Comparative transcription map of the wobbler critical region on mouse chromosome 11 and the homologous region on human chromosome 2p13-14

BACKGROUND: To support the positional cloning of the mouse mutation wobbler (wr) the corresponding regions on human Chr2p13-14 and mouse Chr11 were analyzed in detail and compared with respect to gene content, order, and orientation. RESULTS: The gene content of the investigated regions was highly conserved between the two species: 20 orthologous genes were identified on our BAC/YAC contig comprising 4.5 Mb between REL/Rel and RAB1A/Rab1a. Exceptions were pseudogenes ELP and PX19 whose mouse counterparts were not located within the analyzed region. Two independently isolated genomic clones indicate an inversion between man and mouse with the inverted segment being identical to the wobbler critical interval. We investigated the wobbler critical region by extensive STS/EST mapping and genomic sequencing. Additionally, the full-length cDNA sequences of four newly mapped genes as well as the previously mapped gene Otx1 were established and subjected to mutation analysis. Our data indicate that all genes in the wr critical region have been identified. CONCLUSION: Unexpectedly, neither mutation analysis of cDNAs nor levels of mRNAs indicated which of the candidate genes might be affected by the wr mutation. The possibility arises that there might be hitherto unknown effects of mutations, in addition to structural changes of the mRNA or regulatory abnormalities

Gain and loss of TASK3 channel function and its regulation by novel variation cause KCNK9 imprinting syndrome

Background: Genomics enables individualized diagnosis and treatment, but large challenges remain to functionally interpret rare variants. To date, only one causative variant has been described for KCNK9 imprinting syndrome (KIS). The genotypic and phenotypic spectrum of KIS has yet to be described and the precise mechanism of disease fully understood. Methods: This study discovers mechanisms underlying KCNK9 imprinting syndrome (KIS) by describing 15 novel KCNK9 alterations from 47 KIS-affected individuals. We use clinical genetics and computer-assisted facial phenotyping to describe the phenotypic spectrum of KIS. We then interrogate the functional effects of the variants in the encoded TASK3 channel using sequence-based analysis, 3D molecular mechanic and dynamic protein modeling, and in vitro electrophysiological and functional methodologies. Results: We describe the broader genetic and phenotypic variability for KIS in a cohort of individuals identifying an additional mutational hotspot at p.Arg131 and demonstrating the common features of this neurodevelopmental disorder to include motor and speech delay, intellectual disability, early feeding difficulties, muscular hypotonia, behavioral abnormalities, and dysmorphic features. The computational protein modeling and in vitro electrophysiological studies discover variability of the impact of KCNK9 variants on TASK3 channel function identifying variants causing gain and others causing loss of conductance. The most consistent functional impact of KCNK9 genetic variants, however, was altered channel regulation. Conclusions: This study extends our understanding of KIS mechanisms demonstrating its complex etiology including gain and loss of channel function and consistent loss of channel regulation. These data are rapidly applicable to diagnostic strategies, as KIS is not identifiable from clinical features alone and thus should be molecularly diagnosed. Furthermore, our data suggests unique therapeutic strategies may be needed to address the specific functional consequences of KCNK9 variation on channel function and regulation

Structure-antiproliferative activity studies on L-proline- and homoproline-4-N-pyrrolidine-3-thiosemicarbazone hybrids and their nickel(II), palladium(II) and copper(II) complexes

International audienc

A Combined Deep Eutectic Solvent–Ionic Liquid Process for the Extraction and Separation of Platinum Group Metals (Pt, Pd, Rh)

Recovery of platinum group metals from spent materials is becoming increasingly relevant due to the high value of these metals and their progressive depletion. In recent years, there is an increased interest in developing alternative and more environmentally benign processes for the recovery of platinum group metals, in line with the increased focus on a sustainable future. To this end, ionic liquids are increasingly investigated as promising candidates that can replace state-of-the-art approaches. Specifically, phosphonium-based ionic liquids have been extensively investigated for the extraction and separation of platinum group metals. In this paper, we present the extraction capacity of several phosphonium-based ionic liquids for platinum group metals from model deep eutectic solvent-based acidic solutions. The most promising candidates, P66614Cl and P66614B2EHP, which exhibited the ability to extract Pt, Pd, and Rh quantitively from a mixed model solution, were additionally evaluated for their capacity to recover these metals from a spent car catalyst previously leached into a choline-based deep eutectic solvent. Specifically, P66614Cl afforded extraction of the three target precious metals from the leachate, while their partial separation from the interfering Al was also achieved since a significant amount (approx. 80%) remained in the leachate

Novel thiosalicylate-based ionic liquids for heavy metal extractions

AbstractThis study aims to develop novel ammonium and phosphonium ionic liquids (ILs) with thiosalicylate (TS) derivatives as anions and evaluate their extracting efficiencies towards heavy metals in aqueous solutions. Six ILs were synthesized, characterized, and investigated for their extracting efficacies for cadmium, copper, and zinc. Liquid-liquid extractions of Cu, Zn, or Cd with ILs after 1–24h using model solutions (pH 7; 0.1M CaCl2) were assessed using flame atomic absorption spectroscopy (F-AAS). Phosphonium-based ILs trihexyltetradecylphosphonium 2-(propylthio)benzoate [P66614][PTB] and 2-(benzylthio)benzoate [P66614][BTB] showed best extraction efficiency for copper and cadmium, respectively and zinc was extracted to a high degree by [P66614][BTB] exclusively

