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Regulated splicing of the fibronectin EDA exon is essential for proper skin wound healing and normal lifespan.

Muro AF, Chauhan AK, Gajovic S, Iaconcig A, Porro F, Stanta G, Baralle FE - J. Cell Biol. (2003)

Bottom Line: However, the precise role of the FN isoforms is poorly understood.One of the alternatively spliced exons is the extra domain A (EDA) or extra type III homology that is regulated spatially and temporally during development and aging.Constitutive exon inclusion was obtained by optimizing the splice sites, whereas complete exclusion was obtained after in vivo CRE-loxP-mediated deletion of the exon.

View Article: PubMed Central - PubMed

Affiliation: International Center for Genetic Engineering and Biotechnology (ICGEB), Padriciano 99, 34012 Trieste, Italy.

ABSTRACT
Fibronectins (FNs) are multifunctional high molecular weight glycoproteins present in the blood plasma and in the ECMs of tissues. The FN primary transcript undergoes alternative splicing in three regions generating up to 20 main different variants in humans. However, the precise role of the FN isoforms is poorly understood. One of the alternatively spliced exons is the extra domain A (EDA) or extra type III homology that is regulated spatially and temporally during development and aging. To study its in vivo function, we generated mice devoid of EDA exon-regulated splicing. Constitutive exon inclusion was obtained by optimizing the splice sites, whereas complete exclusion was obtained after in vivo CRE-loxP-mediated deletion of the exon. Homozygous mouse strains with complete exclusion or inclusion of the EDA exon were viable and developed normally, indicating that the alternative splicing at the EDA exon is not necessary during embryonic development. Conversely, mice without the EDA exon in the FN protein displayed abnormal skin wound healing, whereas mice having constitutive inclusion of the EDA exon showed a major decrease in the FN levels in all tissues. Moreover, both mutant mouse strains have a significantly shorter lifespan than the control mice, suggesting that EDA splicing regulation is necessary for efficient long-term maintenance of biological functions.

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Expression of FN in different tissues at different ages. (A) Protein extracts from liver, heart, lung, and brain were prepared from EDAwt/wt, EDA+/+, and EDA−/− mice of different ages (13.5 d p.c., 1, 3, and 14 mo old) and analyzed by Western blot using an anti-FN antibody. (B) The histograms represent the relative amount of FN present in EDA+/+ mice relative to that present in EDAwt/wt mice (considered as 100%) for each tissue and each time point.
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fig4: Expression of FN in different tissues at different ages. (A) Protein extracts from liver, heart, lung, and brain were prepared from EDAwt/wt, EDA+/+, and EDA−/− mice of different ages (13.5 d p.c., 1, 3, and 14 mo old) and analyzed by Western blot using an anti-FN antibody. (B) The histograms represent the relative amount of FN present in EDA+/+ mice relative to that present in EDAwt/wt mice (considered as 100%) for each tissue and each time point.

Mentions: To determine whether the mechanisms that produced the FN decrease were developmentally regulated, we performed a Western blot analysis of samples prepared from mice of different ages. The decrease in FN levels was not present during embryonic development. It started after birth in most of the tissues, and arrived to a plateau in 2-mo-old mice (Fig. 4)Figure 4.


Regulated splicing of the fibronectin EDA exon is essential for proper skin wound healing and normal lifespan.

Muro AF, Chauhan AK, Gajovic S, Iaconcig A, Porro F, Stanta G, Baralle FE - J. Cell Biol. (2003)

Expression of FN in different tissues at different ages. (A) Protein extracts from liver, heart, lung, and brain were prepared from EDAwt/wt, EDA+/+, and EDA−/− mice of different ages (13.5 d p.c., 1, 3, and 14 mo old) and analyzed by Western blot using an anti-FN antibody. (B) The histograms represent the relative amount of FN present in EDA+/+ mice relative to that present in EDAwt/wt mice (considered as 100%) for each tissue and each time point.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172721&req=5

fig4: Expression of FN in different tissues at different ages. (A) Protein extracts from liver, heart, lung, and brain were prepared from EDAwt/wt, EDA+/+, and EDA−/− mice of different ages (13.5 d p.c., 1, 3, and 14 mo old) and analyzed by Western blot using an anti-FN antibody. (B) The histograms represent the relative amount of FN present in EDA+/+ mice relative to that present in EDAwt/wt mice (considered as 100%) for each tissue and each time point.
Mentions: To determine whether the mechanisms that produced the FN decrease were developmentally regulated, we performed a Western blot analysis of samples prepared from mice of different ages. The decrease in FN levels was not present during embryonic development. It started after birth in most of the tissues, and arrived to a plateau in 2-mo-old mice (Fig. 4)Figure 4.

Bottom Line: However, the precise role of the FN isoforms is poorly understood.One of the alternatively spliced exons is the extra domain A (EDA) or extra type III homology that is regulated spatially and temporally during development and aging.Constitutive exon inclusion was obtained by optimizing the splice sites, whereas complete exclusion was obtained after in vivo CRE-loxP-mediated deletion of the exon.

View Article: PubMed Central - PubMed

Affiliation: International Center for Genetic Engineering and Biotechnology (ICGEB), Padriciano 99, 34012 Trieste, Italy.

ABSTRACT
Fibronectins (FNs) are multifunctional high molecular weight glycoproteins present in the blood plasma and in the ECMs of tissues. The FN primary transcript undergoes alternative splicing in three regions generating up to 20 main different variants in humans. However, the precise role of the FN isoforms is poorly understood. One of the alternatively spliced exons is the extra domain A (EDA) or extra type III homology that is regulated spatially and temporally during development and aging. To study its in vivo function, we generated mice devoid of EDA exon-regulated splicing. Constitutive exon inclusion was obtained by optimizing the splice sites, whereas complete exclusion was obtained after in vivo CRE-loxP-mediated deletion of the exon. Homozygous mouse strains with complete exclusion or inclusion of the EDA exon were viable and developed normally, indicating that the alternative splicing at the EDA exon is not necessary during embryonic development. Conversely, mice without the EDA exon in the FN protein displayed abnormal skin wound healing, whereas mice having constitutive inclusion of the EDA exon showed a major decrease in the FN levels in all tissues. Moreover, both mutant mouse strains have a significantly shorter lifespan than the control mice, suggesting that EDA splicing regulation is necessary for efficient long-term maintenance of biological functions.

Show MeSH
Related in: MedlinePlus