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Activation of GSK-3 and phosphorylation of CRMP2 in transgenic mice expressing APP intracellular domain.

Ryan KA, Pimplikar SW - J. Cell Biol. (2005)

Bottom Line: APP is cleaved by gamma-secretase that releases the APP intracellular domain (AICD) in the cytoplasm.In vitro studies have implicated AICD in cell signaling and transcriptional regulation, but its biologic relevance has been uncertain and its in vivo function has not been examined.Our data suggest that AICD is biologically relevant, causes significant alterations in cell signaling, and may play a role in axonal elongation or pathfinding.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Cell Biology Program, Case Western Reserve University, Cleveland, OH 44106, USA.

ABSTRACT
Amyloid precursor protein (APP), implicated in Alzheimer's disease, is a trans-membrane protein of undetermined function. APP is cleaved by gamma-secretase that releases the APP intracellular domain (AICD) in the cytoplasm. In vitro studies have implicated AICD in cell signaling and transcriptional regulation, but its biologic relevance has been uncertain and its in vivo function has not been examined. To investigate its functional role, we generated AICD transgenic mice, and found that AICD causes significant biologic changes in vivo. AICD transgenic mice show activation of glycogen synthase kinase-3beta (GSK-3beta) and phosphorylation of CRMP2 protein, a GSK-3beta substrate that plays a crucial role in Semaphorin3a-mediated axonal guidance. Our data suggest that AICD is biologically relevant, causes significant alterations in cell signaling, and may play a role in axonal elongation or pathfinding.

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Increased KAI1 levels in AICD transgenic mice. (A) The AICD transgenic mice, but not the control or Fe.27 mice, show expression of KAI1 gene. Membrane fractions from indicated mice were probed with anti-KAI1 antibody. FeCγ.25 mice showed higher expression of KAI1 protein compared with FeCγ.12 mice. (B) Membrane fractions from indicated mice were probed with anti-SERCA 2b antibody. No significant changes in SERCA 2b levels were reproducibly observed in FeCγ transgenic mice.
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fig3: Increased KAI1 levels in AICD transgenic mice. (A) The AICD transgenic mice, but not the control or Fe.27 mice, show expression of KAI1 gene. Membrane fractions from indicated mice were probed with anti-KAI1 antibody. FeCγ.25 mice showed higher expression of KAI1 protein compared with FeCγ.12 mice. (B) Membrane fractions from indicated mice were probed with anti-SERCA 2b antibody. No significant changes in SERCA 2b levels were reproducibly observed in FeCγ transgenic mice.

Mentions: Although no bona fide target genes of AICD have been identified genetically, a chromatin immunoprecipitation assay was used recently to show that AICD is recruited to KAI1 gene promoter and stimulates its transcription (Baek et al., 2002; Von Rotz et al., 2004). We sought to validate our animal model by examining the expression of KAI1 gene in the AICD transgenic mice. We probed the membrane fractions from nontransgenic littermate controls, two FeCγ transgenic lines, and Fe.27 animals with anti-KAI1 antibody (Fig. 3 A, top panel). A doublet of KAI1 protein band was visible in FeCγ.12 and FeCγ.25 mice (lanes 3–6), but not in nontransgenic controls (lanes 1 and 2) nor in Fe65 transgenic Fe.27 mice (lanes 7 and 8). When normalized for protein loading (Fig. 3 A, bottom panel), FeCγ.25 seem to express higher levels of KAI1 compared with FeCγ.12. These data confirm and extend in vivo the previous observations that AICD activates KAI1 gene expression (Baek et al., 2002; Von Rotz et al., 2004). These findings also validate the supposition that the γ-secretase cleaved, 59-residue long AICD exhibits biologic effects that are observed when APP is cleaved in vivo to generate AICD (Cao and Sudhof, 2001; Baek et al., 2002).


Activation of GSK-3 and phosphorylation of CRMP2 in transgenic mice expressing APP intracellular domain.

Ryan KA, Pimplikar SW - J. Cell Biol. (2005)

Increased KAI1 levels in AICD transgenic mice. (A) The AICD transgenic mice, but not the control or Fe.27 mice, show expression of KAI1 gene. Membrane fractions from indicated mice were probed with anti-KAI1 antibody. FeCγ.25 mice showed higher expression of KAI1 protein compared with FeCγ.12 mice. (B) Membrane fractions from indicated mice were probed with anti-SERCA 2b antibody. No significant changes in SERCA 2b levels were reproducibly observed in FeCγ transgenic mice.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171208&req=5

fig3: Increased KAI1 levels in AICD transgenic mice. (A) The AICD transgenic mice, but not the control or Fe.27 mice, show expression of KAI1 gene. Membrane fractions from indicated mice were probed with anti-KAI1 antibody. FeCγ.25 mice showed higher expression of KAI1 protein compared with FeCγ.12 mice. (B) Membrane fractions from indicated mice were probed with anti-SERCA 2b antibody. No significant changes in SERCA 2b levels were reproducibly observed in FeCγ transgenic mice.
Mentions: Although no bona fide target genes of AICD have been identified genetically, a chromatin immunoprecipitation assay was used recently to show that AICD is recruited to KAI1 gene promoter and stimulates its transcription (Baek et al., 2002; Von Rotz et al., 2004). We sought to validate our animal model by examining the expression of KAI1 gene in the AICD transgenic mice. We probed the membrane fractions from nontransgenic littermate controls, two FeCγ transgenic lines, and Fe.27 animals with anti-KAI1 antibody (Fig. 3 A, top panel). A doublet of KAI1 protein band was visible in FeCγ.12 and FeCγ.25 mice (lanes 3–6), but not in nontransgenic controls (lanes 1 and 2) nor in Fe65 transgenic Fe.27 mice (lanes 7 and 8). When normalized for protein loading (Fig. 3 A, bottom panel), FeCγ.25 seem to express higher levels of KAI1 compared with FeCγ.12. These data confirm and extend in vivo the previous observations that AICD activates KAI1 gene expression (Baek et al., 2002; Von Rotz et al., 2004). These findings also validate the supposition that the γ-secretase cleaved, 59-residue long AICD exhibits biologic effects that are observed when APP is cleaved in vivo to generate AICD (Cao and Sudhof, 2001; Baek et al., 2002).

Bottom Line: APP is cleaved by gamma-secretase that releases the APP intracellular domain (AICD) in the cytoplasm.In vitro studies have implicated AICD in cell signaling and transcriptional regulation, but its biologic relevance has been uncertain and its in vivo function has not been examined.Our data suggest that AICD is biologically relevant, causes significant alterations in cell signaling, and may play a role in axonal elongation or pathfinding.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Cell Biology Program, Case Western Reserve University, Cleveland, OH 44106, USA.

ABSTRACT
Amyloid precursor protein (APP), implicated in Alzheimer's disease, is a trans-membrane protein of undetermined function. APP is cleaved by gamma-secretase that releases the APP intracellular domain (AICD) in the cytoplasm. In vitro studies have implicated AICD in cell signaling and transcriptional regulation, but its biologic relevance has been uncertain and its in vivo function has not been examined. To investigate its functional role, we generated AICD transgenic mice, and found that AICD causes significant biologic changes in vivo. AICD transgenic mice show activation of glycogen synthase kinase-3beta (GSK-3beta) and phosphorylation of CRMP2 protein, a GSK-3beta substrate that plays a crucial role in Semaphorin3a-mediated axonal guidance. Our data suggest that AICD is biologically relevant, causes significant alterations in cell signaling, and may play a role in axonal elongation or pathfinding.

Show MeSH
Related in: MedlinePlus