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alpha3beta1 Integrin is required for normal development of the epidermal basement membrane.

DiPersio CM, Hodivala-Dilke KM, Jaenisch R, Kreidberg JA, Hynes RO - J. Cell Biol. (1997)

Bottom Line: Laminin-5 and other matrix proteins remained associated with both the dermal and epidermal sides of blisters, suggesting rupture of the basement membrane itself, rather than detachment of the epidermis from the basement membrane as occurs in some blistering disorders such as epidermolysis bullosa.Consistent with this notion, primary keratinocytes from alpha3beta1-deficient skin adhered to laminin-5 through alpha6 integrins.However, alpha3beta1-deficient keratinocytes spread poorly compared with wild-type cells on laminin-5, demonstrating a postattachment requirement for alpha3beta1 and indicating distinct roles for alpha3beta1 and alpha6beta4.

View Article: PubMed Central - PubMed

Affiliation: Center for Cancer Research, and Department of Biology, Massachusetts Institute of Technology, Cambridge 02139, USA.

ABSTRACT
Integrins alpha3beta1 and alpha6beta4 are abundant receptors on keratinocytes for laminin-5, a major component of the basement membrane between the epidermis and the dermis in skin. These integrins are recruited to distinct adhesion structures within keratinocytes; alpha6beta4 is present in hemidesmosomes, while alpha3beta1 is recruited into focal contacts in cultured cells. To determine whether differences in localization reflect distinct functions of these integrins in the epidermis, we studied skin development in alpha3beta1-deficient mice. Examination of extracellular matrix by immunofluorescence microscopy and electron microscopy revealed regions of disorganized basement membrane in alpha3beta1-deficient skin. Disorganized matrix was first detected by day 15.5 of embryonic development and became progressively more extensive as development proceeded. In neonatal skin, matrix disorganization was frequently accompanied by blistering at the dermal-epidermal junction. Laminin-5 and other matrix proteins remained associated with both the dermal and epidermal sides of blisters, suggesting rupture of the basement membrane itself, rather than detachment of the epidermis from the basement membrane as occurs in some blistering disorders such as epidermolysis bullosa. Consistent with this notion, primary keratinocytes from alpha3beta1-deficient skin adhered to laminin-5 through alpha6 integrins. However, alpha3beta1-deficient keratinocytes spread poorly compared with wild-type cells on laminin-5, demonstrating a postattachment requirement for alpha3beta1 and indicating distinct roles for alpha3beta1 and alpha6beta4. Our findings support a novel role for alpha3beta1 in establishment and/or maintenance of basement membrane integrity, while alpha6beta4 is required for stable adhesion of the epidermis to the basement membrane through hemidesmosomes.

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Surface expression of integrins in primary keratinocytes isolated from wild-type (lanes 1–3) or α3- (lanes 4–6),  neonatal mice. Detergent lysates from 125I surface-labeled cells were  immunoprecipitated with antisera against the cytoplasmic domains of the β1, α3, or α6 integrin subunits, as indicated at the  top of each lane. Molecular weight markers are shown to the right  of the autoradiograph. Migratory positions of certain integrin  subunits are indicated to the left, including proteolytic fragments  of β4, a, b, and c, described previously by Hemler et al. (1989).  The β1-associated band in α3- cells that comigrates with α3  (lane 4) represents other integrin α subunits that dimerize with  β1 in keratinocytes, since α3 was not detected in α3- cells  (lane 5). ?, an unidentified band that may represent a proteolytic  fragment of β4 (see text).
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Figure 8: Surface expression of integrins in primary keratinocytes isolated from wild-type (lanes 1–3) or α3- (lanes 4–6), neonatal mice. Detergent lysates from 125I surface-labeled cells were immunoprecipitated with antisera against the cytoplasmic domains of the β1, α3, or α6 integrin subunits, as indicated at the top of each lane. Molecular weight markers are shown to the right of the autoradiograph. Migratory positions of certain integrin subunits are indicated to the left, including proteolytic fragments of β4, a, b, and c, described previously by Hemler et al. (1989). The β1-associated band in α3- cells that comigrates with α3 (lane 4) represents other integrin α subunits that dimerize with β1 in keratinocytes, since α3 was not detected in α3- cells (lane 5). ?, an unidentified band that may represent a proteolytic fragment of β4 (see text).

Mentions: To address directly the importance of α3β1 as a laminin-5 receptor in the epidermis, we isolated primary keratinocytes from wild-type and α3-, neonatal mice (Dlugosz et al., 1995; for details see Materials and Methods). To compare surface expression of integrins that bind to laminin-5 in wild-type and α3β1-deficient keratinocytes, 7-d, primary cultures were surface-iodinated with 125I, and detergent lysates were immunoprecipitated with antibodies against integrin subunits and analyzed by SDS-PAGE under nonreducing conditions (Fig. 8). As expected, antiserum against α3 immunoprecipitated an abundance of the α3 subunit (∼150 kD) associated with the β1 subunit (∼110 kD) from lysates of wild-type cells (lane 2), but did not detect α3β1 in lysates from α3- cells (lane 5). An anti-β1 serum (lane 1) coimmunoprecipitated β1 and associated proteins of ∼150 kD, corresponding to α3 and other comigrating α subunits present in keratinocytes (Watt and Hertle, 1994). Surface levels of β1 integrins dropped significantly in α3 cells compared to wild-type cells (compare lanes 1 and 4), demonstrating that α3β1 constitutes the major proportion of total β1 integrins in mouse keratinocytes.


alpha3beta1 Integrin is required for normal development of the epidermal basement membrane.

