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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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Osteogenesis is compromised by the delay in chondrogenic progression and vascular recruitment in Mia3- bone. (A and B) In contrast to wt embryos, rare Spp1-positive osteoblasts (arrows) are present in the perichondrium of the Mia3−/− humerus at 14.5 dpc. (C and D) Mmp9 and PLVAP/MECA-32 are restricted to a few cells within the perichondrium of Mia3−/− bones at 14.5 dpc. (E and G) At 16.5 dpc, Mmp9-positive cells remain restricted to the presumptive bone collar perichondrium of Mia3 mutant embryos. (F and H) A small number of Spp1-positive osteoblasts reside within the hypertrophic zone. (I–N’) Masson’s trichrome stain (I–L) and hematoxylin/eosin stain (M and N) of wt and Mia3- humeri at 13.5, 15.5, and 18.5 dpc highlight the onset and progression of the chondrodysplastic phenotype. Boxed regions (I–N) are enlarged to the right of each set of panels (I′–N′). Bars, 100 µm.
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fig3: Osteogenesis is compromised by the delay in chondrogenic progression and vascular recruitment in Mia3- bone. (A and B) In contrast to wt embryos, rare Spp1-positive osteoblasts (arrows) are present in the perichondrium of the Mia3−/− humerus at 14.5 dpc. (C and D) Mmp9 and PLVAP/MECA-32 are restricted to a few cells within the perichondrium of Mia3−/− bones at 14.5 dpc. (E and G) At 16.5 dpc, Mmp9-positive cells remain restricted to the presumptive bone collar perichondrium of Mia3 mutant embryos. (F and H) A small number of Spp1-positive osteoblasts reside within the hypertrophic zone. (I–N’) Masson’s trichrome stain (I–L) and hematoxylin/eosin stain (M and N) of wt and Mia3- humeri at 13.5, 15.5, and 18.5 dpc highlight the onset and progression of the chondrodysplastic phenotype. Boxed regions (I–N) are enlarged to the right of each set of panels (I′–N′). Bars, 100 µm.

Mentions: Residual mineral deposition is apparent in the dwarfed mutants at 18.5 dpc (Fig. 2, I and J), but microcomputed tomography (micro-CT) fails to detect ossified tissue, which would indicate secondary mineralization in the mutants (n = 3; Fig. 2, K and L). Secondary mineralization is driven by osteoblasts, and indeed, Mia3 mutants have a severe reduction in osteopontin (Spp1)-positive osteoblasts at 14.5 dpc (Fig. 3, A and B) and show no detectable staining for osteoblast-derived Col1a1 and Igfbp6 (insulin-like growth factor–binding protein) in the perichondrium at this stage (not depicted). The few osteoblasts observed in the mutant bones were found primarily in the periphery of the perichondrium adjacent to the hypertrophic region at the middle of the condensation (Fig. 3 B). In the controls, these cells are well entrenched in the perichondrium and have invaded the remodeling matrix of the hypertrophic zone by 14.5 dpc (Fig. 3 A). Expression of Mmp9 (matrix metallopeptidase 9), a secreted ECM-degrading enzyme, which primes the mineralized matrix for vascular invasion (Vu et al., 1998), is strongly down-regulated (Fig. 3, C and D), and MECA-32–positive vessels are not apparent in the mutant hypertrophic zone. Clearly, the dependence of osteoblasts on Mia3 is not absolute, and by 16.5 dpc more, osteoblasts are apparent within the center of each bone, although their numbers are still drastically reduced with respect to wt littermates (Fig. 3, F and H), and they are restricted to the marrow cavity immediately underlying the Mmp9-positive perichondrium (Fig. 3, E and G). Thus, the absence of secondary mineralization is caused by severe defects in vascular invasion and recruitment of osteoblasts in Mia3 mutants. We hypothesized that this was caused by a general delay in chondrogenic maturation and focused our further analysis on this.


