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The Rab5-Rab11 endosomal pathway is required for BDNF-induced CREB transcriptional regulation in neurons

Andrés G. González, Oscar M. Lazo, Francisca C. Bronfman

Preprint posted on 16 November 2019 https://www.biorxiv.org/content/10.1101/844720v1

Article now published in The Journal of Neuroscience at http://dx.doi.org/10.1523/JNEUROSCI.2063-19.2020

BDNF/TrkB utilizes Rab5/Rab11 signaling endosomes to induce CREB phosphorylation and immediate gene expression in neurons

Selected by Theresa Pohlkamp

Background

The brain derived neurotrophic factor BDNF regulates neuron morphology, connectivity, activity, and survival through the tyrosine receptor kinase TrkB. Dysfunction of the BDNF/TrkB pathway is implicated in various neurodevelopmental and neurodegenerative disorders. BDNF binding to and activation of TrkB involves receptor dimerization and induces three downstream cascades through (1) PI3K/Akt, (2) MAPK/ERK and (3) PLCƴ.

TrkB with BDNF also undergoes endocytosis to control early endosomal membrane trafficking (Fu et al., 2011), which is important for BDNF/TrkB function in neuroprotection (Burk et al., 2017) and dendritic growth (Moya-Alvarado et al., 2018). Members of the Rab family of small GTPases define the identity of endosomal compartments while they are maturing and sorted through the endolysosomal system: Rab5 labels endocytosed vesicles and early endosomes that further mature to fast (Rab4) and slow (Rab11) recycling endosomes or late (Rab7) endosomes. Importantly, a number of studies indicated that neurotrophic-receptor mediated signaling regulates endosomal signaling and vice versa.

Previously, the laboratory of Francisca Bronfman found that internalized BDNF localizes to Rab5 and Rab11 endosomes and regulates their dynamics and function to control dendritic branching (Lazo et al., 2013; Moya-Alvarado et al., 2018). In their current preprint the authors analyze the role of Rab5 and Rab11 vesicles in BDNF mediated regulation of nuclear signaling and gene transcription.

 

Findings

The authors first showed that they are able to alter BDNF/TrkB signaling in primary neurons when disrupting the endosomal pathway by the overexpression of dominant negative (DN) Rab5 or Rab11. They treated Rab5DN, Rab11DN, or EGFP (control) overexpressing neurons with BDNF and analyzed effects on signaling five minutes or two hours after. Even though BDNF-induced phosphorylation of TrkB, Akt, and ERK1/2 was comparably strong in all three groups after five minutes (early activation). The amount of phosphorylated proteins declined after two hours of treatment, which was enhanced for all three targets when Rab5DN or Rab11DN were overexpressed. They further examined ERK1/2 activation after one hour of BDNF treatment and found by immunocytochemistry that in control cells, phosphorylated ERK1/2 localized to the nuclei, cell body, and dendrites. Rab5DN, more than Rab11DN, reduced phosphorylation of ERK1/2 in all three subcellular regions. To assess whether transcription was also affected, they analyzed the phosphorylation status of the transcription factor CREB, which was induced by BDNF after 15 minutes, persisting for an hour. When Rab5DN or Rab11DN were overexpressed, CREB phosphorylation was not detected after 15 or 30 minutes of BDNF treatment and only reduced levels of phosphorylation were measured after one hour of treatment.

BDNF induced phosphorylation of CREB was prevented by the application of pERK1/2 and Trk inhibitors. Importantly, the PI3K inhibitor LY294002, whose activity was demonstrated by the prevention of BDNF mediated Akt phosphorylation (pS473), did not affect CREB phosphorylation. In agreement with previous studies (Finsterwald et al., 2010), overexpression of a dominant negative form of CREB (S133A) prevented BDNF induced dendritic branching. Previous studies also demonstrate that Rab5DN and Rab11DN reduce dendritic branching (Moya-Alvarado et al., 2018). Thus, the authors conclude that BDNF/TrkB signaling requires the MAPK/ERK pathway but not the PI3K/Akt pathway to regulate nuclear signaling and dendritic branching. To further analyze neuronal gene expression alterations upon disrupting the endosomal system, the authors focused on Rab11. BDNF signaling and CREB activation leads to enhanced expression in neurons, which was reduced by Rab11DN. The mRNA transcripts of several other CREB-regulated immediate early genes were also altered by Rab11DN. In summary, Rab5/Rab11 signaling endosomes propagate BDNF/TrkB to the nucleus to phosphorylate CREB and activate immediate early genes in an ERK1/2 dependent fashion.

 

Why I chose this preprint

The endolysosomal system is of critical importance to control neuronal plasticity by regulating the quantity, diversity, and distribution of surface receptors on neurons. GWAS studies demonstrate that Alzheimer’s disease risk genes, particularly of the more common late-onset form, are commonly related to endocytic transport regulations. Apolipoprotein E isoforms differentially affect endosomal trafficking (Nuriel et al., 2017) and ApoE4, the most important genetic risk factor for Alzheimer’s disease, prevents Reelin mediated Apoer2 and glutamate receptor surface recruitment, which affects synaptic plasticity (Chen et al., 2010). Therefore, a broader understanding of endosomal trafficking and signaling in neurons in health and disease is of critical importance.

