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Anti-apoptotic function of Xbp1 as an IL-3 signaling molecule in hematopoietic cells.

Kurata M, Yamazaki Y, Kanno Y, Ishibashi S, Takahara T, Kitagawa M, Nakamura T - Cell Death Dis (2011)

Bottom Line: Inhibition of IL-3 signaling as well as knockdown of Xbp1-induced apoptosis in BaF3 cells.Expression of apoptosis-, cell cycle- and differentiation-related genes was modulated by Xbp1S expression.These results indicate that the proper transcriptional and splicing regulation of Xbp1 by IL-3 signaling is important in homeostasis of hematopoietic cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Carcinogenesis, The Cancer Institute, Japanese Foundation for Cancer Research, 3-8-31 Ariake, Koto-ku, Tokyo 135-8550, Japan.

ABSTRACT
Cytokine signaling is critical for proliferation, survival and differentiation of hematopoietic cell, and interleukin-3 (IL-3) is required for maintenance of many hematopoietic cell lines, such as BaF3. We have isolated apoptosis-resistant clones of BaF3 using retroviral insertional mutagenesis and the Xbp1 locus was identified as a retroviral integration site. Expression and splicing of the Xbp1 transcript was conserved in the resistant clone but was promptly disappeared on IL-3 withdrawal in parental BaF3. IL-3 stimulation of BaF3 cells enhanced Xbp1 promoter activity and induced phosphorylation of the endoplasmic reticulum stress sensor protein IRE1, resulting in the increase in Xbp1S that activates unfolded protein response. When downstream signaling from IL-3 was blocked by LY294002 and/or dn-Stat5, Xbp1 expression was downregulated and IRE1 phosphorylation was suppressed. Inhibition of IL-3 signaling as well as knockdown of Xbp1-induced apoptosis in BaF3 cells. In contrast, constitutive expression of Xbp1S protected BaF3 from apoptosis during IL-3 depletion. However, cell cycle arrest at the G1 stage was observed in BaF3 and myeloid differentiation was induced in IL-3-dependent 32Dcl3 cells. Expression of apoptosis-, cell cycle- and differentiation-related genes was modulated by Xbp1S expression. These results indicate that the proper transcriptional and splicing regulation of Xbp1 by IL-3 signaling is important in homeostasis of hematopoietic cells.

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Xbp1S modulates cell cycle and differentiation. (a) Cell cycle arrest was induced in BaF3 cells by Xbp1S expression. Proportions of cells at G0/G1 and G2/M phases are indicated. (b) A list of downregulated genes related to cell cycle progression in Xbp1S transduced 32Dcl3 cells. (c) Wright–Giemsa-staining of cytospin samples. 32Dcl3 cells show granulocytic differentiation by Xbp1S expression, while 32Dcl3 cells transduced with pMIG or pMYs-Xbp1U remain immature. (d) Increased intensity of Mac-1 expression in 32Dcl3 cells by GFP-positive population with using pMY-Xbp1S-GFP vector (**P<0.05). (e) A list of upregulated genes associated with myeloid cell differentiation
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fig6: Xbp1S modulates cell cycle and differentiation. (a) Cell cycle arrest was induced in BaF3 cells by Xbp1S expression. Proportions of cells at G0/G1 and G2/M phases are indicated. (b) A list of downregulated genes related to cell cycle progression in Xbp1S transduced 32Dcl3 cells. (c) Wright–Giemsa-staining of cytospin samples. 32Dcl3 cells show granulocytic differentiation by Xbp1S expression, while 32Dcl3 cells transduced with pMIG or pMYs-Xbp1U remain immature. (d) Increased intensity of Mac-1 expression in 32Dcl3 cells by GFP-positive population with using pMY-Xbp1S-GFP vector (**P<0.05). (e) A list of upregulated genes associated with myeloid cell differentiation

Mentions: Flowcytometric analyses revealed that Xbp1S-expressing BaF3 cells showed 1.7-fold increase in the G1 phase population and 40% reduction of the G2 population (Figure 6a), indicating that constitutive expression of Xbp1S induces cell cycle arrest at the G1 phase. Downregulation of cell cycle-related genes was evident in Xbp1S-expressing 32Dcl3 cells compared with those transduced with an empty vector or Xbp1U retrovirus. Significant downregulation of seven cyclin genes as well as six members of the Mcm gene family and four Cdc-related genes was observed (Figure 6b).


