Pseudodominant Inheritance of Goitrous Congenital Hypothyroidism Caused by TPO Mutations: Molecular and in Silico Studies

Context and Objective: Most cases of goitrous congenital hypothyroidism (CH) from thyroid dyshormonogenesis 1) follow a recessive mode of inheritance and 2) are due to mutations in the thyroid peroxidase gene (TPO). We report the genetic mechanism underlying the apparently dominant inheritance of goitrous CH in a nonconsanguineous family of French Canadian origin. Design, Setting, and Participants: Two brothers identified by newborn TSH screening had severe hypothyroidism and a goiter with increased (99m)Tc uptake. The mother was euthyroid, but the father and two paternal uncles had also been diagnosed with goitrous CH. After having excluded PAX8 gene mutations, we hypothesized that the underlying defect could be TPO mutations. Results: Both compound heterozygous siblings had inherited a mutant TPO allele carried by their mother (c.1496delC; p.Pro499Argfs2X), and from their father, one brother had inherited a missense mutation (c.1978C-->G; p.Gln660Glu) and the other an insertion (c.1955insT; p.Phe653Valfs15X). The thyroid gland of one uncle who is a compound heterozygote for TPO mutations (p.Phe653Valfs15X/p.Gln660Glu) was removed because of concurrent multiple endocrine neoplasia type 2A. Immunohistochemistry revealed normal TPO staining, implying that Gln660Glu TPO is expressed properly. Modeling of this mutant in silico suggests that its three-dimensional structure is conserved, whereas the electrostatic binding energy between the Gln660Glu TPO and its heme group becomes repulsive. Conclusion: We report a pedigree presenting with pseudodominant goitrous CH due to segregation of three different TPO mutations. Although goitrous CH generally follows a recessive mode of inheritance, the high frequency of TPO mutations carriers may lead to pseudodominant inheritance.

Clinical, structural and functional implications of mutations and polymorphisms in human NADPH P450 oxidoreductase

Cytochrome P450 proteins are involved in metabolism of drugs and xenobiotics. In the endoplasmic reticulum a single nicotinamide adenine dinucleotide phosphate (NADPH) P450 oxidoreductase (POR) supplies electrons to all microsomal P450s for catalytic activity. POR is a flavoprotein that contains both flavin mononucleotide and flavin adenine dinucleotide as cofactors and uses NADPH as the source of electrons. We have recently reported a number of POR mutations in the patients with disordered steroidogenesis. In the first report we had described missense mutations (A287P, R457H, V492E, C569Y, and V608F) identified in four patients with defects in steroid production. Each POR variant was produced as recombinant N-27 form of the enzyme in bacteria and as full-length form in yeast. Membranes from bacteria or yeast expressing normal or variant POR were purified and their activities were characterized in cytochrome c and CYP17A1 assays. Later we have published a larger study that described a whole range of POR mutations and characterized the mutants/polymorphisms A115V, T142A, M263V, Y459H, A503V, G539R, L565P, R616X, V631I, and F646del from the sequencing of patient DNA. We also studied POR variants Y181D, P228L, R316W, G413S, and G504R that were available in public databases or published literature. Three-dimensional structure of rat POR is known and we have used this structure to deduce the structure-function correlation of POR mutations in human. The missense mutations found in patients with disordered steroidogenesis are generally in the co-factor binding and functionally important domains of POR and the apparent polymorphisms are found in regions with lesser structural importance. A variation in POR can alter the activity of all microsomal P450s, and therefore, can affect the metabolism of drugs and xenobiotics even when the P450s involved are otherwise normal. It is important to study the genetic and biochemical basis of POR variants in human population to gain information about possible differences in P450 mediated reactions among the individuals carrying a variant or polymorphic form of POR that could impact their metabolism.