Rab family GTPases play an important role in defining the identity of endosomes, allowing them to mature for recycling, sorting, transport, or degradation of the respective compartments and/or their contents. Overall, several members of the Rab family (Rab4,5,6,7a,10,11,27) have been linked to Alzheimer’s disease (Guadagno and Progida, 2019). Rab10, which functions as regulator between early and recycling endosomes (Chen et al., 2006), has been identified as an Alzheimer’s resilience locus (Tavana et al., 2018), and Rab10 phosphorylation was identified as a pathological feature in Alzheimer’s disease (Yan et al., 2018). The preprint co-author Dr. Oscar Lazo recently presented a poster suggesting a mechanism in which Rab10 regulates TrkB sorting to retrograde axonal transport (Lazo and Schiavo, 2019). Intense studies on BDNF/TrkB signaling provide a perfect window into understanding the transport dynamics of endosomes which potentially translate to other ligand-receptor systems, like ApoE/Reelin/Apoer2. Interestingly, Reelin and BDNF share downstream effectors including PI3K, Akt, ERK1/2 and CREB. Furthermore, both control dendritic branching and the expression of immediate genes like Arc. Overall, the precise endocytic and endosomal regulatory functions of Rab family members need further investigation to improve the understanding of the implication of the endolysosomal system in neurodegeneration and neurological disorders to help uncover new therapeutic targets.

 

Questions for the authors:

  1. You demonstrated that BDNF mediated phosphorylation of CREB is dependent on Trk and ERK, but independent of PI3K. Do you think that p75 or PLCƴ could cross-interact in this pathway? Especially since BDNF/TrkB induced PLCƴ acts on CREB phosphorylation through Calcium/Calmodulin?
  2. In Figure 5B and C you compare the fold change of transcripts after BDNF treatment. Did you also look at the baseline of EGFP and Rab11DN neurons without BDNF treatment? Are there any differences in the tested transcripts?
  3. Signaling endosomes are important to spatiotemporally propagate neurotrophin signaling. Do you think Rab5/11 endosomes provide a general pathway after receptor mediated endocytosis to induce endosomal signaling in neurons?

 

References

Burk, K., Murdoch, J.D., Freytag, S., Koenig, M., Bharat, V., Markworth, R., Burkhardt, S., Fischer, A., Dean, C., 2017. EndophilinAs regulate endosomal sorting of BDNF-TrkB to mediate survival signaling in hippocampal neurons. Sci. Rep. 7, 2149. https://doi.org/10.1038/s41598-017-02202-4

Chen, C.C.-H., Schweinsberg, P.J., Vashist, S., Mareiniss, D.P., Lambie, E.J., Grant, B.D., 2006. RAB-10 is required for endocytic recycling in the Caenorhabditis elegans intestine. Mol. Biol. Cell 17, 1286–1297. https://doi.org/10.1091/mbc.e05-08-0787

Chen, Y., Durakoglugil, M.S., Xian, X., Herz, J., 2010. ApoE4 reduces glutamate receptor function and synaptic plasticity by selectively impairing ApoE receptor recycling. Proc. Natl. Acad. Sci. U. S. A. 107, 12011–12016. https://doi.org/10.1073/pnas.0914984107

Finsterwald, C., Fiumelli, H., Cardinaux, J.-R., Martin, J.-L., 2010. Regulation of dendritic development by BDNF requires activation of CRTC1 by glutamate. J. Biol. Chem. 285, 28587–28595. https://doi.org/10.1074/jbc.M110.125740

Fu, X., Yang, Y., Xu, C., Niu, Y., Chen, T., Zhou, Q., Liu, J.-J., 2011. Retrolinkin cooperates with endophilin A1 to mediate BDNF-TrkB early endocytic trafficking and signaling from early endosomes. Mol. Biol. Cell 22, 3684–3698. https://doi.org/10.1091/mbc.E11-04-0308

Guadagno, N.A., Progida, C., 2019. Rab GTPases: Switching to Human Diseases. Cells 8. https://doi.org/10.3390/cells8080909

Lazo, O.M., Gonzalez, A., Ascaño, M., Kuruvilla, R., Couve, A., Bronfman, F.C., 2013. BDNF regulates Rab11-mediated recycling endosome dynamics to induce dendritic branching. J. Neurosci. Off. J. Soc. Neurosci. 33, 6112–6122. https://doi.org/10.1523/JNEUROSCI.4630-12.2013

Lazo, O.M., Schiavo, G., 2019. Rab10 as a novel target to manipulate axonal transport of neurotrophic signalling endosomes.

Moya-Alvarado, G., Gonzalez, A., Stuardo, N., Bronfman, F.C., 2018. Brain-Derived Neurotrophic Factor (BDNF) Regulates Rab5-Positive Early Endosomes in Hippocampal Neurons to Induce Dendritic Branching. Front. Cell. Neurosci. 12, 493. https://doi.org/10.3389/fncel.2018.00493

Nuriel, T., Peng, K.Y., Ashok, A., Dillman, A.A., Figueroa, H.Y., Apuzzo, J., Ambat, J., Levy, E., Cookson, M.R., Mathews, P.M., Duff, K.E., 2017. The Endosomal–Lysosomal Pathway Is Dysregulated by APOE4 Expression in Vivo. Front. Neurosci. 11. https://doi.org/10.3389/fnins.2017.00702

Tavana, J.P., Rosene, M., Jensen, N.O., Ridge, P.G., Kauwe, J.S., Karch, C.M., 2018. RAB10: an Alzheimer’s disease resilience locus and potential drug target. Clin. Interv. Aging 14, 73–79. https://doi.org/10.2147/CIA.S159148

Yan, T., Wang, L., Gao, J., Siedlak, S.L., Huntley, M.L., Termsarasab, P., Perry, G., Chen, S.G., Wang, X., 2018. Rab10 Phosphorylation is a Prominent Pathological Feature in Alzheimer’s Disease. J. Alzheimers Dis. JAD 63, 157–165. https://doi.org/10.3233/JAD-180023

Tags: alzheimer's disease, bdnf, endocytosis, nuclear signaling, primary neurons, recycling, reelin, synaptic plasticity

Posted on: 20 December 2019

doi: https://doi.org/10.1242/prelights.15926

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