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XBP1: a link between the unfolded protein response, lipid biosynthesis, and biogenesis of the endoplasmic reticulum.

Sriburi R, Jackowski S, Mori K, Brewer JW - J. Cell Biol. (2004)

Bottom Line: When the protein folding capacity of the endoplasmic reticulum (ER) is challenged, the unfolded protein response (UPR) maintains ER homeostasis by regulating protein synthesis and enhancing expression of resident ER proteins that facilitate protein maturation and degradation.Cells overexpressing XBP1(S) exhibit elevated levels of membrane phospholipids, increased surface area and volume of rough ER, and enhanced activity of the cytidine diphosphocholine pathway of phosphatidylcholine biosynthesis.These data suggest that XBP1(S) links the mammalian UPR to phospholipid biosynthesis and ER biogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University Chicago, Maywood, IL 60153, USA.

ABSTRACT
When the protein folding capacity of the endoplasmic reticulum (ER) is challenged, the unfolded protein response (UPR) maintains ER homeostasis by regulating protein synthesis and enhancing expression of resident ER proteins that facilitate protein maturation and degradation. Here, we report that enforced expression of XBP1(S), the active form of the XBP1 transcription factor generated by UPR-mediated splicing of XBP1 mRNA, is sufficient to induce synthesis of phosphatidylcholine, the primary phospholipid of the ER membrane. Cells overexpressing XBP1(S) exhibit elevated levels of membrane phospholipids, increased surface area and volume of rough ER, and enhanced activity of the cytidine diphosphocholine pathway of phosphatidylcholine biosynthesis. These data suggest that XBP1(S) links the mammalian UPR to phospholipid biosynthesis and ER biogenesis.

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Microscopy analysis of the ER. (A) At 48 h after transduction with the indicated retroviral vectors, thin sections were prepared from NIH-3T3 cells and examined by transmission EM. Representative micrographs of increasing magnification (from top to bottom) are shown with scale bar (1 μm) in top right corner: middle panels, arrow indicates representative ER; bottom panels, arrow indicates membrane-bound ribosomes. N, nucleus. (B) Stereological analysis of RER volume (top) and RER surface area (bottom) was performed on electron micrographs and the mean ± SEM are plotted (n = 4 stereological sets with 2–16 micrographs per set; *, P < 0.01). (C) Immunofluorescence analysis (using anti-KDEL mAb) and confocal microscopy was performed on NIH-3T3 cells grown on coverslips and transduced with the indicated retroviral vectors for 48 h.
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fig2: Microscopy analysis of the ER. (A) At 48 h after transduction with the indicated retroviral vectors, thin sections were prepared from NIH-3T3 cells and examined by transmission EM. Representative micrographs of increasing magnification (from top to bottom) are shown with scale bar (1 μm) in top right corner: middle panels, arrow indicates representative ER; bottom panels, arrow indicates membrane-bound ribosomes. N, nucleus. (B) Stereological analysis of RER volume (top) and RER surface area (bottom) was performed on electron micrographs and the mean ± SEM are plotted (n = 4 stereological sets with 2–16 micrographs per set; *, P < 0.01). (C) Immunofluorescence analysis (using anti-KDEL mAb) and confocal microscopy was performed on NIH-3T3 cells grown on coverslips and transduced with the indicated retroviral vectors for 48 h.

Mentions: Electron microscopy revealed that XBP1(S)-transduced cells contained an increased number of intracellular membrane–bound structures that were frequently studded with ribosomes, indicating their identity as RER (Fig. 2 A). Indeed, the surface area and volume of RER were significantly increased in XBP1(S)-transduced cells, 2.5- and 3.4-fold, respectively (Fig. 2 B). Immunofluorescence microscopy demonstrated that KDEL-containing proteins were distributed throughout the cytoplasm of XBP1(S)-transduced cells (Fig. 2 C), consistent with an overall increase in the amount of ER. In addition, XBP1(S) induced a 44% increase in cell size (empty vector cells, 103.9 ± 1.8 μm2; XBP1(S) cells, 149.8 ± 2.5 μm2). Cells overexpressing XBP1(S) exhibited a slower rate of proliferation 24 h after transduction, but their viability was not affected through 48 h. Importantly, while this manuscript was under review, Shaffer et al. (2004) reported that XBP1(S) can mediate increased cell size and expansion of intracellular organelles including the ER, thereby corroborating our findings.


