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Global defects in collagen secretion in a Mia3/TANGO1 knockout mouse.

Wilson DG, Phamluong K, Li L, Sun M, Cao TC, Liu PS, Modrusan Z, Sandoval WN, Rangell L, Carano RA, Peterson AS, Solloway MJ - J. Cell Biol. (2011)

Bottom Line: These changes are associated with intracellular accumulation of collagen and the induction of a strong unfolded protein response, primarily within the developing skeleton.Chondrocyte maturation and bone mineralization are severely compromised in Mia3- embryos, leading to dwarfism and neonatal lethality.Thus, Mia3's role in protein secretion is much broader than previously realized, and it may, in fact, be required for the efficient secretion of all collagen molecules in higher organisms.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Biology, Genentech, South San Francisco, CA 94080, USA.

ABSTRACT
Melanoma inhibitory activity member 3 (MIA3/TANGO1) [corrected] is an evolutionarily conserved endoplasmic reticulum resident transmembrane protein. Recent in vitro studies have shown that it is required for the loading of collagen VII, but not collagen I, into COPII-coated transport vesicles. In this paper, we show that mice lacking Mia3 are defective for the secretion of numerous collagens, including collagens I, II, III, IV, VII, and IX, from chondrocytes, fibroblasts, endothelial cells, and mural cells. Collagen deposition by these cell types is abnormal, and extracellular matrix composition is compromised. These changes are associated with intracellular accumulation of collagen and the induction of a strong unfolded protein response, primarily within the developing skeleton. Chondrocyte maturation and bone mineralization are severely compromised in Mia3- embryos, leading to dwarfism and neonatal lethality. Thus, Mia3's role in protein secretion is much broader than previously realized, and it may, in fact, be required for the efficient secretion of all collagen molecules in higher organisms.

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Collagen and Comp retention and reduced fibril formation in Mia3- embryonic tissues. (A–H) Matrix deposition of Col2a1 (A–D), Col3a1 (E and F), and Col1 (G and H) is reduced around chondrocytes within 14.5-dpc Mia3- bones. Boxed regions are enlarged in adjacent panels. Col2a1 overlaps exclusively with the ER marker Hsp47 in  chondrocytes (C and D), with similar results seen with Col1 and Col3a1 (not depicted). (G and H) WGA labeling of the plasma membrane reveals intracellular accumulation of Col1 in  cells. Yellow arrows in all panels highlight these collagen aggregates. (I–L) Deposition of Col1 (I and J) and Col3a1 (K and L) within collagen fibrils is reduced in Mia3- subdermal mesenchymal cells with associated intracellular accumulation. (M and N) Vascular Col4a1 is present within intracellular aggregates in the Mia3 knockouts. (O–R) Colabeling of Col4a1 and MECA-32 (O and P) or smooth muscle actin (Q and R) demonstrates abnormal collagen accumulation within both endothelial (yellow arrows) and mural cells (pink arrow) of the limb vasculature. (S and T) Col4a1 is disrupted in the basement membrane of the Mia3 mutant muscle. (U–V″) Comp is predominantly ECM associated in wt hypertrophic chondrocytes (enlarged in U′) and resting chondrocytes (U″) of the 14.5-dpc humerus, whereas in Mia3−/− cartilage condensates, COMP is found both within and without the hypertrophic chondrocytes (V′) and is predominantly intracellular in the growth plate (V″). (W–X″) Antibody labeling of Comp intracellular aggregates (W and X) and Col2a1 aggregates (W′ and X′) within Mia3- chondrocytes overlaps (W″ and X″) with cell-retained Col2a1, indicating that both factors are trapped within the ER. Bars: (A–T and W–X″) 10 µm; (U–V″) 100 µm.
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fig6: Collagen and Comp retention and reduced fibril formation in Mia3- embryonic tissues. (A–H) Matrix deposition of Col2a1 (A–D), Col3a1 (E and F), and Col1 (G and H) is reduced around chondrocytes within 14.5-dpc Mia3- bones. Boxed regions are enlarged in adjacent panels. Col2a1 overlaps exclusively with the ER marker Hsp47 in chondrocytes (C and D), with similar results seen with Col1 and Col3a1 (not depicted). (G and H) WGA labeling of the plasma membrane reveals intracellular accumulation of Col1 in cells. Yellow arrows in all panels highlight these collagen aggregates. (I–L) Deposition of Col1 (I and J) and Col3a1 (K and L) within collagen fibrils is reduced in Mia3- subdermal mesenchymal cells with associated intracellular accumulation. (M and N) Vascular Col4a1 is present within intracellular aggregates in the Mia3 knockouts. (O–R) Colabeling of Col4a1 and MECA-32 (O and P) or smooth muscle actin (Q and R) demonstrates abnormal collagen accumulation within both endothelial (yellow arrows) and mural cells (pink arrow) of the limb vasculature. (S and T) Col4a1 is disrupted in the basement membrane of the Mia3 mutant muscle. (U–V″) Comp is predominantly ECM associated in wt hypertrophic chondrocytes (enlarged in U′) and resting chondrocytes (U″) of the 14.5-dpc humerus, whereas in Mia3−/− cartilage condensates, COMP is found both within and without the hypertrophic chondrocytes (V′) and is predominantly intracellular in the growth plate (V″). (W–X″) Antibody labeling of Comp intracellular aggregates (W and X) and Col2a1 aggregates (W′ and X′) within Mia3- chondrocytes overlaps (W″ and X″) with cell-retained Col2a1, indicating that both factors are trapped within the ER. Bars: (A–T and W–X″) 10 µm; (U–V″) 100 µm.