Lifshitz holographic renormalization from anti-de Sitter 1

Lifshitz space–times with critical exponent z = 2 can be obtained by dimensional reduction of Schrödinger space–times with critical exponent z = 0. The latter space–times are asymptotically anti-de Sitter (AdS) solutions of AdS gravity coupled to an axion–dilaton system (or even just a massless scalar field). This basic observation is used to perform holographic renormalization for four-dimensional asymptotically locally Lifshitz space–times by dimensional reduction of the corresponding problem of holographic renormalization for five-dimensional asymptotically AdS space–times coupled to an axion–dilaton system. In this setup the four-dimensional structure of the Lifshitz – Fefferman-Graham expansion and the structure of the counterterm action, including the scale anomaly, will be summarized.

Autosomal dominant neurohypophyseal diabetes insipidus in a Swiss family, caused by a novel mutation (C59Delta/A60W) in the neurophysin moiety of prepro-vasopressin-neurophysin II (AVP-NP II).

OBJECTIVE To study clinical, morphological and molecular characteristics in a Swiss family with autosomal dominant familial neurohypophyseal diabetes insipidus (adFNDI). PARTICIPANTS AND METHODS A 15-month-old girl presenting with symptoms of polydipsia and polyuria was investigated by water deprivation test. Evaluation of the family revealed three further family members with symptomatic vasopressin-deficient diabetes insipidus. T1-weighted magnetic resonance images of the posterior pituitary were taken in two affected adult family members and molecular genetic analysis was performed in all affected individuals. RESULTS The water deprivation test in the 15-month-old child confirmed the diagnosis of vasopressin-deficient diabetes insipidus and the pedigree was consistent with autosomal dominant inheritance. The characteristic bright spot of the normal vasopressin-containing neurophypophysis was absent in both adults with adFNDI. Direct sequence analysis revealed a new deletion (177-179DeltaCGC) in exon 2 of the AVP-NP II gene in all affected individuals. At the amino acid level, this deletion eliminates cysteine 59 (C59Delta) and substitutes alanine 60 by tryptophan (A60W) in the AVP-NP II precursor; interestingly, the remainder of the reading frame remains unchanged. According to the three-dimensional structure of neurophysin, C59 is involved in a disulphide bond with C65. CONCLUSIONS Deletion of C59 and substitution of A60W in the AVP-NP II precursor is predicted to disrupt one of the seven disulphide bridges required for correct folding of the neurophysin moiety and thus disturb the function of neurophysin as the vasopressin transport protein. These data are in line with the clinical and morphological findings in the reported family with adFNDI.

Quantitative 3D strain analysis in analogue experiments: Integration of X-ray computed tomography and digital volume correlation techniques

The combination of scaled analogue experiments, material mechanics, X-ray computed tomography (XRCT) and Digital Volume Correlation techniques (DVC) is a powerful new tool not only to examine the 3 dimensional structure and kinematic evolution of complex deformation structures in scaled analogue experiments, but also to fully quantify their spatial strain distribution and complete strain history. Digital image correlation (DIC) is an important advance in quantitative physical modelling and helps to understand non-linear deformation processes. Optical non-intrusive (DIC) techniques enable the quantification of localised and distributed deformation in analogue experiments based either on images taken through transparent sidewalls (2D DIC) or on surface views (3D DIC). X-ray computed tomography (XRCT) analysis permits the non-destructive visualisation of the internal structure and kinematic evolution of scaled analogue experiments simulating tectonic evolution of complex geological structures. The combination of XRCT sectional image data of analogue experiments with 2D DIC only allows quantification of 2D displacement and strain components in section direction. This completely omits the potential of CT experiments for full 3D strain analysis of complex, non-cylindrical deformation structures. In this study, we apply digital volume correlation (DVC) techniques on XRCT scan data of “solid” analogue experiments to fully quantify the internal displacement and strain in 3 dimensions over time. Our first results indicate that the application of DVC techniques on XRCT volume data can successfully be used to quantify the 3D spatial and temporal strain patterns inside analogue experiments. We demonstrate the potential of combining DVC techniques and XRCT volume imaging for 3D strain analysis of a contractional experiment simulating the development of a non-cylindrical pop-up structure. Furthermore, we discuss various options for optimisation of granular materials, pattern generation, and data acquisition for increased resolution and accuracy of the strain results. Three-dimensional strain analysis of analogue models is of particular interest for geological and seismic interpretations of complex, non-cylindrical geological structures. The volume strain data enable the analysis of the large-scale and small-scale strain history of geological structures.

