Floxed allele for conditional inactivation of the GABAB(1) geneHaller, Corinne; Casanova, Emilio; Müller, Matthias; Vacher, Claire‐Marie; Vigot, Rejan; Doll, Thierry; Barbieri, Samuel; Gassmann, Martin; Bettler, Bernhard
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20073pmid: 15493018
GABAB receptors are the G‐protein‐coupled receptors for the neurotransmitter GABA. GABAB receptors are broadly expressed in the nervous system. Their complete absence in mice causes premature lethality or—when mice are viable—epilepsy, impaired memory, hyperalgesia, hypothermia, and hyperactivity. A spatially and temporally restricted loss of GABAB function would allow addressing how the absence of GABAB receptors leads to these diverse phenotypes. To permit a conditional gene inactivation, we flanked critical exons of the GABAB(1) gene with lox511 sites. GABAB(1)lox511/lox511 mice exhibit normal levels of GABAB(1) protein, are fertile, and do not display any behavioral phenotype. We crossed GABAB(1)lox511/lox511 with Cre‐deleter mice to produce mice with an unrestricted GABAB receptor elimination. These GABAB(1)−/− mice no longer synthesize GABAB(1) protein and exhibit the expected behavioral abnormalities. The conditional GABAB(1) allele described here is therefore suitable for generating mice with a site‐ and time‐specific loss of GABAB function. genesis 40:125–130, 2004. © 2004 Wiley‐Liss, Inc.
Cre recombinase specificity defined by the tau locusKorets‐Smith, Ella; Lindemann, Lothar; Tucker, Kerry Lee; Jiang, Caoyang; Kabacs, Nikolett; Belteki, Gusztav; Haigh, Jody; Gertsenstein, Marina; Nagy, Andras
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20074pmid: 15493019
We generated a transgenic mouse line (tau::Cre) by targeting the Cre to the tau locus (Mapt). Based on previous reports on the expression of Tau during development, we expected the Cre recombinase to be expressed in a neuron‐specific and pan‐neuronal manner. However, intercrosses between the tau::Cre and the Cre‐activatable reporter animals resulted in offspring with recombination either restricted to the nervous system or throughout the entire conceptus, indicating expression of Tau early in development. The percentage of neuron‐specific excision was dependent on the Cre reporter used representing different Cre target sites in the mouse genome. In spite of the observed variability, our data suggest that the tau::Cre mouse line can be used for pan‐neuronal recombination of floxed alleles when it is used with caution. genesis 40:131–138, 2004. © 2004 Wiley‐Liss, Inc.
CRBP‐III:lacZ expression pattern reveals a novel heterogeneity of vascular endothelial cellsCaprioli, Arianna; Zhu, Hongxin; Sato, Thomas N.
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20075pmid: 15493015
Vascular endothelial cells are structurally and functionally heterogeneous. However, the molecular basis of this heterogeneity remains poorly defined. We used subtractive and differential screening to identify genes that exhibit heterogeneous expression patterns among vascular endothelial cells. One such gene is cellular retinol binding protein III (CRBP‐III/Rbp7). Analysis of the lacZ knockin line for this gene (CRBP‐III:lacZ) revealed a novel organ‐specific vascular endothelial expression pattern. LacZ was expressed in vascular endothelial cells in heart, skeletal muscle, adipose tissues, thymus, and salivary gland. However, it was not detected in other tissues such as brain, liver, and lung. Furthermore, the expression within each organ was primarily restricted to small capillary endothelial cells, but could not be detected in larger vessels. This organ‐specific vascular endothelial expression of CRPB:lacZ is relatively resistant to the changes of organ microenvironment. However, the level of expression can be modified by vitamin A deficiency. Therefore, our results provide novel molecular evidence for the heterogeneity of vascular endothelial cells. genesis 40:139–145, 2004. © 2004 Wiley‐Liss, Inc.
X‐inactivation is stably maintained in mouse embryos deficient for histone methyl transferase G9aOhhata, Tatsuya; Tachibana, Makoto; Tada, Masako; Tada, Takashi; Sasaki, Hiroyuki; Shinkai, Yoichi; Sado, Takashi
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20077pmid: 15493016
Summary: One of the two X chromosomes becomes inactivated during early development of female mammals. Recent studies demonstrate that the inactive X chromosome is rich in histone H3 methylated at Lys‐9 and Lys‐27, suggesting an important role for these modifications in X‐inactivation. It has been shown that in the mouse Eed is required for maintenance of X‐inactivation in the extraembryonic lineages. Interestingly, Eed associates with Ezh2 to form a complex possessing histone methyltransferase activity predominantly for H3 Lys‐27. We previously showed that G9a is one of the histone methyltransferases specific for H3 Lys‐9 and is essential for embryonic development. Here we examined X‐inactivation in mouse embryos deficient for G9a. Expression of Xist, which is crucial for the initiation of X‐inactivation, was properly regulated and the inactivated X chromosome was stably maintained even in the absence of G9a. These results demonstrate that G9a is not essential for X‐inactivation. genesis 40:151–156, 2004. © 2004 Wiley‐Liss, Inc.