Mentions: To evaluate collagen secretion defects in vivo, we looked at the distribution of collagen in the developing bone at 14.5 dpc. Col2a1 is highly expressed, but extracellular staining is significantly reduced and accompanied by intracellular accumulation in the knockout animals (Fig. 6, A–D). Expression of Col3a1, a prototypical fibrillar collagen, is significantly up-regulated in the mutant chondrocytes but is only detectable intracellularly rather than in the ECM (Fig. 6, E and F). Col1, another fibrillar collagen, is abundantly expressed in the mutant but mislocalized to an intracellular compartment (Fig. 6, G and H). Costaining of the plasma membrane with WGA further emphasizes the significant reduction of Col1 incorporation into extracellular collagen fibrils (Fig. 6, G′ and H′).


Global defects in collagen secretion in a Mia3/TANGO1 knockout mouse.

Wilson DG, Phamluong K, Li L, Sun M, Cao TC, Liu PS, Modrusan Z, Sandoval WN, Rangell L, Carano RA, Peterson AS, Solloway MJ - J. Cell Biol. (2011)

Collagen and Comp retention and reduced fibril formation in Mia3- embryonic tissues. (A–H) Matrix deposition of Col2a1 (A–D), Col3a1 (E and F), and Col1 (G and H) is reduced around chondrocytes within 14.5-dpc Mia3- bones. Boxed regions are enlarged in adjacent panels. Col2a1 overlaps exclusively with the ER marker Hsp47 in  chondrocytes (C and D), with similar results seen with Col1 and Col3a1 (not depicted). (G and H) WGA labeling of the plasma membrane reveals intracellular accumulation of Col1 in  cells. Yellow arrows in all panels highlight these collagen aggregates. (I–L) Deposition of Col1 (I and J) and Col3a1 (K and L) within collagen fibrils is reduced in Mia3- subdermal mesenchymal cells with associated intracellular accumulation. (M and N) Vascular Col4a1 is present within intracellular aggregates in the Mia3 knockouts. (O–R) Colabeling of Col4a1 and MECA-32 (O and P) or smooth muscle actin (Q and R) demonstrates abnormal collagen accumulation within both endothelial (yellow arrows) and mural cells (pink arrow) of the limb vasculature. (S and T) Col4a1 is disrupted in the basement membrane of the Mia3 mutant muscle. (U–V″) Comp is predominantly ECM associated in wt hypertrophic chondrocytes (enlarged in U′) and resting chondrocytes (U″) of the 14.5-dpc humerus, whereas in Mia3−/− cartilage condensates, COMP is found both within and without the hypertrophic chondrocytes (V′) and is predominantly intracellular in the growth plate (V″). (W–X″) Antibody labeling of Comp intracellular aggregates (W and X) and Col2a1 aggregates (W′ and X′) within Mia3- chondrocytes overlaps (W″ and X″) with cell-retained Col2a1, indicating that both factors are trapped within the ER. Bars: (A–T and W–X″) 10 µm; (U–V″) 100 µm.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3105544&req=5

