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Hematopoietic stem cells lacking Ott1 display aspects associated with aging and are unable to maintain quiescence during proliferative stress.

ABSTRACT: Aging degrades hematopoietic stem cell (HSC) functions, including stress response; however, the involved molecular pathways are incompletely defined. Murine BM conditionally deleted for One-Twenty-Two-1 (Ott1), is able to maintain lifelong hematopoiesis and has preserved numbers of long-term HSCs, yet cannot repopulate nor sustain itself after transplantation against a competitor even when Ott1 is excised after engraftment. We show, specifically under replicative stress, that Ott1-deleted HSCs have a significant reduction of the G(0) cell-cycle fraction associated with self-renewal and undergo early failure. Therefore, Ott1 is required to preserve HSC quiescence during stress but not steady-state hematopoiesis. Reduced tolerance of replicative stress, increased myeloid potential, and greater absolute numbers are mutual characteristics of both Ott1-deleted and aged HSCs, and comparison of their gene expression profiles reveals a shared signature. Ott1-deleted HSCs share multiple aging-associated physiologic changes, including increases in NF-?B activation and DNA damage. Loss of Ott1 causes increased reactive oxygen species; however, antioxidant treatment does not rescue the competitive defect, indicating the existence of additional essential Ott1-dependent HSC pathways. In conclusion, our data establish a requirement for Ott1 in stress hematopoiesis and suggest that Ott1-dependent processes may converge with those affected by aging.

SUBMITTER: Xiao N

PROVIDER: S-EPMC3367894 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Hematopoietic stem cells lacking Ott1 display aspects associated with aging and are unable to maintain quiescence during proliferative stress.

Xiao Nan N Jani Kaushal K Morgan Kelly K Okabe Rachel R Cullen Dana E DE Jesneck Jonathan L JL Raffel Glen D GD

Blood 20120406 21

Aging degrades hematopoietic stem cell (HSC) functions, including stress response; however, the involved molecular pathways are incompletely defined. Murine BM conditionally deleted for One-Twenty-Two-1 (Ott1), is able to maintain lifelong hematopoiesis and has preserved numbers of long-term HSCs, yet cannot repopulate nor sustain itself after transplantation against a competitor even when Ott1 is excised after engraftment. We show, specifically under replicative stress, that Ott1-deleted HSCs h ...[more]

PMID: 22490678

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Project description:The infant leukemia-associated gene, Ott1 (Rbm15), has broad regulatory effects on embryonic development and hematopoiesis. Embryonic deletion of Ott1 results in defects to the placenta, spleen and heart. Conditional deletion within the adult hematopoietic compartment demonstrates a requirement in pre-B development and inhibitory roles in myeloid progenitor and megakaryocyte populations. Ott1-deleted bone marrow has an expansion of the Lin- Sca-1+ c-Kit+ (LSK) population which includes the hematopoietic stem cell (HSC) population. Functional HSC testing through competitive repopulation of irradiated recipients demonstrated however, a severe defect in Ott1-deficient HSCs, despite adequate numbers of immunophenotypically identified long term HSCs. Although mice deleted in situ for Ott1 are able to maintain hematopoiesis in steady state over a normal lifetime, but when subjected to proliferative stress, the HSC population loses the self-renewing, G0 fraction and undergoes bone marrow failure. Therefore, Ott1 is required for maintaining HSC quiescence during proliferative stress. To investigate the downstream effects of Ott1 loss, LSK cells were sorted from Ott1-deleted mice and controls then global gene expression profiles were generated through hybridization of total RNA onto microarrays. Gene set enrichment analysis showed significant enrichment with genes altered in aged HSCs. Ott1-deficient HSCs were found to have other aging-associated features such as increased DNA damage, NF kappa B activation and p38Mapk activation, suggesting Ott1 controls pathways also involved in the aging process. Six week old Ott1 flox/null Tg-Mx1cre mice and littermate controls possessing a wild type Ott1 allele were injected with 250 mcg intraperitoneal pIpC every other day for 3 doses. Six weeks post-pIpC, bone marrow was harvested, labeled with lineage markers (B220, CD19, CD4, CD8, CD3, IL7R, Terr119 and Gr-1) and depleted using magnetic beads. The remaining cells were labeled for c-Kit and Sca-1 and the Lin- Sca-1+ c-Kit+ (LSK) population was sorted using a FACSAria cell sorter (BD Biosciences). Approximately 10,000 LSK cells per mouse were isolated from 3 Ott1 flox/null Tg-Mx1cre mice and 3 controls. RNA was extracted using the RNeasy Micro kit (Qiagen, Valencia, CA) and treated with RNAse-free DNAse (Qiagen, Valencia, CA). 50 ng of purified total RNA was linearly amplified using the Ovation RNA amplification system V2 (Nugen, San Carlos, CA) and biotinylated with the FL-Ovation Biotin Module V2 (Nugen, San Carlos, CA) per the supplied protocol. cDNA was then hybridized to Affymetrix Mouse expression array 430A2.0 chips by the Dana Farber Microarray Core Facility (Boston, MA). The raw gene expression values were preprocessed with robust multi-array analysis (RMA) algorithm using BioConductor software.

Dataset Information

Hematopoietic stem cells lacking Ott1 display aspects associated with aging and are unable to maintain quiescence during proliferative stress.

Publications

Hematopoietic stem cells lacking Ott1 display aspects associated with aging and are unable to maintain quiescence during proliferative stress.

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