In cis TP53 and RAD51C pathogenic variants may predispose to sebaceous gland carcinomas

Pathogenic variants in TP53 have been classically thought to cause Li-Fraumeni syndrome (LFS), a cancer predisposition with high risks for various childhood- and adult-onset malignancies. However, increased genetic testing has lately revealed, that pathogenic variant carriers exhibit a broader range of phenotypes and that penetrance may be dependent both on variant type and modifiers. Using next generation sequencing and short tandem repeat analysis, we identified germline pathogenic variants in TP53 and RAD51C located in cis on chromosome 17 in a 43-year-old male, who has developed a rare sebaceous gland carcinoma (SGC) but so far no tumors of the LFS spectrum. This course mirrors a Trp53-Rad51c-double-mutant cis mouse-model, which similarly develops SGC, while the characteristic Trp53-associated tumor spectrum occurs with significantly lower frequency. Therefore, we propose that co-occurent pathogenic variants in RAD51C and TP53 may predispose to SGC, reminiscent of Muir-Torre syndrome. Further, this report supports the diversity of clinical presentations associated with germline TP53 alterations, and thus, the proposed expansion of LFS to heritable TP53-related cancer syndrome

Efficient immunoglobulin gene disruption and targeted replacement in rabbit using zinc finger nucleases.

Rabbits are widely used in biomedical research, yet techniques for their precise genetic modification are lacking. We demonstrate that zinc finger nucleases (ZFNs) introduced into fertilized oocytes can inactivate a chosen gene by mutagenesis and also mediate precise homologous recombination with a DNA gene-targeting vector to achieve the first gene knockout and targeted sequence replacement in rabbits. Two ZFN pairs were designed that target the rabbit immunoglobulin M (IgM) locus within exons 1 and 2. ZFN mRNAs were microinjected into pronuclear stage fertilized oocytes. Founder animals carrying distinct mutated IgM alleles were identified and bred to produce offspring. Functional knockout of the immunoglobulin heavy chain locus was confirmed by serum IgM and IgG deficiency and lack of IgM(+) and IgG(+) B lymphocytes. We then tested whether ZFN expression would enable efficient targeted sequence replacement in rabbit oocytes. ZFN mRNA was co-injected with a linear DNA vector designed to replace exon 1 of the IgM locus with ∼1.9 kb of novel sequence. Double strand break induced targeted replacement occurred in up to 17% of embryos and in 18% of fetuses analyzed. Two major goals have been achieved. First, inactivation of the endogenous IgM locus, which is an essential step for the production of therapeutic human polyclonal antibodies in the rabbit. Second, establishing efficient targeted gene manipulation and homologous recombination in a refractory animal species. ZFN mediated genetic engineering in the rabbit and other mammals opens new avenues of experimentation in immunology and many other research fields

DataSheet_2_Consequences of somatic mutations of GIRK1 detected in primary malign tumors on expression and function of G-protein activated, inwardly rectifying, K+ channels.pdf

A search in the GDC Data Portal revealed 304 documented somatic mutations of the KCNJ3 gene in primary tumors (out of 10.202 cases). Most affected tumor types were carcinomas from uterus, skin and lung, while breast cancer exerted the lowest number of somatic mutations. We focused our research on 15 missense mutations within the region between TM1 and TM2, comprising the pore helix and ion selectivity signature. Expression was measured by confocal laser scan microscopy of eGFP tagged GIRK1 subunits, expressed with and without GIRK4 in oocytes of Xenopus laevis. GIRK ion currents were activated via coexpressed m2Rs and measured by the Two Electrode Voltage Clamp technique. Magnitude of the total GIRK current, as well as the fraction of current inducible by the agonist, were measured. Ion selectivity was gauged by assessment of the PNa+/PK+ ratio, calculated by the GIRK current reversal potential in extracellular media at different Na+ and K+ concentrations. None of the tested mutations was able to form functional GIRK1 homooligomeric ion channels. One of the mutations, G145A, which locates directly to the ion selectivity signature, exerted an increased PNa+/PK+ ratio. Generally, the missense mutations studied can be categorized into three groups: (i) normal/reduced expression accompanied by reduced/absent function (S132Y, F136L, E139K, G145A, R149Q, R149P, G178D, S185Y, Q186R), (ii) normal/increased expression as well as increased function (E140M, A142T, M184I) and (iii) miniscule expression but increased function relative to expression levels (I151N, G158S). We conclude, that gain of function mutations, identical or similar to categories (ii) and (iii), may potentially be involved in genesis and progression of malignancies in tissues that exert a high rate of occurrence of somatic mutations of KCNJ3.</p