fig6: Collagen and Comp retention and reduced fibril formation in Mia3- embryonic tissues. (A–H) Matrix deposition of Col2a1 (A–D), Col3a1 (E and F), and Col1 (G and H) is reduced around chondrocytes within 14.5-dpc Mia3- bones. Boxed regions are enlarged in adjacent panels. Col2a1 overlaps exclusively with the ER marker Hsp47 in chondrocytes (C and D), with similar results seen with Col1 and Col3a1 (not depicted). (G and H) WGA labeling of the plasma membrane reveals intracellular accumulation of Col1 in cells. Yellow arrows in all panels highlight these collagen aggregates. (I–L) Deposition of Col1 (I and J) and Col3a1 (K and L) within collagen fibrils is reduced in Mia3- subdermal mesenchymal cells with associated intracellular accumulation. (M and N) Vascular Col4a1 is present within intracellular aggregates in the Mia3 knockouts. (O–R) Colabeling of Col4a1 and MECA-32 (O and P) or smooth muscle actin (Q and R) demonstrates abnormal collagen accumulation within both endothelial (yellow arrows) and mural cells (pink arrow) of the limb vasculature. (S and T) Col4a1 is disrupted in the basement membrane of the Mia3 mutant muscle. (U–V″) Comp is predominantly ECM associated in wt hypertrophic chondrocytes (enlarged in U′) and resting chondrocytes (U″) of the 14.5-dpc humerus, whereas in Mia3−/− cartilage condensates, COMP is found both within and without the hypertrophic chondrocytes (V′) and is predominantly intracellular in the growth plate (V″). (W–X″) Antibody labeling of Comp intracellular aggregates (W and X) and Col2a1 aggregates (W′ and X′) within Mia3- chondrocytes overlaps (W″ and X″) with cell-retained Col2a1, indicating that both factors are trapped within the ER. Bars: (A–T and W–X″) 10 µm; (U–V″) 100 µm.
Mentions: To evaluate collagen secretion defects in vivo, we looked at the distribution of collagen in the developing bone at 14.5 dpc. Col2a1 is highly expressed, but extracellular staining is significantly reduced and accompanied by intracellular accumulation in the knockout animals (Fig. 6, A–D). Expression of Col3a1, a prototypical fibrillar collagen, is significantly up-regulated in the mutant chondrocytes but is only detectable intracellularly rather than in the ECM (Fig. 6, E and F). Col1, another fibrillar collagen, is abundantly expressed in the mutant but mislocalized to an intracellular compartment (Fig. 6, G and H). Costaining of the plasma membrane with WGA further emphasizes the significant reduction of Col1 incorporation into extracellular collagen fibrils (Fig. 6, G′ and H′).

Bottom Line: These changes are associated with intracellular accumulation of collagen and the induction of a strong unfolded protein response, primarily within the developing skeleton.Chondrocyte maturation and bone mineralization are severely compromised in Mia3- embryos, leading to dwarfism and neonatal lethality.Thus, Mia3's role in protein secretion is much broader than previously realized, and it may, in fact, be required for the efficient secretion of all collagen molecules in higher organisms.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Biology, Genentech, South San Francisco, CA 94080, USA.

ABSTRACT
Melanoma inhibitory activity member 3 (MIA3/TANGO1) [corrected] is an evolutionarily conserved endoplasmic reticulum resident transmembrane protein. Recent in vitro studies have shown that it is required for the loading of collagen VII, but not collagen I, into COPII-coated transport vesicles. In this paper, we show that mice lacking Mia3 are defective for the secretion of numerous collagens, including collagens I, II, III, IV, VII, and IX, from chondrocytes, fibroblasts, endothelial cells, and mural cells. Collagen deposition by these cell types is abnormal, and extracellular matrix composition is compromised. These changes are associated with intracellular accumulation of collagen and the induction of a strong unfolded protein response, primarily within the developing skeleton. Chondrocyte maturation and bone mineralization are severely compromised in Mia3- embryos, leading to dwarfism and neonatal lethality. Thus, Mia3's role in protein secretion is much broader than previously realized, and it may, in fact, be required for the efficient secretion of all collagen molecules in higher organisms.

Show MeSH
Related in: MedlinePlus