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)

Osteogenesis is compromised by the delay in chondrogenic progression and vascular recruitment in Mia3- bone. (A and B) In contrast to wt embryos, rare Spp1-positive osteoblasts (arrows) are present in the perichondrium of the Mia3−/− humerus at 14.5 dpc. (C and D) Mmp9 and PLVAP/MECA-32 are restricted to a few cells within the perichondrium of Mia3−/− bones at 14.5 dpc. (E and G) At 16.5 dpc, Mmp9-positive cells remain restricted to the presumptive bone collar perichondrium of Mia3 mutant embryos. (F and H) A small number of Spp1-positive osteoblasts reside within the hypertrophic zone. (I–N’) Masson’s trichrome stain (I–L) and hematoxylin/eosin stain (M and N) of wt and Mia3- humeri at 13.5, 15.5, and 18.5 dpc highlight the onset and progression of the chondrodysplastic phenotype. Boxed regions (I–N) are enlarged to the right of each set of panels (I′–N′). Bars, 100 µm.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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

fig3: Osteogenesis is compromised by the delay in chondrogenic progression and vascular recruitment in Mia3- bone. (A and B) In contrast to wt embryos, rare Spp1-positive osteoblasts (arrows) are present in the perichondrium of the Mia3−/− humerus at 14.5 dpc. (C and D) Mmp9 and PLVAP/MECA-32 are restricted to a few cells within the perichondrium of Mia3−/− bones at 14.5 dpc. (E and G) At 16.5 dpc, Mmp9-positive cells remain restricted to the presumptive bone collar perichondrium of Mia3 mutant embryos. (F and H) A small number of Spp1-positive osteoblasts reside within the hypertrophic zone. (I–N’) Masson’s trichrome stain (I–L) and hematoxylin/eosin stain (M and N) of wt and Mia3- humeri at 13.5, 15.5, and 18.5 dpc highlight the onset and progression of the chondrodysplastic phenotype. Boxed regions (I–N) are enlarged to the right of each set of panels (I′–N′). Bars, 100 µm.
Mentions: Residual mineral deposition is apparent in the dwarfed mutants at 18.5 dpc (Fig. 2, I and J), but microcomputed tomography (micro-CT) fails to detect ossified tissue, which would indicate secondary mineralization in the mutants (n = 3; Fig. 2, K and L). Secondary mineralization is driven by osteoblasts, and indeed, Mia3 mutants have a severe reduction in osteopontin (Spp1)-positive osteoblasts at 14.5 dpc (Fig. 3, A and B) and show no detectable staining for osteoblast-derived Col1a1 and Igfbp6 (insulin-like growth factor–binding protein) in the perichondrium at this stage (not depicted). The few osteoblasts observed in the mutant bones were found primarily in the periphery of the perichondrium adjacent to the hypertrophic region at the middle of the condensation (Fig. 3 B). In the controls, these cells are well entrenched in the perichondrium and have invaded the remodeling matrix of the hypertrophic zone by 14.5 dpc (Fig. 3 A). Expression of Mmp9 (matrix metallopeptidase 9), a secreted ECM-degrading enzyme, which primes the mineralized matrix for vascular invasion (Vu et al., 1998), is strongly down-regulated (Fig. 3, C and D), and MECA-32–positive vessels are not apparent in the mutant hypertrophic zone. Clearly, the dependence of osteoblasts on Mia3 is not absolute, and by 16.5 dpc more, osteoblasts are apparent within the center of each bone, although their numbers are still drastically reduced with respect to wt littermates (Fig. 3, F and H), and they are restricted to the marrow cavity immediately underlying the Mmp9-positive perichondrium (Fig. 3, E and G). Thus, the absence of secondary mineralization is caused by severe defects in vascular invasion and recruitment of osteoblasts in Mia3 mutants. We hypothesized that this was caused by a general delay in chondrogenic maturation and focused our further analysis on this.

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