Limits...
Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the glomerular basement membrane.

Kikkawa Y, Virtanen I, Miner JH - J. Cell Biol. (2003)

Bottom Line: In addition, podocytes exhibited their typical arrangement in a single cell layer epithelium adjacent to the GBM, but convolution of glomerular capillaries did not occur.Finally, in vitro studies showed that integrin alpha3beta1 and the Lutheran glycoprotein mediate adhesion of mesangial cells to laminin alpha5.Our results elucidate a mechanism whereby mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the GBM.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, USA.

ABSTRACT
In developing glomeruli, laminin alpha5 replaces laminin alpha1 in the glomerular basement membrane (GBM) at the capillary loop stage, a transition required for glomerulogenesis. To investigate domain-specific functions of laminin alpha5 during glomerulogenesis, we produced transgenic mice that express a chimeric laminin composed of laminin alpha5 domains VI through I fused to the human laminin alpha1 globular (G) domain, designated Mr51. Transgene-derived protein accumulated in many basement membranes, including the developing GBM. When bred onto the Lama5 -/- background, Mr51 supported GBM formation, preventing the breakdown that normally occurs in Lama5 -/- glomeruli. In addition, podocytes exhibited their typical arrangement in a single cell layer epithelium adjacent to the GBM, but convolution of glomerular capillaries did not occur. Instead, capillaries were distended and exhibited a ballooned appearance, a phenotype similar to that observed in the total absence of mesangial cells. However, here the phenotype could be attributed to the lack of mesangial cell adhesion to the GBM, suggesting that the G domain of laminin alpha5 is essential for this adhesion. Analysis of an additional chimeric transgene allowed us to narrow the region of the alpha5 G domain essential for mesangial cell adhesion to alpha5LG3-5. Finally, in vitro studies showed that integrin alpha3beta1 and the Lutheran glycoprotein mediate adhesion of mesangial cells to laminin alpha5. Our results elucidate a mechanism whereby mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the GBM.

Show MeSH

Related in: MedlinePlus

Structure of wild-type and chimeric laminin chains. The domains present in full-length laminin α5 (A), in the chimeric laminin α chains (B and C), and in full–length human α1 (D) are shown. (B) Mr51 contains human laminin α1 G domain linked to domains VI through I of mouse laminin α5. (C) Mr5G2 contains laminin α5 domains VI through α5 LG2 fused to the human laminin α1LG3-5 domain. Anti–mouse laminin α5 (*) and anti–human laminin α1 LG1–2 (**) antibody epitopes are indicated.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2172883&req=5

fig2: Structure of wild-type and chimeric laminin chains. The domains present in full-length laminin α5 (A), in the chimeric laminin α chains (B and C), and in full–length human α1 (D) are shown. (B) Mr51 contains human laminin α1 G domain linked to domains VI through I of mouse laminin α5. (C) Mr5G2 contains laminin α5 domains VI through α5 LG2 fused to the human laminin α1LG3-5 domain. Anti–mouse laminin α5 (*) and anti–human laminin α1 LG1–2 (**) antibody epitopes are indicated.

Mentions: To begin to examine domain-specific functions of laminin α5, we produced transgenic mice expressing two different full-length chimeric laminin α chains. These encoded laminin α5 domains VI through I and VI through LG2 fused to the complete human laminin α1 G domain and α1LG3-5, designated Mr51 and Mr5G2, respectively (Fig. 2, B and C) . We chose to use the human rather than mouse α1 G domain because of the availability of mouse monoclonal antibodies specific for the human domain (Virtanen et al., 2000); thus, transgene-derived proteins could be specifically localized in transgenic mouse tissues. A transgene encoding the full-length mouse α5 chain, designated Mr5 (Fig. 2 A), served as a control. The widely active regulatory element miw (Suemori et al., 1990) was used to drive transgene expression. As described in our previous papers, transgene-derived laminin levels were significantly increased in heart and skeletal muscle (Moulson et al., 2001; Kikkawa et al., 2002). Crossing of the Mr5 transgene onto the Lama5 −/− background revealed that transgene-derived laminin α5 was deposited widely in basement membranes. Expression was sufficient to fully rescue all known Lama5 −/− embryonic defects in two independent lines, and the resulting Lama5 −/−; Mr5 mice are viable and fertile (unpublished observations). These results show that the miw regulatory element directs expression of the transgene in a manner sufficient to replace the missing endogenous α5 wherever it is necessary.


Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the glomerular basement membrane.

