Project description:Long-term memory formation is attributed to experience-dependent gene expression. Dynamic changes in histone methylation status are essential for epigenetic regulation of memory consolidation-related genes. Here, we demonstrated that plant homeodomain finger protein 2 (PHF2) histone demethylase is upregulated in the mouse hippocampus during experience and plays an essential role in memory formation. QuantSeq analysis was performed to examine differential gene expression in the hippocampus of WT and PHF2 t/g mice. Transgenic mice were created by injecting the CMV-Flag-PHF2 vector into fertilized eggs from C57BL6 mice. Transgenic lines were established from 9 founders that were identified via PCR-based genotyping. Among the transgenic lines, mice with high PHF2 expressed brains were singled out for breeding. First, selected heterozygote males were bred with heterozygote females which produced homozygote, heterozygote and wild littermates. Since it is difficult to differentiate between hetero and homo mice with genotyping, we separated homo mice from hetero mice by breeding each of hetero and homo mice with wild type mice and confirming the genotypes of their offspring. In other words, hetero mice bred with wild type mice would have both wild type and hetero genotype of offspring, while homo mice bred with wild type mice would only have hetero genotype of offspring. Through this process, we particularly selected the homo mice and this transgenic line was used and maintained for this experiment.
Project description:Total RNA-seq analyses for the differentially expressed genes (DEGs) among WT (c42), Lin28 R192G hetero mut hESC (c26_5), and Lin28 R192G homo mut hESC (c38) To gain further molecular insight into the observed Lin28R192G mutation in human ESCs, we performed total RNA-sequencing (RNA-seq) analysis of the WT, Lin28 R192G hetero mut hESC, and Lin28 R192G homo mut hESC. The genes that are differentially expressed (DEGs) in Lin28 R192G hetero mut and Lin28 R192G homo mut hES compared to WT hESC.
Project description:Transcriptional profiling of mouse postnatal SVZ NSCs comparing WT NSCs with KO NSCs under proliferating/undifferentiated states as well as differentiating conditions. Goal was to determine Dnmt3a-dependent gene expression changes in postnatal SVZ NSCs Two-condition experiment with a dye-swap design, WT NSCs vs. KO NSCs. Biological replicates: 4 replicates under proliferating/undifferentiation conditions, 2 replicates under differentiating conditions.
Project description:Transcriptional profiling of mouse postnatal SVZ NSCs comparing WT NSCs with KO NSCs under proliferating/undifferentiated states as well as differentiating conditions. Goal was to determine Dnmt3a-dependent gene expression changes in postnatal SVZ NSCs
Project description:Primitive neural stem cells (NSCs) could be derived from pluripotent mouse embryonic stem (ES) cells, and then differentiate into definitive-type neural stem cells which resemble NSCs obtained from the central nervous system. Hence, primitive NSCs define an early stage of neural induction and provide a model to understand the mechanism that controls initial neural commitment. In this study, we performed microarray assay to analyze the global transcriptional profiles in mouse ES cell-derived primitive and definitive NSCs and to depict the molecular changes during the multi-staged neural differentiation process. Primitive NSCs derived directly from ESCs in Lif (p-NSC_L), primitive NSCs that were sub-cultured in the presence of Lif and FGF (p-NSC_LF), as well as definitive NSCs derived from primitive NSCs in medium containing FGF and EGF, were collected for RNA extraction and hybridization on Affymetrix microarrays. Mouse ESCs and NSCs obtained from mouse embryonic brain (E11.5) were included for controls. For each cell type, we collected two biological replicate samples for microarray analysis.
Project description:Primitive neural stem cells (NSCs) could be derived from pluripotent mouse embryonic stem (ES) cells, and then differentiate into definitive-type neural stem cells which resemble NSCs obtained from the central nervous system. Hence, primitive NSCs define an early stage of neural induction and provide a model to understand the mechanism that controls initial neural commitment. In this study, we performed microarray assay to analyze the global transcriptional profiles in mouse ES cell-derived primitive and definitive NSCs and to depict the molecular changes during the multi-staged neural differentiation process.
