at this position. A cysteine residue in the catalytic center could assist in enzymatic action by providing a nucleophilic sulfide or by participating in redox reactions with a cofactor or enzyme residues. We have investigated whether D2 mutants with a cysteine (A131C) or serine (A131S) two-residue amino terminal to the SeC are enzymatically active and have characterized these mutants with regard to substrate affinity, reducing cofactor interaction and inhibitor profile. COS cells were transfected with expression vectors encoding wild-type (wt) D2, D2 A131C, or D2 A131S proteins. Kinetic analysis was performed on homogenates with dithiothreitol (DTT) as reducing cofactor. The D2 A131C and A131S mutants displayed similar Michaelis-Menten constant values for T(4) (5 nM) and reverse T(3) (9 nM) as the wt D2 enzyme. The limiting Michaelis-Menten constant for DTT of the D2 A131C enzyme was 3-fold lower than that of the wt D2 enzyme. The wt and mutant D2 enzymes are essentially insensitive to propylthiouracil [concentration inhibiting 50% of activity (IC(50)) > 2 mM] in the presence of 20 mM DTT, but when tested in the presence of 0.2 mM DTT the IC(50) value for propylthiouracil is reduced to about 0.1 mM. During incubations of intact COS cells expressing wt D2, D2 A131C, or D2 A131S, addition of increasing amounts of unlabeled T(4) resulted in the saturation of [(125)I]T(4) deiodination, as reflected in a decrease of [(125)I]T(3) release into the medium. Saturation first appeared at medium T(4) concentrations between 1 and 10 nM. In conclusion: substitution of cysteine for a conserved alanine residue in the catalytic center of the D2 protein does not inactivate the enzyme in vitro and in situ, but rather improves the interaction with the reducing cofactor DTT in vitro

PyPedia:using the wiki paradigm as crowd sourcing environment for bioinformatics protocols

Background: Today researchers can choose from many bioinformatics protocols for all types of life sciences research, computational environments and coding languages. Although the majority of these are open source, few of them possess all virtues to maximize reuse and promote reproducible science. Wikipedia has proven a great tool to disseminate information and enhance collaboration between users with varying expertise and background to author qualitative content via crowdsourcing. However, it remains an open question whether the wiki paradigm can be applied to bioinformatics protocols. Results: We piloted PyPedia, a wiki where each article is both implementation and documentation of a bioinformatics computational protocol in the python language. Hyperlinks within the wiki can be used to compose complex workflows and induce reuse. A RESTful API enables code execution outside the wiki. Initial content of PyPedia contains articles for population statistics, bioinformatics format conversions and genotype imputation. Use of the easy to learn wiki syntax effectively lowers the barriers to bring expert programmers and less computer savvy researchers on the same page. Conclusions: PyPedia demonstrates how wiki can provide a collaborative development, sharing and even execution environment for biologists and bioinformaticians that complement existing resources, useful for local and multi-center research teams. Availability: PyPedia is available online at: http://www.pypedia.com. The source code and installation instructions are available at: https://github.com/kantale/PyPedia_server. The PyPedia python library is available at: https://github.com/kantale/pypedia. PyPedia is open-source, available under the BSD 2-Clause License

Inclination Effects and Beaming in Black Hole X-ray Binaries

We investigate the dependence of observational properties of black hole X-ray binaries on the inclination angle i of their orbits. We find the following: (1) Transient black hole binaries show no trend in their quiescent X-ray luminosities as a function of i, suggesting that the radiation is not significantly beamed. This is consistent with emission from an accretion disk. If the X-rays are from a jet, then the Lorentz factor gamma of the jet is less than 1.24 at the 90% confidence level. (2) The X-ray binary 4U1543-47 with i of order 21 degrees has a surprisingly strong fluorescent iron line in the high soft state. Quantifying an earlier argument by Park et al. (2004), we conclude that if the continuum X-ray emission in this source is from a jet, then gamma < 1.04. (3) None of the known binaries has cos i 75 degrees. This fact, plus the lack of eclipses among the 20 black hole binaries in our sample, strongly suggests at the 99.5% confidence level that systems with large inclination angles are hidden from view. The obscuration could be the result of disk flaring, as suggested by Milgrom (1978) for neutron star X-ray binaries. (4) Transient black hole binaries with i ~ 70-75 degrees have significantly more complex X-ray light curves than systems with i < 65 degrees. This may be the result of variable obscuration and/or variable height above the disk of the radiating gas.Comment: 26 pages, to appear in The Astrophysical Journal, vol. 624, May 1, 200