Anti-apoptotic function of Xbp1 as an IL-3 signaling molecule in hematopoietic cells.

Kurata M, Yamazaki Y, Kanno Y, Ishibashi S, Takahara T, Kitagawa M, Nakamura T - Cell Death Dis (2011)

Xbp1S modulates cell cycle and differentiation. (a) Cell cycle arrest was induced in BaF3 cells by Xbp1S expression. Proportions of cells at G0/G1 and G2/M phases are indicated. (b) A list of downregulated genes related to cell cycle progression in Xbp1S transduced 32Dcl3 cells. (c) Wright–Giemsa-staining of cytospin samples. 32Dcl3 cells show granulocytic differentiation by Xbp1S expression, while 32Dcl3 cells transduced with pMIG or pMYs-Xbp1U remain immature. (d) Increased intensity of Mac-1 expression in 32Dcl3 cells by GFP-positive population with using pMY-Xbp1S-GFP vector (**P<0.05). (e) A list of upregulated genes associated with myeloid cell differentiation
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101701&req=5

fig6: Xbp1S modulates cell cycle and differentiation. (a) Cell cycle arrest was induced in BaF3 cells by Xbp1S expression. Proportions of cells at G0/G1 and G2/M phases are indicated. (b) A list of downregulated genes related to cell cycle progression in Xbp1S transduced 32Dcl3 cells. (c) Wright–Giemsa-staining of cytospin samples. 32Dcl3 cells show granulocytic differentiation by Xbp1S expression, while 32Dcl3 cells transduced with pMIG or pMYs-Xbp1U remain immature. (d) Increased intensity of Mac-1 expression in 32Dcl3 cells by GFP-positive population with using pMY-Xbp1S-GFP vector (**P<0.05). (e) A list of upregulated genes associated with myeloid cell differentiation
Mentions: Flowcytometric analyses revealed that Xbp1S-expressing BaF3 cells showed 1.7-fold increase in the G1 phase population and 40% reduction of the G2 population (Figure 6a), indicating that constitutive expression of Xbp1S induces cell cycle arrest at the G1 phase. Downregulation of cell cycle-related genes was evident in Xbp1S-expressing 32Dcl3 cells compared with those transduced with an empty vector or Xbp1U retrovirus. Significant downregulation of seven cyclin genes as well as six members of the Mcm gene family and four Cdc-related genes was observed (Figure 6b).

Bottom Line: Inhibition of IL-3 signaling as well as knockdown of Xbp1-induced apoptosis in BaF3 cells.Expression of apoptosis-, cell cycle- and differentiation-related genes was modulated by Xbp1S expression.These results indicate that the proper transcriptional and splicing regulation of Xbp1 by IL-3 signaling is important in homeostasis of hematopoietic cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Carcinogenesis, The Cancer Institute, Japanese Foundation for Cancer Research, 3-8-31 Ariake, Koto-ku, Tokyo 135-8550, Japan.

ABSTRACT
Cytokine signaling is critical for proliferation, survival and differentiation of hematopoietic cell, and interleukin-3 (IL-3) is required for maintenance of many hematopoietic cell lines, such as BaF3. We have isolated apoptosis-resistant clones of BaF3 using retroviral insertional mutagenesis and the Xbp1 locus was identified as a retroviral integration site. Expression and splicing of the Xbp1 transcript was conserved in the resistant clone but was promptly disappeared on IL-3 withdrawal in parental BaF3. IL-3 stimulation of BaF3 cells enhanced Xbp1 promoter activity and induced phosphorylation of the endoplasmic reticulum stress sensor protein IRE1, resulting in the increase in Xbp1S that activates unfolded protein response. When downstream signaling from IL-3 was blocked by LY294002 and/or dn-Stat5, Xbp1 expression was downregulated and IRE1 phosphorylation was suppressed. Inhibition of IL-3 signaling as well as knockdown of Xbp1-induced apoptosis in BaF3 cells. In contrast, constitutive expression of Xbp1S protected BaF3 from apoptosis during IL-3 depletion. However, cell cycle arrest at the G1 stage was observed in BaF3 and myeloid differentiation was induced in IL-3-dependent 32Dcl3 cells. Expression of apoptosis-, cell cycle- and differentiation-related genes was modulated by Xbp1S expression. These results indicate that the proper transcriptional and splicing regulation of Xbp1 by IL-3 signaling is important in homeostasis of hematopoietic cells.

Show MeSH
Related in: MedlinePlus