Instability of the insertional mutation in Cftr(TgH(neoim)Hgu )cystic fibrosis mouse model

BACKGROUND: A major boost to the cystic fibrosis disease research was given by the generation of various mouse models using gene targeting in embryonal stem cells. Moreover, the introduction of the same mutation on different inbred strains generating congenic strains facilitated the search for modifier genes. From the original Cftr(TgH(neoim)Hgu )CF mouse model we have generated using strict brother × sister mating two inbred Cftr(TgH(neoim)Hgu )mouse lines (CF/1 and CF/3). Thereafter, the insertional mutation was introgressed from CF/3 into three inbred backgrounds (C57BL/6, BALB/c, DBA/2J) generating congenic animals. In every backcross cycle germline transmission of the insertional mutation was monitored by direct probing the insertion via Southern RFLP. In order to bypass this time consuming procedure we devised an alternative PCR based protocol whereby mouse strains are differentiated at the Cftr locus by Cftr intragenic microsatellite genotypes that are tightly linked to the disrupted locus. RESULTS: Using this method we were able to identify animals carrying the insertional mutation based upon the differential haplotypic backgrounds of the three inbred strains and the mutant Cftr(TgH(neoim)Hgu )at the Cftr locus. Moreover, this method facilitated the identification of the precise vector excision from the disrupted Cftr locus in two out of 57 typed animals. This reversion to wild type status took place without any loss of sequence revealing the instability of insertional mutations during the production of congenic animals. CONCLUSIONS: We present intragenic microsatellite markers as a tool for fast and efficient identification of the introgressed locus of interest in the recipient strain during congenic animal breeding. Moreover, the same genotyping method allowed the identification of a vector excision event, posing questions on the stability of insertional mutations in mice

Spontaneous rescue from cystic fibrosis in a mouse model

BACKGROUND: From the original Cftr(TgH(neoim)Hgu )mutant mouse model with a divergent genetic background (129P2, C57BL/6, MF1) we have generated two inbred Cftr(TgH(neoim)Hgu )mutant strains named CF/1-Cftr(TgH(neoim)Hgu )and CF/3-Cftr(TgH(neoim)Hgu), which are fertile and show normal growth and lifespan. Initial genome wide scan analysis with microsatellite markers indicated that the two inbred strains differed on the genetic level. In order to further investigate whether these genetic differences have an impact on the disease phenotype of cystic fibrosis we characterised the phenotype of the two inbred strains. RESULTS: Reduced amounts, compared to wild type control animals, of correctly spliced Cftr mRNA were detected in the nasal epithelia, lungs and the intestine of both inbred Cftr(TgH(neoim)Hgu )strains, with higher residual amount observed for CF/1-Cftr(TgH(neoim)Hgu )than CF/3-Cftr(TgH(neoim)Hgu )for every investigated tissue. Accordingly the amounts of wild type Cftr protein in the intestine were 9% for CF/1-Cftr(TgH(neoim)Hgu )and 4% for CF/3-Cftr(TgH(neoim)Hgu). Unlike the apparent strain and/or tissue specific regulation of Cftr mRNA splicing, short circuit current measurements in the respiratory and intestinal epithelium revealed that both strains have ameliorated the basic defect of cystic fibrosis with a presentation of a normal electrophysiology in both tissues. CONCLUSION: Unlike the outbred Cftr(TgH(neoim)Hgu )insertional mouse model, which displayed the electrophysiological defect in the gastrointestinal and respiratory tracts characteristic of cystic fibrosis, both inbred Cftr(TgH(neoim)Hgu )strains have ameliorated the electrophysiological defect. On the basis of these findings both CF/1-Cftr(TgH(neoim)Hgu )and CF/3-Cftr(TgH(neoim)Hgu )offer an excellent model whereby determination of the minimal levels of protein required for the restoration of the basic defect of cystic fibrosis can be studied, along with the modulating factors which may affect this outcome

