Prolonged cultivation of Saccharomyces cerevisiae in aerobic, glucose-limited chemostat cultures (dilution rate, 010 h1) resulted in a progressive decrease of the residual glucose concentration (from 20 to 8 mg l1 after 200 generations). This increase in the affinity for glucose was accompanied by a fivefold decrease of fermentative capacity, and changes in cellular morphology. These phenotypic changes were retained when single-cell isolates from prolonged cultures were used to inoculate fresh chemostat cultures, indicating that genetic changes were involved. Kinetic analysis of glucose transport in an evolved strain revealed a decreased Km, while Vmax was slightly increased relative to the parental strain. Apparently, fermentative capacity in the evolved strain was not controlled by glucose uptake. Instead, enzyme assays in cell extracts of the evolved strain revealed strongly decreased capacities of enzymes in the lower part of glycolysis. This decrease was corroborated by genome-wide transcriptome analysis using DNA microarrays. In aerobic batch cultures on 20 g glucose l1, the specific growth rate of the evolved strain was lower than that of the parental strain (028 and 037 h1, respectively). Instead of the characteristic instantaneous production of ethanol that is observed when aerobic, glucose-limited cultures of wild-type S. cerevisiae are exposed to excess glucose, the evolved strain exhibited a delay of 90 min before aerobic ethanol formation set in. This study demonstrates that the effects of selection in glucose-limited chemostat cultures extend beyond glucose-transport kinetics. Although extensive physiological analysis offered insight into the underlying cellular processes, the evolutionary driving force for several of the observed changes remains to be elucidated
Prolonged selection in aerobic, glucose-limited chemostat cultures of Saccharomyces cerevisiae causes a partial loss of glycolytic capacity.
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View SamplesWe performed RNA-Seq and compared expression levels of genes of reactivated LCMV.GP66-77 specific CD4 T cells isolated from bone marrow (BM) and spleen of LCMV.GP61-80 primed C57BL/6 mice. Cells were isolated 3 days after antigenic re-challenge Overall design: C57BL/6 mice were primed at day 0 with LCMV.GP61-80-NP-MSA + poly(I:C) and immunized again at day 14 with LCMV.GP61-80 + poly(I:C). 60 days later, C57BL/6 mice were boosted with LCMV.GP61-80-NP-MSA + poly(I:C) and 3 days after the boost, LCMV specific CD4 T cells were isolated from BM and spleen
Nonfollicular reactivation of bone marrow resident memory CD4 T cells in immune clusters of the bone marrow.
Age, Specimen part, Cell line, Subject
View SamplesWe compared gene expression profiles of Th cells, macrophages and monocytes isolated from the inflamed colon of colitis induced by the transfer of WT versus Tbx21-/- Th cells in Rag1-/- recipients.
T-bet expression by Th cells promotes type 1 inflammation but is dispensable for colitis.
Specimen part
View SamplesDetermination of the molecular properties of genetically targeted cell types has led to fundamental insights into mouse brain function and dysfunction. Here, we report an efficient strategy for precise exploration of gene expression events in specific cell types in a broad range of species. We demonstrate that classically defined, homologous neuronal and glial cell types differ between rodent and human by the expression of hundreds of orthologous, cell specific genes. Confirmation that these genes are differentially active was obtained using epigenetic mapping, quantitative PCR, and immunofluorescence localization. Studies of sixteen human postmortem brains revealed cell-specific molecular responses to aging, and the induction of a shared, robust response to an unknown external event experienced by three donors. Our data establish a comprehensive approach for analysis of unique molecular events associated with specific circuits and cell types in a wide variety of human conditions. Overall design: RNA purified from nuclei or cytoplasm from mouse, rat, or human cerebellum. ATAC-seq was also performed using cerebellar nuclei from the three species.
Species and cell-type properties of classically defined human and rodent neurons and glia.