Directing pluripotent cell differentiation using “diced RNA” in transient transfectionCarpenter, Lee; Zernicka‐Goetz, Magdalena
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20078pmid: 15515021
Embryonic stem (ES) and embryonic carcinoma (EC) cells are pluripotent and have the capacity to differentiate into many cell types. The ability to direct their differentiation should have considerable practical applications. Here, we first report the use of diced short interfering RNAi against Oct4 in a transient approach, to direct differentiation of ES towards the trophectoderm lineage. We then apply this approach to downregulate Smad4 in mouse P19 EC cells. We have found that this leads to an increase in the levels of Pax6 (a neuroectoderm marker), reduction in the levels of Brachyury (a mesoderm marker), and a 3‐fold increase in the number of βIII tubulin‐positive colonies when these cells were allowed to differentiate. This indicates a redirection of cell fate towards the neuroectoderm lineage. Thus, transient RNAi could provide a valuable tool to direct pluripotent cells along specific pathways of differentiation while circumventing permanent genetic changes. genesis 40:157–163, 2004. © 2004 Wiley‐Liss, Inc.
Drosophila tudor is essential for polar granule assembly and pole cell specification, but not for posterior patterningThomson, Travis; Lasko, Paul
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20079pmid: 15495201
Pole cells and posterior segmentation in Drosophila are specified by maternally encoded genes whose products accumulate at the posterior pole of the oocyte. Among these genes is tudor (tud). Progeny of hypomorphic tud mothers lack pole cells and have variable posterior patterning defects. We have isolated a null allele to further investigate tud function. While no pole cells are ever observed in embryos from tud‐null mothers, 15% of these embryos have normal posterior patterning. OSKAR (OSK) and VASA (VAS) proteins, and nanos (nos) RNA, all initially localize to the pole plasm of tud‐null oocytes and embryos from tud‐null mothers, while localization of germ cell‐less (gcl) and polar granule component (pgc), is undetectable or severely reduced. In embryos from tud‐null mothers, polar granules are greatly reduced in number, size, and electron density. Thus, tud is dispensable for somatic patterning, but essential for pole cell specification and polar granule formation. genesis 40:164–170, 2004. © 2004 Wiley‐Liss, Inc.
Sox10‐rtTA mouse line for tetracycline‐inducible expression of transgenes in neural crest cells and oligodendrocytesLudwig, Andreas; Schlierf, Beate; Schardt, Anke; Nave, Klaus‐Armin; Wegner, Michael
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20083pmid: 15493017
Using gene targeting, we inserted a high‐affinity variant of the reverse tetracycline controlled transactivator (rtTA) into the genomic Sox10 locus. This rtTA transgene faithfully recapitulated Sox10 expression in the emerging neural crest, several of its derivatives, and in oligodendrocytes. It was furthermore able to induce expression of a tetracycline inducible transgenic reporter gene in a doxycycline‐dependent manner. Induction was fast, with substantial reporter gene expression visible 6 h after the onset of doxycycline treatment. Shut‐off, in contrast, exhibited delayed kinetics, which probably correlated with doxycycline clearance rates. This mouse provides a useful tool for generating tetracycline‐controlled gene expression in neural crest and oligodendrocytes. genesis 40:171–175, 2004. © 2004 Wiley‐Liss, Inc.
Isolation of mutations with dumpy‐like phenotypes and of collagen genes in the nematode Pristionchus pacificusKenning, Charlotte; Kipping, Isabel; Sommer, Ralf J.
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20084pmid: 15493014
The nematode Pristionchus pacificus was developed as a satellite system in evolutionary developmental biology and forward and reverse genetic approaches allow a detailed comparison of various developmental processes between P. pacificus and Caenorhabditis elegans. To facilitate map‐based cloning in P. pacificus, a genome map was generated including a genetic linkage map of ∼300 molecular markers and a physical map of 10,000 BAC clones. Here, we describe the isolation and characterization of more than 40 morphological mutations that can be used as genetic markers. These mutations fall into 12 Dumpy genes and one Roller gene that represent morphological markers for all six P. pacificus chromosomes. Using an in silico approach, we identified ∼150 hits of P. pacificus collagen genes in the available EST, BAC‐end, and fosmid‐end sequences. However, 1:1 orthologs could only be identified for fewer than 20 collagen genes. genesis 40:176–183, 2004. © 2004 Wiley‐Liss, Inc.
Positional cloning and characterization of mouse mei8, a disrupted allele of the meiotic cohesin Rec8Bannister, Laura A.; Reinholdt, Laura G.; Munroe, Robert J.; Schimenti, John C.
2004 Genesis: the Journal of Genetics and Development
doi: 10.1002/gene.20085pmid: 15515002
A novel mutation, mei8, was isolated in a forward genetic screen for infertility mutations induced by chemical mutagenesis of ES cells. Homozygous mutant mice are sterile. Mutant females exhibit ovarian dysgenesis and lack ovarian follicles at reproductive maturity. Affected males have small testes due to arrest of spermatogenesis during meiotic prophase I. Genetic mapping and positional cloning of mei8 led to the identification of a mutation in Rec8, a homolog of the yeast meiosis‐specific cohesin gene REC8. Analysis of meiosis in Rec8mei8/Rec8mei8 spermatocytes showed that, while initiation of recombination and synapsis occurs, REC8 is required for the completion and/or maintenance of synapsis, cohesion of sister chromatids, and the formation of chiasmata, as it is in other organisms. However, unlike yeast and Caenorhabditis elegans, localization of REC8 on meiotic chromosomes is not required for the assembly of axial elements. genesis 40:184–194, 2004. © 2004 Wiley‐Liss, Inc.