Bag1 is essential for differentiation and survival of hematopoietic and neuronal cells

Abstract

Bag1 is a cochaperone for the heat-shock protein Hsp70 that interacts with C-Raf, B-Raf, Akt, Bcl-2, steroid hormone receptors and other proteins. Here we use targeted gene disruption in mice to show that Bag1 has an essential role in the survival of differentiating neurons and hematopoietic cells. Cells of the fetal liver and developing nervous system in Bag1−/− mice underwent massive apoptosis. Lack of Bag1 did not disturb the primary function of Akt or Raf, as phosphorylation of the forkhead transcription factor FKHR and activation of extracellular signal–regulated kinase (Erk)-1/2 were not affected. However, the defect was associated with the disturbance of a tripartite complex formed by Akt, B-Raf and Bag1, in addition to the absence of Bad phosphorylation at Ser136. We also observed reduced expression of members of the inhibitor of apoptosis (IAP) family. Our data show that Bag1 is a physiological mediator of extracellular survival signals linked to the cellular mechanisms that prevent apoptosis in hematopoietic and neuronal progenitor cells.

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Figure 1: Targeted disruption of Bag1 by homologous recombination.
Figure 2: Bag1 mutant embryos show severe defects in the forebrain and liver.
Figure 3: Differentiation and survival of spinal motoneurons is impaired in Bag1−/− embryos.
Figure 4: Enhanced apoptosis of differentiating neurons from Bag1-mutant neural stem cells.
Figure 5: Enhanced apoptosis of differentiating neurons from spinal cords of Bag1 mutants.
Figure 6: Specific lack of Bad phosphorylation at Ser136 is not caused by inhibition of MAPK and Akt in Bag1−/− embryos.
Figure 7: Bag1 is necessary to form a complex of Bag1, Hsp70, Raf and Akt.
Figure 8: Loss of Bag1 leads to changes in mitochondrial localization of Akt and Raf in isolated motoneurons.

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Acknowledgements

We thank M. Pfister, J. Marcano and K. Kalus for excellent technical assistance; R. McKay (US National Institutes of Health) for providing the nestin-specific antibody; T. Jessell for providing the islet-1/2 hybridoma (39.4D5) through the Developmental Studies Hybridoma Bank; and R. Timpl for the kind donation of laminin. This work was supported by the Deutsche Forschungsgemeinschaft (SE 697/3-3, SFB487, TP C4 and SFB581 TP B4 and B17), the US National Institutes of Health (CA67385) and the Hermann and Lilly Schilling Foundation.

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Correspondence to Michael Sendtner.

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Supplementary information

Supplementary Fig. 1

Activated caspase 3 immunoreactivity is increased in E10.5 and E11.5 bag-1−/− embryonic forebrain. (PDF 7732 kb)

Supplementary Fig. 2

Enhanced apoptosis of differentiating neurons from bag-1−/− Pax6-positive neural stem cells. (PDF 3237 kb)

Supplementary Fig. 3

Knockdown of Bag-1 by RNAi results in loss of Bag-1 immunohistochemistry. (PDF 775 kb)

Supplementary Fig. 4

Enhanced apoptosis of differentiating neurons in the forebrain of bag-1−/− mice. (PDF 8622 kb)

Supplementary Fig. 5

Model for the interaction of a complex formed by Bag-1, Hsp70, B-Raf and Akt and its substrate Bad in wild-type mice and disturbance of this process in bag-1−/− mice. (PDF 157 kb)

Supplementary Table 1

Number of apoptotic cells in spinal cord and brain sections of bag-1+/+ and bag-1−/− mice. (PDF 36 kb)

Supplementary Methods (PDF 96 kb)

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Götz, R., Wiese, S., Takayama, S. et al. Bag1 is essential for differentiation and survival of hematopoietic and neuronal cells. Nat Neurosci 8, 1169–1178 (2005). https://doi.org/10.1038/nn1524

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