Sex, Age, Specimen part, Subject
View SamplesDetermination of the molecular properties of genetically targeted cell types has led to fundamental insights into mouse brain function and dysfunction. Here, we report an efficient strategy for precise exploration of gene expression events in specific cell types in a broad range of species. We demonstrate that classically defined, homologous neuronal and glial cell types differ between rodent and human by the expression of hundreds of orthologous, cell specific genes. Confirmation that these genes are differentially active was obtained using epigenetic mapping, quantitative PCR, and immunofluorescence localization. Studies of sixteen human postmortem brains revealed cell-specific molecular responses to aging, and the induction of a shared, robust response to an unknown external event experienced by three donors. Our data establish a comprehensive approach for analysis of unique molecular events associated with specific circuits and cell types in a wide variety of human conditions. Overall design: RNA purified from nuclei or cytoplasm from mouse, rat, or human cerebellum. ATAC-seq was also performed using cerebellar nuclei from the three species.
Species and cell-type properties of classically defined human and rodent neurons and glia.
Specimen part, Cell line, Subject
View SamplesPulmonary alveoli are complex architectural units thought to undergo endogenous or pharmacologically induced programs of regeneration and degeneration. To study the molecular mechanism of alveoli loss mice were calorie restricted at different timepoints. Lungs were harvested and processed for RNA extraction.
Calorie-related rapid onset of alveolar loss, regeneration, and changes in mouse lung gene expression.
Time
View SamplesIt has been shown that dexamethasone (Dex) impairs the normal lung septation that occurs in the early postnatal period. Treatment with retinoic acid (ATRA) abrogates the effects of Dex. To understand the molecular basis for the Dex indiced inhibition of the formation of the alveoli and the ability of ATRA to prevent the inhibition of septation, gene expression was analyzed in 4-day old mice treated with diluent (control), Dex-treated and ATRA+Dex-treated.
DNA microarray analysis of neonatal mouse lung connects regulation of KDR with dexamethasone-induced inhibition of alveolar formation.
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View SamplesThe normally virulent type-I RH parasite is rendered avirulent when lacking ROP5. The avirulent phenotype is a consequence of interaction with the host innate immune system. We sought to understand if ROP5 alters host gene expression in order to escape host defenses. We saw no gene expression differences between host cells infected with wt (RHku80) or RHku80rop5 parasites. We have included uninfected HFF samples that were harvested in parallel with the infected samples.
The polymorphic pseudokinase ROP5 controls virulence in Toxoplasma gondii by regulating the active kinase ROP18.
Specimen part
View SamplesWe performed total RNA-Seq of murine Th1 cells which were four times reactivated in vitro in the presence of irradiated APC'srepeatedly activated in vitro. Overall design: CD4+CD62Lhi (naive) cells were isolated from C57BL/6 mice, activated with aCD3 and aCD28 an cultured under Th1 polarizing conditions in the presence of irradiated APCs. Every sixth day cells were harvested, restimulated with aCD3 and aCD28 and cultured under Th1 polarizing conditions in the presence of irradiated APCs APCs. After four rounds of restimulation, total RNA was extracted and cDNA libraries for total RNA sequencing were generated using “TruSeq® Stranded Total RNA Library” kit (Illumina, San Diego, CA, USA).
MicroRNA-31 Reduces the Motility of Proinflammatory T Helper 1 Lymphocytes.
Specimen part, Subject
View SamplesTo identify transcription factors critically involved in the Tfh cell transcriptional network, antigen-specific Tfh and non-Tfh cells from a day 8 immune response were analyzed by RNA-seq. Overall design: Ovalbumin-specific T cells from OT-II TCR-transgenic mice were transferred into C57BL/6 recipients, which were immunized subcutaneously with nitrophenol coupled to ovalbumin. Eight days after immunization, transgenic T cells from pooled inguinal lymph nodes were sorted for a CD44hi CXCR5+ PD-1+ Tfh and CD44hi CXCR5- PD-1- non-Tfh cell phenotype for analysis by RNA-seq.
Bach2 Controls T Follicular Helper Cells by Direct Repression of Bcl-6.
Specimen part, Subject
View Samples