Kikkawa Y, Virtanen I, Miner JH - J. Cell Biol. (2003)

Structure of wild-type and chimeric laminin chains. The domains present in full-length laminin α5 (A), in the chimeric laminin α chains (B and C), and in full–length human α1 (D) are shown. (B) Mr51 contains human laminin α1 G domain linked to domains VI through I of mouse laminin α5. (C) Mr5G2 contains laminin α5 domains VI through α5 LG2 fused to the human laminin α1LG3-5 domain. Anti–mouse laminin α5 (*) and anti–human laminin α1 LG1–2 (**) antibody epitopes are indicated.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172883&req=5

fig2: Structure of wild-type and chimeric laminin chains. The domains present in full-length laminin α5 (A), in the chimeric laminin α chains (B and C), and in full–length human α1 (D) are shown. (B) Mr51 contains human laminin α1 G domain linked to domains VI through I of mouse laminin α5. (C) Mr5G2 contains laminin α5 domains VI through α5 LG2 fused to the human laminin α1LG3-5 domain. Anti–mouse laminin α5 (*) and anti–human laminin α1 LG1–2 (**) antibody epitopes are indicated.
Mentions: To begin to examine domain-specific functions of laminin α5, we produced transgenic mice expressing two different full-length chimeric laminin α chains. These encoded laminin α5 domains VI through I and VI through LG2 fused to the complete human laminin α1 G domain and α1LG3-5, designated Mr51 and Mr5G2, respectively (Fig. 2, B and C) . We chose to use the human rather than mouse α1 G domain because of the availability of mouse monoclonal antibodies specific for the human domain (Virtanen et al., 2000); thus, transgene-derived proteins could be specifically localized in transgenic mouse tissues. A transgene encoding the full-length mouse α5 chain, designated Mr5 (Fig. 2 A), served as a control. The widely active regulatory element miw (Suemori et al., 1990) was used to drive transgene expression. As described in our previous papers, transgene-derived laminin levels were significantly increased in heart and skeletal muscle (Moulson et al., 2001; Kikkawa et al., 2002). Crossing of the Mr5 transgene onto the Lama5 −/− background revealed that transgene-derived laminin α5 was deposited widely in basement membranes. Expression was sufficient to fully rescue all known Lama5 −/− embryonic defects in two independent lines, and the resulting Lama5 −/−; Mr5 mice are viable and fertile (unpublished observations). These results show that the miw regulatory element directs expression of the transgene in a manner sufficient to replace the missing endogenous α5 wherever it is necessary.

Bottom Line: In addition, podocytes exhibited their typical arrangement in a single cell layer epithelium adjacent to the GBM, but convolution of glomerular capillaries did not occur.Finally, in vitro studies showed that integrin alpha3beta1 and the Lutheran glycoprotein mediate adhesion of mesangial cells to laminin alpha5.Our results elucidate a mechanism whereby mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the GBM.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Washington University School of Medicine, St Louis, MO 63110, USA.

ABSTRACT
In developing glomeruli, laminin alpha5 replaces laminin alpha1 in the glomerular basement membrane (GBM) at the capillary loop stage, a transition required for glomerulogenesis. To investigate domain-specific functions of laminin alpha5 during glomerulogenesis, we produced transgenic mice that express a chimeric laminin composed of laminin alpha5 domains VI through I fused to the human laminin alpha1 globular (G) domain, designated Mr51. Transgene-derived protein accumulated in many basement membranes, including the developing GBM. When bred onto the Lama5 -/- background, Mr51 supported GBM formation, preventing the breakdown that normally occurs in Lama5 -/- glomeruli. In addition, podocytes exhibited their typical arrangement in a single cell layer epithelium adjacent to the GBM, but convolution of glomerular capillaries did not occur. Instead, capillaries were distended and exhibited a ballooned appearance, a phenotype similar to that observed in the total absence of mesangial cells. However, here the phenotype could be attributed to the lack of mesangial cell adhesion to the GBM, suggesting that the G domain of laminin alpha5 is essential for this adhesion. Analysis of an additional chimeric transgene allowed us to narrow the region of the alpha5 G domain essential for mesangial cell adhesion to alpha5LG3-5. Finally, in vitro studies showed that integrin alpha3beta1 and the Lutheran glycoprotein mediate adhesion of mesangial cells to laminin alpha5. Our results elucidate a mechanism whereby mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin alpha5 in the GBM.

Show MeSH
Related in: MedlinePlus