The androgen receptor: Functional structure and expression in transplanted human prostate tumors and prostate tumor cell lines

Abstract The growth of the majority of prostate tumors is androgen-dependent, for which the presence of a functional androgen receptor is a prerequisite. Tumor growth can be inhibited by blockade of androgen receptor action. However, this inhibition is transient. To study the role of the androgen receptor in androgen-dependent and androgen-independent prostate tumor cell growth, androgen receptor mRNA expression was monitored in six different human prostate tumor cell lines and tumors, which were grown either in vitro or by transplantation on (male) nude mice. Androgen receptor mRNA was clearly detectable in three androgen-dependent (sensitive) tumors and absent or low in three androgen-independent tumors. Growth of the LNCaP prostate tumor cell line can be stimulated both by androgens and by fetal calf serum. In the former situation androgen receptor mRNA expression is downregulated, whereas in the latter no effect on androgen receptor mRNA levels can be demonstrated. Sequence analysis showed that the androgen receptor gene from LNCaP cells contains a point mutation in the region encoding the steroid-binding domain, which confers an ACT coVon encoding a threonine residue to GCT, encoding alanine

Androgen receptor abnormalities

The human androgen receptor is a member of the superfamily of steroid hormone receptors. Proper functioning of this protein is a prerequisite for normal male sexual differentiation and development. The cloning of the human androgen receptor cDNA and the elucidation of the genomic organization of the corresponding gene has enabled us to study androgen receptors in subjects with the clinical manifestation of androgen insensitivity and in a human prostate carcinoma cell line (LNCaP). Using PCR amplification, subcloning and sequencing of exons 2–8, we identified a G → T mutation in the androgen receptor gene of a subject with the complete form of androgen insensitivity, which inactivates the splice donor site at the exon 4/intron 4 boundary. This mutation causes the inactivation of a cryptic splice donor site in exon 4, which results in the deletion of 41 amino acids from the steroid binding domain. In two other independently arising cases we identified two different nucleotide alterations in codon 686 (GAC; aspartic acid) located in exon 4. One mutation (G → C) results in an aspartic acid → histidine substitution (with negligible androgen binding), whereas the other mutation (G → A) leads to an aspartic acid → asparagine substitution (normal androgen binding, but a rapidly dissociating androgen receptor complex). Sequence analysis of the androgen receptor in human LNCaP-cells (lymph node carcinoma of the prostate) revealed a point mutation (A → G) in codon 868 in exon 8 resulting in the substitution of threonine by alanine. This mutation is the cause of the altered steroid binding specificity of the LNCaP-cell androgen receptor. The functional consequences of the observed mutations with respect to protein expression, specific ligand binding and transcriptional activation, were established after transient expression of the mutant receptors in COS and HeLa cells. These findings illustrate that functional error

Distinct modulation of cellular immunopeptidome by the allosteric regulatory site of ER aminopeptidase 1

ER aminopeptidase 1 (ERAP1) is an ER-resident aminopeptidase that excises N-terminal residues of peptides that then bind onto Major Histocompatibility Complex I molecules (MHC-I) and indirectly modulates adaptive immune responses. ERAP1 contains an allosteric regulatory site that accommodates the C-terminus of at least some peptide substrates, raising questions about its exact influence on antigen presentation and the potential of allosteric inhibition for cancer immunotherapy. We used an inhibitor that targets this regulatory site to study its effect on the immunopeptidome of a human cancer cell line. The immunopeptidomes of allosterically inhibited and ERAP1 KO cells contain high-affinity peptides with sequence motifs consistent with the cellular HLA class I haplotypes but are strikingly different in peptide composition. Compared to KO cells, allosteric inhibition did not affect the length distribution of peptides and skewed the peptide repertoire both in terms of sequence motifs and HLA allele utilization, indicating significant mechanistic differences between the two ways of disrupting ERAP1 function. These findings suggest that the regulatory site of ERAP1 plays distinct roles in antigenic peptide selection, which should be taken into consideration when designing therapeutic interventions targeting the cancer immunopeptidome