Project description:Long-term hematopoietic output is dependent on the hematopoietic stem cell (HSC) homeostasis which is maintained by a complex network of molecules. Among these, microRNAs (miRNAs) play crucial roles, while the underlying molecular basis have not been fully demonstrated. Here, we found that miR-21 is enriched in murine HSCs. Then, we generated a polyinosinic:polycytidylic acid (pIpC)-inducible mouse model (miR-21flox/flox:Mx1-Cre) to obtain a specific deletion of miR-21 in hematopoietic system. It was found that mice with conditional knockout of miR-21 exhibit an obvious perturbation of normal hematopoiesis.Further researches reveal that miR-21 deficiency affect HSC homeostasis and function. We used microarrays to detail the global programme of gene expression, and identified distinct classes of up-regulated and down-regulated genes in murine HSCs after miR-21 conditional knockout.
Project description:MicroRNAs influence hematopoietic differentiation, but little is known about their effects on the stem cell state. Here, we report that the microRNA processing enzyme Dicer is essential for stem cell persistence in vivo and a specific microRNA, miR-125a controls the size of the stem cell population by regulating stem/progenitor cell (HSPC) apoptosis. Conditional deletion of Dicer revealed an absolute dependence for the multipotent HSPC population in a cell autonomous manner, with increased HSPC apoptosis in mutant animals. An evolutionarily conserved microRNA cluster containing miR-99b, let-7e and miR-125a was preferentially expressed in long term HSCs. miR-125a alone was capable of increasing the number of hematopoietic stem cells in vivo by more than eight fold. This was accomplished through a differentiation stage-specific reduction of apoptosis in immature hematopoietic progenitors, possibly through targeting multiple pro-apoptotic genes. Bak1 was directly down-regulated by miR-125a and expression of a 3’UTR-less Bak1 blocked miR-125a-induced hematopoietic expansion in vivo. These data demonstrate cell-state-specific regulation by microRNA and identify a unique microRNA functioning to regulate the stem cell pool size. Bone marrow populations were FACS-sorted and profiled using a bead-based profiling platform. Long-term HSCs, short-term HSCs, multipotent progenitors, Lin-Kit+Sca+ cells, Lin-Kit+Sca- cells, Lin-Kit-Sca+ cells, Lin- cells and unfractionated whole bone marrow cells were prepared for total RNA using TriZol (Invitrogen) in replicates. For rare populations, cells from multiple mice were pooled. To perform microRNA profiling, 60 ng of total RNA were used for each sample.
Project description:MicroRNAs influence hematopoietic differentiation, but little is known about their effects on the stem cell state. Here, we report that the microRNA processing enzyme Dicer is essential for stem cell persistence in vivo and a specific microRNA, miR-125a controls the size of the stem cell population by regulating stem/progenitor cell (HSPC) apoptosis. Conditional deletion of Dicer revealed an absolute dependence for the multipotent HSPC population in a cell autonomous manner, with increased HSPC apoptosis in mutant animals. An evolutionarily conserved microRNA cluster containing miR-99b, let-7e and miR-125a was preferentially expressed in long term HSCs. miR-125a alone was capable of increasing the number of hematopoietic stem cells in vivo by more than eight fold. This was accomplished through a differentiation stage-specific reduction of apoptosis in immature hematopoietic progenitors, possibly through targeting multiple pro-apoptotic genes. Bak1 was directly down-regulated by miR-125a and expression of a 3’UTR-less Bak1 blocked miR-125a-induced hematopoietic expansion in vivo. These data demonstrate cell-state-specific regulation by microRNA and identify a unique microRNA functioning to regulate the stem cell pool size.