Interaction of the Trinuclear Triangular Secondary Building Unit [Cu 3 (μ 3 -OH)(μ-pz) 3 ] 2+ with 4,4′-Bipyridine. Structural Characterizations of New Coordination Polymers and Hexanuclear Cu II Clusters. 2°

By reacting 4,4′-bipyridine (bpy) with selected trinuclear triangular CuII complexes, [Cu3(μ3-OH)(μ-pz)3(RCOO)2(LL′)] [pz = pyrazolate anion; R = CH3, CH3CH2, CH2═CH, CH2═C(CH3); L, L′ = Hpz, H2O, MeOH] in MeOH, the substitution of monotopic ligands by ditopic bpy was observed. Depending on the stoichiometric reaction ratios, different compounds were isolated and structurally characterized. One- and two-dimensional coordination polymers (CPs), as well as two hexanuclear CuII clusters were identified. One of the hexanuclear clusters self-assembles into a supramolecular three-dimensional structure, and its crystal packing shows the presence of two intersecting channels, one of which is almost completely occupied by guest bpy, while in the second one guest water molecules are present. This compound also shows a reversible, thermally induced, single-crystal-to-single-crystal transition.

Mikro- versus Makroperspektive der retributiven Gerechtigkeit, Strafziele und die Forderung nach Strafe

Wie Personen auf Straftaten reagieren und welche Strafziele sie dabei verfolgen, war bisher kaum Gegenstand psychologischer Forschung. In der vorliegenden Arbeit wird die dimensionale Struktur der Präferenz von Strafzielen untersucht. In zwei Befragungen wurden juristischen Laien Fallgeschichten zur Beurteilung vorgelegt (Raubüberfall, Körperverletzung, Betrug, Vergewaltigung). Die multivariaten Auswertungen weisen eine hohe Übereinstimmung auf: Strafziele lassen sich durch die zwei voneinander unabhängigen Dimensionen Strafhärte und Mikro- versus Makroperspektive klassifizieren. Wird eine Makroperspektive eingenommen, so ist dies mit einer stärkeren Gewichtung von Gesellschaftsinteressen verbunden sowie mit der Präferenz für das Strafziel der positiven Generalprävention. Die Mikroperspektive ist hingegen mit der Überzeugung verbunden, dass Gerechtigkeit insbesondere aus der Perspektive der konkret Beteiligten herzustellen ist. Je nach geforderter Strafhärte geht die Mikroperspektive mit einem Bias entweder für das Opfer und das Strafziel der Vergeltung oder aber für den Täter und das Strafziel der Resozialisierung einher.

Luminal particles within cellular microtubules.

The regulation of microtubule dynamics is attributed to microtubule-associated proteins that bind to the microtubule outer surface, but little is known about cellular components that may associate with the internal side of microtubules. We used cryoelectron tomography to investigate in a quantitative manner the three dimensional structure of microtubules in intact mammalian cells. We show that the lumen of microtubules in this native state is filled with discrete, globular particles with a diameter of 7 nm and spacings between 8 and 20 nm in neuronal cells. Cross-sectional views of microtubules confirm the presence of luminal material in vitreous sections of brain tissue. Most of the luminal particles had connections to the microtubule wall, as revealed in tomograms. A higher accumulation of particles was seen near the retracting plus ends of microtubules. The luminal particles were abundant in neurons, but were also observed in other cells, such as astrocytes and stem cells.

Neospora caninum calcium-dependent protein kinase 1 is an effective drug target for neosporosis therapy.