Project description:The early onset of microglia activation in the Rp58 hetero-KO mice raised the possibility that chronic inflammation in the brain was induced by mutation even in the early life stage. To test this possibility, we examined gene expression profiles in the hippocampus of the Rp58 hetero-KO and the control wild-type mice in adulthood and old age months. Among the genes whose expression level was down-regulated in the aged wild-type mice, 341 genes were down-regulated in the adulthood Rp58 hetero-KO mice when both were compared with the adulthood wild-type mice. Conversely, the expression levels of 103 genes were up-regulated both in the adulthood Rp58 hetero-KO and the aged wild-type mice. In the pathway enrichment analysis, these 103 up-regulated genes included Cxcl10, Oas2, Oas1a, and Oasl2, which are associated with a pathway of immune response interferon gamma action on extracellular matrix and cell differentiation.
Project description:Three types of phenotypic expression of M-CM-^_-lactam resistance has been reported in MRSA: heterogeneous-, homogeneous-, and Eagle-type resistance. Heterogeneous to homogeneous (hetero-to-homo) conversion of M-CM-^_-lactam resistance is postulated to be caused by a chromosomal mutation (chr*) together with mecA-gene expression. The Eagle-type resistance is a special pattern of chr* expression in the pre-MRSA strain N315 under the strong mecI-gene mediated repression of mecA gene transcription. Here, for the identification of chr*, experiments were conducted using an in-vitro derived homogeneously imipenem-resistant MRSA strain N315M-bM-^HM-^FIPH5 (M-bM-^HM-^FIPH5). The strain was selected with imipenem 8 mg/L from the heterogeneously imipenem-resistant MRSA strain N315M-bM-^HM-^FIP (M-bM-^HM-^FIP). The whole genome sequencing of M-bM-^HM-^FIPH5 revealed the presence of a unique mutation in the rpoB gene, rpoB(N967I), causing the amino-acid (AA) substitution of Asp by Ile at the 967th AA position of the RNA polymerase M-CM-^_ subunit. The effect of the mutation on the phenotypic change was confirmed by constructing and studying the phenotype of the strains H5rpoB(I967N), a M-bM-^HM-^FIPH5-derived strain cured of the rpoB mutation, and N315rpoB(N967I), a N315-derived strain introduced with the mutated rpoB gene. H5rpoB(I967N) regained the hetero-MRSA phenotype, and the mutant strain N315rpoB(N967I) showed an Eagle-type phenotype similar to that of N315h4. Furthermore, subsequent whole genome sequencing revealed that N315h4 also had a missense mutation in the rpoB gene rpoB(R644H). The rpoB mutations caused decreased autolysis, prolonged doubling-time, and tolerance to bactericidal concentrations of methicillin. We concluded that the certain rpoB mutations are chr* responsible for the hetero-to-homo phenotypic conversion of MRSA. We compared the gene expression profiles of the wild-type strain and rpoB mutant (N967I) using a 60mer oligo array.
Project description:Three types of phenotypic expression of ß-lactam resistance has been reported in MRSA: heterogeneous-, homogeneous-, and Eagle-type resistance. Heterogeneous to homogeneous (hetero-to-homo) conversion of ß-lactam resistance is postulated to be caused by a chromosomal mutation (chr*) together with mecA-gene expression. The Eagle-type resistance is a special pattern of chr* expression in the pre-MRSA strain N315 under the strong mecI-gene mediated repression of mecA gene transcription. Here, for the identification of chr*, experiments were conducted using an in-vitro derived homogeneously imipenem-resistant MRSA strain N315∆IPH5 (∆IPH5). The strain was selected with imipenem 8 mg/L from the heterogeneously imipenem-resistant MRSA strain N315∆IP (∆IP). The whole genome sequencing of ∆IPH5 revealed the presence of a unique mutation in the rpoB gene, rpoB(N967I), causing the amino-acid (AA) substitution of Asp by Ile at the 967th AA position of the RNA polymerase ß subunit. The effect of the mutation on the phenotypic change was confirmed by constructing and studying the phenotype of the strains H5rpoB(I967N), a ∆IPH5-derived strain cured of the rpoB mutation, and N315rpoB(N967I), a N315-derived strain introduced with the mutated rpoB gene. H5rpoB(I967N) regained the hetero-MRSA phenotype, and the mutant strain N315rpoB(N967I) showed an Eagle-type phenotype similar to that of N315h4. Furthermore, subsequent whole genome sequencing revealed that N315h4 also had a missense mutation in the rpoB gene rpoB(R644H). The rpoB mutations caused decreased autolysis, prolonged doubling-time, and tolerance to bactericidal concentrations of methicillin. We concluded that the certain rpoB mutations are chr* responsible for the hetero-to-homo phenotypic conversion of MRSA.