DiPersio CM, Hodivala-Dilke KM, Jaenisch R, Kreidberg JA, Hynes RO - J. Cell Biol. (1997)

Surface expression of integrins in primary keratinocytes isolated from wild-type (lanes 1–3) or α3- (lanes 4–6),  neonatal mice. Detergent lysates from 125I surface-labeled cells were  immunoprecipitated with antisera against the cytoplasmic domains of the β1, α3, or α6 integrin subunits, as indicated at the  top of each lane. Molecular weight markers are shown to the right  of the autoradiograph. Migratory positions of certain integrin  subunits are indicated to the left, including proteolytic fragments  of β4, a, b, and c, described previously by Hemler et al. (1989).  The β1-associated band in α3- cells that comigrates with α3  (lane 4) represents other integrin α subunits that dimerize with  β1 in keratinocytes, since α3 was not detected in α3- cells  (lane 5). ?, an unidentified band that may represent a proteolytic  fragment of β4 (see text).
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Related In: Results  -  Collection

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Figure 8: Surface expression of integrins in primary keratinocytes isolated from wild-type (lanes 1–3) or α3- (lanes 4–6), neonatal mice. Detergent lysates from 125I surface-labeled cells were immunoprecipitated with antisera against the cytoplasmic domains of the β1, α3, or α6 integrin subunits, as indicated at the top of each lane. Molecular weight markers are shown to the right of the autoradiograph. Migratory positions of certain integrin subunits are indicated to the left, including proteolytic fragments of β4, a, b, and c, described previously by Hemler et al. (1989). The β1-associated band in α3- cells that comigrates with α3 (lane 4) represents other integrin α subunits that dimerize with β1 in keratinocytes, since α3 was not detected in α3- cells (lane 5). ?, an unidentified band that may represent a proteolytic fragment of β4 (see text).
Mentions: To address directly the importance of α3β1 as a laminin-5 receptor in the epidermis, we isolated primary keratinocytes from wild-type and α3-, neonatal mice (Dlugosz et al., 1995; for details see Materials and Methods). To compare surface expression of integrins that bind to laminin-5 in wild-type and α3β1-deficient keratinocytes, 7-d, primary cultures were surface-iodinated with 125I, and detergent lysates were immunoprecipitated with antibodies against integrin subunits and analyzed by SDS-PAGE under nonreducing conditions (Fig. 8). As expected, antiserum against α3 immunoprecipitated an abundance of the α3 subunit (∼150 kD) associated with the β1 subunit (∼110 kD) from lysates of wild-type cells (lane 2), but did not detect α3β1 in lysates from α3- cells (lane 5). An anti-β1 serum (lane 1) coimmunoprecipitated β1 and associated proteins of ∼150 kD, corresponding to α3 and other comigrating α subunits present in keratinocytes (Watt and Hertle, 1994). Surface levels of β1 integrins dropped significantly in α3 cells compared to wild-type cells (compare lanes 1 and 4), demonstrating that α3β1 constitutes the major proportion of total β1 integrins in mouse keratinocytes.

Bottom Line: Laminin-5 and other matrix proteins remained associated with both the dermal and epidermal sides of blisters, suggesting rupture of the basement membrane itself, rather than detachment of the epidermis from the basement membrane as occurs in some blistering disorders such as epidermolysis bullosa.Consistent with this notion, primary keratinocytes from alpha3beta1-deficient skin adhered to laminin-5 through alpha6 integrins.However, alpha3beta1-deficient keratinocytes spread poorly compared with wild-type cells on laminin-5, demonstrating a postattachment requirement for alpha3beta1 and indicating distinct roles for alpha3beta1 and alpha6beta4.

View Article: PubMed Central - PubMed

Affiliation: Center for Cancer Research, and Department of Biology, Massachusetts Institute of Technology, Cambridge 02139, USA.

ABSTRACT
Integrins alpha3beta1 and alpha6beta4 are abundant receptors on keratinocytes for laminin-5, a major component of the basement membrane between the epidermis and the dermis in skin. These integrins are recruited to distinct adhesion structures within keratinocytes; alpha6beta4 is present in hemidesmosomes, while alpha3beta1 is recruited into focal contacts in cultured cells. To determine whether differences in localization reflect distinct functions of these integrins in the epidermis, we studied skin development in alpha3beta1-deficient mice. Examination of extracellular matrix by immunofluorescence microscopy and electron microscopy revealed regions of disorganized basement membrane in alpha3beta1-deficient skin. Disorganized matrix was first detected by day 15.5 of embryonic development and became progressively more extensive as development proceeded. In neonatal skin, matrix disorganization was frequently accompanied by blistering at the dermal-epidermal junction. Laminin-5 and other matrix proteins remained associated with both the dermal and epidermal sides of blisters, suggesting rupture of the basement membrane itself, rather than detachment of the epidermis from the basement membrane as occurs in some blistering disorders such as epidermolysis bullosa. Consistent with this notion, primary keratinocytes from alpha3beta1-deficient skin adhered to laminin-5 through alpha6 integrins. However, alpha3beta1-deficient keratinocytes spread poorly compared with wild-type cells on laminin-5, demonstrating a postattachment requirement for alpha3beta1 and indicating distinct roles for alpha3beta1 and alpha6beta4. Our findings support a novel role for alpha3beta1 in establishment and/or maintenance of basement membrane integrity, while alpha6beta4 is required for stable adhesion of the epidermis to the basement membrane through hemidesmosomes.

Show MeSH
Related in: MedlinePlus