Despite the enormous economic importance of Neospora caninum related veterinary diseases, the number of effective therapeutic agents is relatively small. Development of new therapeutic strategies to combat the economic impact of neosporosis remains an important scientific endeavor. This study demonstrates molecular, structural and phenotypic evidence that N. caninum calcium-dependent protein kinase 1 (NcCDPK1) is a promising molecular target for neosporosis drug development. Recombinant NcCDPK1 was expressed, purified and screened against a select group of bumped kinase inhibitors (BKIs) previously shown to have low IC50s against Toxoplasma gondii CDPK1 and T. gondii tachyzoites. NcCDPK1 was inhibited by low concentrations of BKIs. The three-dimensional structure of NcCDPK1 in complex with BKIs was studied crystallographically. The BKI-NcCDPK1 structures demonstrated the structural basis for potency and selectivity. Calcium-dependent conformational changes in solution as characterized by small-angle X-ray scattering are consistent with previous structures in low Calcium-state but different in the Calcium-bound active state than predicted by X-ray crystallography. BKIs effectively inhibited N. caninum tachyzoite proliferation in vitro. Electron microscopic analysis of N. caninum cells revealed ultra-structural changes in the presence of BKI compound 1294. BKI compound 1294 interfered with an early step in Neospora tachyzoite host cell invasion and egress. Prolonged incubation in the presence of 1294 interfered produced observable interference with viability and replication. Oral dosing of BKI compound 1294 at 50 mg/kg for 5 days in established murine neosporosis resulted in a 10-fold reduced cerebral parasite burden compared to untreated control. Further experiments are needed to determine the PK, optimal dosage, and duration for effective treatment in cattle and dogs, but these data demonstrate proof-of-concept for BKIs, and 1294 specifically, for therapy of bovine and canine neosporosis.

Novel sst2-selective somatostatin agonists. Three-dimensional consensus structure by NMR

The 3D NMR structures of six octapeptide agonist analogues of somatostatin (SRIF) in the free form are described. These analogues, with the basic sequence H-DPhe/Phe2-c[Cys3-Xxx7-DTrp8-Lys9-Thr10-Cys14]-Thr-NH2 (the numbering refers to the position in native SRIF), with Xxx7 being Ala/Aph, exhibit potent and highly selective binding to human SRIF type 2 (sst2) receptors. The backbone of these sst2-selective analogues have the usual type-II' beta-turn reported in the literature for sst2/3/5-subtype-selective analogues. Correlating the biological results and NMR studies led to the identification of the side chains of DPhe2, DTrp8, and Lys9 as the necessary components of the sst2 pharmacophore. This is the first study to show that the aromatic ring at position 7 (Phe7) is not critical for sst2 binding and that it plays an important role in sst3 and sst5 binding. This pharmacophore is, therefore, different from that proposed by others for sst2/3/5 analogues.

Three-dimensional consensus structure of sst2-selective somatostatin (SRIF) antagonists by NMR

The three-dimensional NMR structures of seven octapeptide analogs of somatostatin (SRIF), based on octreotide, with the basic sequence H-Cpa/Phe2-c[DCys3-Xxx7-DTrp/DAph(Cbm)8-Lys9-Thr10-Cys14]-Yyy-NH2 (the numbering refers to the position in native SRIF), with Xxx7 being Aph(Cbm)/Tyr/Agl(NMe,benzoyl) and Yyy being Nal/DTyr/Thr, are presented here. Most of these analogs exhibit potent and highly selective binding to sst2 receptors, and all of the analogs are antagonists inhibiting receptor signaling. Based on their consensus 3D structure, the pharmacophore of the sst2-selective antagonist has been defined. The pharmacophore involves the side chains of Cpa2, DTrp/DAph(Cbm)8, and Lys9, with the backbone for most of the sst2-selective antagonists comprised a Type-II' beta-turn. Hence, the sst2-selective antagonist pharmacophore is very similar to the sst2-selective agonist pharmacophore previously described.

Magnetic structure of two- and three-dimensional supramolecular compounds

Supramolecular chiral networks of oxalato-bridged transition metals show either two- or three-dimensional structural features. The magnetic structures of such compounds have been investigated by means of elastic neutron powder diffraction.