XBP1: a link between the unfolded protein response, lipid biosynthesis, and biogenesis of the endoplasmic reticulum.

Sriburi R, Jackowski S, Mori K, Brewer JW - J. Cell Biol. (2004)

Microscopy analysis of the ER. (A) At 48 h after transduction with the indicated retroviral vectors, thin sections were prepared from NIH-3T3 cells and examined by transmission EM. Representative micrographs of increasing magnification (from top to bottom) are shown with scale bar (1 μm) in top right corner: middle panels, arrow indicates representative ER; bottom panels, arrow indicates membrane-bound ribosomes. N, nucleus. (B) Stereological analysis of RER volume (top) and RER surface area (bottom) was performed on electron micrographs and the mean ± SEM are plotted (n = 4 stereological sets with 2–16 micrographs per set; *, P < 0.01). (C) Immunofluorescence analysis (using anti-KDEL mAb) and confocal microscopy was performed on NIH-3T3 cells grown on coverslips and transduced with the indicated retroviral vectors for 48 h.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2172532&req=5

fig2: Microscopy analysis of the ER. (A) At 48 h after transduction with the indicated retroviral vectors, thin sections were prepared from NIH-3T3 cells and examined by transmission EM. Representative micrographs of increasing magnification (from top to bottom) are shown with scale bar (1 μm) in top right corner: middle panels, arrow indicates representative ER; bottom panels, arrow indicates membrane-bound ribosomes. N, nucleus. (B) Stereological analysis of RER volume (top) and RER surface area (bottom) was performed on electron micrographs and the mean ± SEM are plotted (n = 4 stereological sets with 2–16 micrographs per set; *, P < 0.01). (C) Immunofluorescence analysis (using anti-KDEL mAb) and confocal microscopy was performed on NIH-3T3 cells grown on coverslips and transduced with the indicated retroviral vectors for 48 h.
Mentions: Electron microscopy revealed that XBP1(S)-transduced cells contained an increased number of intracellular membrane–bound structures that were frequently studded with ribosomes, indicating their identity as RER (Fig. 2 A). Indeed, the surface area and volume of RER were significantly increased in XBP1(S)-transduced cells, 2.5- and 3.4-fold, respectively (Fig. 2 B). Immunofluorescence microscopy demonstrated that KDEL-containing proteins were distributed throughout the cytoplasm of XBP1(S)-transduced cells (Fig. 2 C), consistent with an overall increase in the amount of ER. In addition, XBP1(S) induced a 44% increase in cell size (empty vector cells, 103.9 ± 1.8 μm2; XBP1(S) cells, 149.8 ± 2.5 μm2). Cells overexpressing XBP1(S) exhibited a slower rate of proliferation 24 h after transduction, but their viability was not affected through 48 h. Importantly, while this manuscript was under review, Shaffer et al. (2004) reported that XBP1(S) can mediate increased cell size and expansion of intracellular organelles including the ER, thereby corroborating our findings.

Bottom Line: When the protein folding capacity of the endoplasmic reticulum (ER) is challenged, the unfolded protein response (UPR) maintains ER homeostasis by regulating protein synthesis and enhancing expression of resident ER proteins that facilitate protein maturation and degradation.Cells overexpressing XBP1(S) exhibit elevated levels of membrane phospholipids, increased surface area and volume of rough ER, and enhanced activity of the cytidine diphosphocholine pathway of phosphatidylcholine biosynthesis.These data suggest that XBP1(S) links the mammalian UPR to phospholipid biosynthesis and ER biogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University Chicago, Maywood, IL 60153, USA.

ABSTRACT
When the protein folding capacity of the endoplasmic reticulum (ER) is challenged, the unfolded protein response (UPR) maintains ER homeostasis by regulating protein synthesis and enhancing expression of resident ER proteins that facilitate protein maturation and degradation. Here, we report that enforced expression of XBP1(S), the active form of the XBP1 transcription factor generated by UPR-mediated splicing of XBP1 mRNA, is sufficient to induce synthesis of phosphatidylcholine, the primary phospholipid of the ER membrane. Cells overexpressing XBP1(S) exhibit elevated levels of membrane phospholipids, increased surface area and volume of rough ER, and enhanced activity of the cytidine diphosphocholine pathway of phosphatidylcholine biosynthesis. These data suggest that XBP1(S) links the mammalian UPR to phospholipid biosynthesis and ER biogenesis.

Show MeSH
Related in: MedlinePlus