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Identification of a cytokine-induced antiapoptotic molecule anamorsin essential for definitive hematopoiesis.

Shibayama H, Takai E, Matsumura I, Kouno M, Morii E, Kitamura Y, Takeda J, Kanakura Y - J. Exp. Med. (2004)

Bottom Line: Using an expression cloning method, we identified a novel antiapoptotic molecule named Anamorsin, which does not show any homology to known apoptosis regulatory molecules such as Bcl-2 family, caspase family, or signal transduction molecules.Also, Anamorsin-/- erythroid cells initiated apoptosis during terminal maturation.Thus, Anamorsin is considered to be a necessary molecule for hematopoiesis that mediates antiapoptotic effects of various cytokines.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology and Oncology, Osaka University Graduate School of Medicine, Yamada-oka, Suita, 565-0871, Japan.

ABSTRACT
Many growth factors and cytokines prevent apoptosis. Using an expression cloning method, we identified a novel antiapoptotic molecule named Anamorsin, which does not show any homology to known apoptosis regulatory molecules such as Bcl-2 family, caspase family, or signal transduction molecules. The expression of Anamorsin was completely dependent on stimulation with growth factors such as interleukin 3, stem cell factor, and thrombopoietin in factor-dependent hematopoietic cell lines, and forced expression of Anamorsin conferred resistance to apoptosis caused by growth factor deprivation in vitro. Furthermore, Anamorsin was found to act as an antiapoptotic molecule in vivo because Anamorsin-/- mice die in late gestation due to defective definitive hematopoiesis in the fetal liver (FL). Although the number of hematopoietic stem/progenitor cells in the FL did not decrease in these mice, myeloid, and particularly erythroid colony formation in response to cytokines, was severely disrupted. Also, Anamorsin-/- erythroid cells initiated apoptosis during terminal maturation. As for the mechanism of Anamorsin-mediated cell survival, a microarray analysis revealed that the expression of Bcl-xL and Jak2 was severely impaired in the FL of Anamorsin-/- mice. Thus, Anamorsin is considered to be a necessary molecule for hematopoiesis that mediates antiapoptotic effects of various cytokines.

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RT-PCR analysis on the expression of Jak2 and Bcl-xL mRNA in Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. Total RNA was isolated from E14.5 Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. 1.5 μg total RNA was reverse transcribed to first strand cDNA and subjected to PCR reactions. After the indicated cycles of PCR reaction, PCR products were electrophoresed on agarose gels and visualized with ethidium bromide staining.
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fig6: RT-PCR analysis on the expression of Jak2 and Bcl-xL mRNA in Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. Total RNA was isolated from E14.5 Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. 1.5 μg total RNA was reverse transcribed to first strand cDNA and subjected to PCR reactions. After the indicated cycles of PCR reaction, PCR products were electrophoresed on agarose gels and visualized with ethidium bromide staining.

Mentions: To characterize the antiapoptotic function of Anamorsin, we compared gene expression profiles between Anamorsin−/− and Anamorsin+/+ FL cells at E14.5 using a cDNA microarray. Among 4289 genes, including Bcl-2 family, caspases, cytokines, and signal transduction molecules, 184 genes were significantly down-regulated and 40 were up-regulated in Anamorsin−/− FL. Among apoptosis-related genes, Bcl-xL and Jak2 were down-regulated most significantly. We also confirmed that expression of these two molecules was decreased in Anamorsin−/− FL cells compared with Anamorsin+/+ FL cells by semiquantitative RT-PCR assays (Fig. 6 and Table S1, available at http://www.jem.org/cgi/content/full/jem.20031858/DC1).


Identification of a cytokine-induced antiapoptotic molecule anamorsin essential for definitive hematopoiesis.

Shibayama H, Takai E, Matsumura I, Kouno M, Morii E, Kitamura Y, Takeda J, Kanakura Y - J. Exp. Med. (2004)

RT-PCR analysis on the expression of Jak2 and Bcl-xL mRNA in Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. Total RNA was isolated from E14.5 Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. 1.5 μg total RNA was reverse transcribed to first strand cDNA and subjected to PCR reactions. After the indicated cycles of PCR reaction, PCR products were electrophoresed on agarose gels and visualized with ethidium bromide staining.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211823&req=5

fig6: RT-PCR analysis on the expression of Jak2 and Bcl-xL mRNA in Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. Total RNA was isolated from E14.5 Anamorsin−/−, Anamorsin+/−, and Anamorsin+/+ FL cells. 1.5 μg total RNA was reverse transcribed to first strand cDNA and subjected to PCR reactions. After the indicated cycles of PCR reaction, PCR products were electrophoresed on agarose gels and visualized with ethidium bromide staining.
Mentions: To characterize the antiapoptotic function of Anamorsin, we compared gene expression profiles between Anamorsin−/− and Anamorsin+/+ FL cells at E14.5 using a cDNA microarray. Among 4289 genes, including Bcl-2 family, caspases, cytokines, and signal transduction molecules, 184 genes were significantly down-regulated and 40 were up-regulated in Anamorsin−/− FL. Among apoptosis-related genes, Bcl-xL and Jak2 were down-regulated most significantly. We also confirmed that expression of these two molecules was decreased in Anamorsin−/− FL cells compared with Anamorsin+/+ FL cells by semiquantitative RT-PCR assays (Fig. 6 and Table S1, available at http://www.jem.org/cgi/content/full/jem.20031858/DC1).

Bottom Line: Using an expression cloning method, we identified a novel antiapoptotic molecule named Anamorsin, which does not show any homology to known apoptosis regulatory molecules such as Bcl-2 family, caspase family, or signal transduction molecules.Also, Anamorsin-/- erythroid cells initiated apoptosis during terminal maturation.Thus, Anamorsin is considered to be a necessary molecule for hematopoiesis that mediates antiapoptotic effects of various cytokines.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology and Oncology, Osaka University Graduate School of Medicine, Yamada-oka, Suita, 565-0871, Japan.

ABSTRACT
Many growth factors and cytokines prevent apoptosis. Using an expression cloning method, we identified a novel antiapoptotic molecule named Anamorsin, which does not show any homology to known apoptosis regulatory molecules such as Bcl-2 family, caspase family, or signal transduction molecules. The expression of Anamorsin was completely dependent on stimulation with growth factors such as interleukin 3, stem cell factor, and thrombopoietin in factor-dependent hematopoietic cell lines, and forced expression of Anamorsin conferred resistance to apoptosis caused by growth factor deprivation in vitro. Furthermore, Anamorsin was found to act as an antiapoptotic molecule in vivo because Anamorsin-/- mice die in late gestation due to defective definitive hematopoiesis in the fetal liver (FL). Although the number of hematopoietic stem/progenitor cells in the FL did not decrease in these mice, myeloid, and particularly erythroid colony formation in response to cytokines, was severely disrupted. Also, Anamorsin-/- erythroid cells initiated apoptosis during terminal maturation. As for the mechanism of Anamorsin-mediated cell survival, a microarray analysis revealed that the expression of Bcl-xL and Jak2 was severely impaired in the FL of Anamorsin-/- mice. Thus, Anamorsin is considered to be a necessary molecule for hematopoiesis that mediates antiapoptotic effects of various cytokines.

Show MeSH