Very mild disease phenotype of congenic CftrTgH(neoim)Hgu cystic fibrosis mice

Background: A major boost to cystic fibrosis disease research was given by the generation of various mouse models using gene targeting in embryonal stem cells. Moreover, the introduction of the same mutation on different inbred strains generating congenic strains facilitated the search for modifier genes. From the original CftrTgH(neoim)Hgumouse model with a divergent genetic background (129/Sv, C57BL/6, HsdOla:MF1) two inbred mutant mouse strains CF/1-CftrTgH(neoim)Hguand CF/3-CftrTgH(neoim)Hguhad been generated using strict brother × sister mating. CF/1-CftrTgH(neoim)Hguand CF/3-CftrTgH(neoim)Hgumice were fertile and showed normal growth and lifespan. In this work the CftrTgH(neoim)Hguinsertional mutation was backcrossed from CF/3-CftrTgH(neoim)Hguonto the inbred backgrounds C57BL/6J and DBA/2J generating congenic animals in order to clarify the differential impact of the Cftr mutation and the genetic background on the disease phenotype of the cystic fibrosis mutant mice. Clinical and electrophysiological features of the two congenic strains were compared with those of CF/1-CftrTgH(neoim)Hguand CF/3-CftrTgH(neoim)Hguand wild type controls. Results: Under the standardized housing conditions of the animal facility, the four mouse strains CF/1-CftrTgH(neoim)Hgu, CF/3-CftrTgH(neoim)Hgu, D2.129P2(CF/3)-CftrTgH(neoim)Hguand B6.129P2(CF/3)-CftrTgH(neoim)Hguexhibited normal life expectancy. Growth of congenic cystic fibrosis mice was comparable with that of wild type controls. All mice but D2.129P2(CF/3)-CftrTgH(neoim)Hgufemales were fertile. Short circuit current measurements revealed characteristic response profiles of the HsdOla:MF1, DBA/2J and C57BL/6J backgrounds in nose, ileum and colon. All cystic fibrosis mouse lines showed the disease-typical hyperresponsiveness to amiloride in the respiratory epithelium. The mean chloride secretory responses to carbachol or forskolin were 15-100% of those of the cognate wild type control animals. Conclusion: The amelioration of the clinical features and of the basic defect that had emerged during the generation of CF/3-CftrTgH(neoim)Hgumice was retained in the congenic mice indicating that the Cftr linkage group or other loci shared between the inbred strains contain(s) the major modifier(s) of attenuation of cystic fibrosis symptoms

Towards Our Common Digital Future. Flagship Report.

In the report “Towards Our Common Digital Future”, the WBGU makes it clear that sustainability strategies and concepts need to be fundamentally further developed in the age of digitalization. Only if digital change and the Transformation towards Sustainability are synchronized can we succeed in advancing climate and Earth-system protection and in making social progress in human development. Without formative political action, digital change will further accelerate resource and energy consumption, and exacerbate damage to the environment and the climate. It is therefore an urgent political task to create the conditions needed to place digitalization at the service of sustainable development

The ter Mutation in the Rat Dnd1 Gene Initiates Gonadal Teratomas and Infertility in Both Genders

A spontaneous mutation leading to the formation of congenital ovarian and testicular tumors was detected in the WKY/Ztm rat strain. The histological evaluation revealed derivatives from all three germ layers, thereby identifying these tumors as teratomas. Teratocarcinogenesis was accompanied by infertility and the underlying mutation was termed ter. Linkage analysis of 58 (WKY-ter×SPRD-Cu3) F2 rats associated the ter mutation with RNO18 (LOD = 3.25). Sequencing of candidate genes detected a point mutation in exon 4 of the dead-end homolog 1 gene (Dnd1), which introduces a premature stop codon assumed to cause a truncation of the Dnd1 protein. Genotyping of the recessive ter mutation revealed a complete penetrance of teratocarcinogenesis and infertility in homozygous ter rats of both genders. Morphologically non-tumorous testes of homozygous ter males were reduced in both size and weight. This testicular malformation was linked to a lack of spermatogenesis using immunohistochemical and histological staining. Our WKY-Dnd1ter/Ztm rat is a novel animal model to investigate gonadal teratocarcinogenesis and the molecular mechanisms involved in germ cell development of both genders

Rethinking Land in the Anthropocene: from Separation to Integration

Nur wenn sich unser Umgang mit Land grundlegend ändert, können die Klimaschutzziele erreicht, der dramatische Verlust der biologischen Vielfalt abgewendet und das globale Ernährungssystem nachhaltig gestaltet werden. Der WBGU schlägt fünf exemplarische Mehrgewinnstrategien vor, um Konkurrenzen zwischen Nutzungsansprüchen zu überwinden. Diese sollten durch fünf Governance-Strategien vorangetrieben werden, darunter insbesondere die Setzung geeigneter Rahmenbedingungen, eine Neuorientierung der EUPolitik und die Errichtung von Gemeinschaften gleichgesinnter Staaten.Only if there is a fundamental change in the way we manage land can we reach the targets of climate-change mitigation, avert the dramatic loss of biodiversity and make the global food system sustainable. The WBGU proposes five multiple-benefit strategies illustrating ways of overcoming competition between rival claims to the use of land. These should be promoted by five governance strategies, especially by setting suitable framework conditions, reorienting EU policy and establishing alliances of like-minded states

Unsere gemeinsame digitale Zukunft

Das Gutachten „Unsere gemeinsame digitale Zukunft“ macht deutlich, dass Nachhaltigkeitsstrategien und -konzepte im Zeitalter der Digitalisierung grundlegend weiterentwickelt werden müssen. Nur wenn der digitale Wandel und die Transformation zur Nachhaltigkeit konstruktiv verzahnt werden, kann es gelingen, Klima- und Erdsystemschutz sowie soziale Fortschritte menschlicher Entwicklung voranzubringen. Ohne aktive politische Gestaltung wird der digitale Wandel den Ressourcen- und Energieverbrauch sowie die Schädigung von Umwelt und Klima weiter beschleunigen. Daher ist es eine vordringliche politische Aufgabe, Bedingungen dafür zu schaffen, die Digitalisierung in den Dienst nachhaltiger Entwicklung zu stellen