Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other
Project description:We have used a retroviral integration screen to search for novel genes that regulate hematopoietic stem cell (HSC) function. One of the genes that conferred HSC dominance when overexpressed due to an adjacent retroviral insertion was Musashi 2 (Msi2), an RNA binding protein that can act as a translational inhibitor. A gene trap mouse model that inactivates the gene shows that Msi2 is more highly expressed in long term (LT) and short term (ST) HSCs as well as in lymphoid myeloid primed progenitors (LMPPs), but much less in intermediate progenitors and mature cells. Mice lacking Msi2 are fully viable up to more than a year but exhibit severe defects in primitive precursors, most significantly a reduction in the number of ST-HSCs and LMPPs and a decrease of leukocyte numbers, effects that are exacerbated with age. Cell cycle and gene expression analyses suggest that the main hematopoietic defect in Msi2 defective mice consists in a decreased proliferation capacity of ST-HSCs and LMPPs. In addition, HSCs lacking Msi2 are severely impaired in competitive repopulation experiments, being overgrown by wild type cells even when mutant cells were provided in excess. Our data indicate that Msi2 maintains the stem cell compartment mainly by regulating the proliferation of primitive progenitors downstream of LT-HSCs. RNA from 2 different cell populations (LSK cells and KIT+ cells) were analyzed for both wildtype and Msi2-/- null samples. Two technical replicates were analyzed for the KIT+ samples and three technical replicates were analyzed for the LSK samples.
Project description:We demonstrate that Msi2 is the predominant form expressed in hematopoietic stem cells (HSC), and its knockdown leads to reduced engraftment and depletion of HSCs in vivo. Overexpression of Msi2 in a mouse model increases HSC cell cycle progression and cooperates with BCR-ABL1 to induce an aggressive leukemia. MSI2 is over-expressed in human myeloid leukemia, and expression levels directly correlate with decreased patient survival, thereby defining MSI2 expression as a novel prognostic marker in acute myeloid leukemia (AML). Depletion of MSI2 in human myeloid leukemia cells leads to decreased proliferation and apoptosis. These data implicate the MSI2 RNA binding protein in myeloid leukemogenesis and identify a novel potential target for therapy in AML. Mouse hematopoietic stem cells (defined as LineageLow, Sca1+, Kit+; LSK; n=3) as well as committed progenitor cells (LineageLow, Sca1-, Kit+; LK; n=3) were sorted and RNA was extracted using Qiagen RNeasy Micro Kit according to manufacturers instruction. cDNA was fragmented and biotinylated before hybridization onto Affymetrix Mouse Expression Array 430 2.0.
Project description:To understand the mechanisms through which JunB regulates Tregs-mediated immune regulation, we examined the global gene expression profiles in the JunB WT and KO Tregs by performing RNA sequencing (RNA-seq) analysis.