Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse spleens of wild type mice and mutants lacking Nix, a proapoptotic BH3-only factor that is transcriptionally induced during erythropoiesis

ABSTRACT: Normal erythropoiesis requires a critical balance between proapoptotic and antipaoptotic pathways. Bcl-xl, an antiapoptotic protein is induced at end-stages of differentiation of erythroid precursors in response to erythropoietin. The details of the proapoptotic pathway and the critical proapoptotic proteins inhibited by Bcl-xl in erythropoiesis are not well understood. We employed gene targeting to ablate Nix, a proapoptotic BH3-domain only Bcl2 family protein, which is known to be transcriptionally induced during erythropoiesis. Nix null mice exhibited reticulocytosis and thrombocytosis in the peripheral blood; and profound splenomegaly with erythroblastosis in the spleen and bone marrow despite normal erythropoietin levels and blood oxygen tension. In vivo apoptosis was diminished in erythroblast precursors from Nix null spleens. To define the molecular consequences of Nix ablation on apoptosis and erythropoiesis, we conducted a detailed comparative analysis of gene expression in spleens from 8 week old Nix null mice and wild type controls. Of 45,101 genes analyzed, 514 were significantly upregulated and 386 down-regulated in Nix-/- splenocytes. Functional cluster analysis delineated the ten most highly regulated gene sets, revealing increased levels of cell cycle and erythroid genes, with decreased levels of cell death and B-cell genes. Experiment Overall Design: In the study, we hybridized RNA from 8 week old spleens of wild type (WT) control and Nix null mice (Nix-/-) to Affymetrix MOE430A GeneChipÂ® arrays containing 45,101 well characterized mouse genes/ESTs.

ORGANISM(S): Mus musculus

SUBMITTER: Bruce Aronow

PROVIDER: E-GEOD-7020 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Unrestrained erythroblast development in Nix-/- mice reveals a mechanism for apoptotic modulation of erythropoiesis.

Diwan Abhinav A Koesters Andrew G AG Odley Amy M AM Pushkaran Suvarnamala S Baines Christopher P CP Spike Benjamin T BT Daria Diedre D Jegga Anil G AG Geiger Hartmut H Aronow Bruce J BJ Molkentin Jeffery D JD Macleod Kay F KF Kalfa Theodosia A TA Dorn Gerald W GW

Proceedings of the National Academy of Sciences of the United States of America 20070409 16

Normal production of RBCs requires that the antiapoptotic protein Bcl-xl be induced at end stages of differentiation in response to erythropoietin (Epo) signaling. The critical proapoptotic pathways inhibited by Bcl-xl in erythroblasts are unknown. We used gene targeting in the mouse to evaluate the BH3-only factor Nix, which is transcriptionally up-regulated during Epo-stimulated in vitro erythrocyte differentiation. Nix null mice are viable and fertile. Peripheral blood counts revealed a profo ...[more]

PMID: 17420462

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Project description:Understanding the pattern of gene expression and identifying the specific genes expressed during erythropoiesis is crucial for a synthesis of erythroid developmental biology. Here we have isolated four distinct populations of erythroblasts at successive erythropoietin-dependent stages of erythropoiesis including the terminal, pyknotic stage. The transcriptome has been determined using Affymetrix arrays. First, we show that cells sorted by surface expression profile express not only significantly fewer genes than unsorted cells, but also significantly more differences in the expression levels of particular genes between stages than unsorted cells, demonstrating the importance of working with defined cell populations to identify lineage and temporally-specific patterns of gene expression. Second, using standard software and matched filtering we identify eleven differentially regulated genes and one continuously expressed gene previously undetected in erythroid expression studies with unknown roles in erythropoiesis (CA3, CALB1, CTSL2, FKBP1B, GSDMB, ITLN1, LIN7B, RRAD, RUNDC3A, UNQ1887, ZNF805, MYL12B). Finally, using transcription factor binding site analysis we identify potential transcription factors that may regulate gene expression during terminal erythropoiesis. Our stringent lists of differentially regulated and continuously expressed transcripts are a resource for functional studies of erythropoietic protein function and gene regulation. PBMC-derived erythroblasts grown in vitro were harvested at the CFU-E, Pro-E, Int-E and Late-E stages and FACS sorted based on expression of cell surface markers. RNA was extracted from those populations which met rigorous specifications of time cultured in the presence of erythropoietin, morphology and CD36, CD71 and CD235a expression. For each stage, 3 chips were hybridised, each representing a pool of 4 different samples. In addition to these 12 chips, one chip representing a pool of RNA extracted from unsorted cells at each stages was also hybridised.

Dataset Information

Transcription profiling of mouse spleens of wild type mice and mutants lacking Nix, a proapoptotic BH3-only factor that is transcriptionally induced during erythropoiesis

Publications

Unrestrained erythroblast development in Nix-/- mice reveals a mechanism for apoptotic modulation of erythropoiesis.

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