The chemokine CXCL12 promotes survival of postmitotic neurons by regulating Rb protein

Abstract

Postmitotic neurons need to keep their cell cycle under control to survive and maintain a differentiated state. This study aims to test the hypothesis that the chemokine CXCL12 regulates neuronal survival and differentiation by promoting Rb function, as suggested by previous studies showing that CXCL12 protects neurons from apoptosis induced by Rb loss. To this end, the effect of CXCL12 on Rb expression and transcriptional activity and the role of Rb in CXCL12-induced neuronal survival were studied. CXCL12 increases Rb protein and RNA levels in rat cortical neurons. The chemokine also stimulates an exogenous Rb promoter expressed in these neurons and counteracts the inhibition of the Rb promoter induced by E2F1 overexpression. Furthermore CXCL12 stimulates Rb activity as a transcription repressor. The effects of CXCL12 are mediated by its specific receptor CXCR4, and do not require the presence of glia. Finally, shRNA studies show that Rb expression is crucial to the neuroprotective activity of CXCL12 as indicated by NMDA-neurotoxicity assays. These findings suggest that proper CXCR4 stimulation in the mature CNS can prevent impairment of the Rb-E2F pathway and support neuronal survival. This is important to maintain CNS integrity in physiological conditions and prevent neuronal injury and loss typical of many neurodegenerative and neuroinflammatory conditions. Cell Death and Differentiation ( 1,2 Stimulation of CXCR4 by its endogenous ligand leads to activation of intracellular pathways affecting neuronal survival, migration, and neurotransmission.1,3 For instance, CXCR4 stimulates the PI3K/Akt pathway and regulates cell-cycle proteins in neurons. [3][4][5][6] Under pathological conditions at least some of these essential CXCR4 functions are compromised, leading to neuronal dysfunction/death. 1,7 Thus, a complete understanding of the effects of CXCR4 activation in the brain has important physiological and pathological implications.Rb is a well-known transcriptional repressor, which controls cell-cycle progression, differentiation/survival, and genomic integrity. Rb is also implicated in fundamental CNS developmental processes such as, neuronal migration, differentiation, and neurite extension. 8,9 The effects of Rb on cell survival are primarily due to regulation of members of the E2F family of transcription factors, though Rb can also directly inactivate proapoptotic proteins, such as pp32.10 E2F proteins stimulate expression of genes promoting cell-cycle progression in proliferating cells, whereas they induce apoptotic genes in differentiated cells, including neurons.11 Rb is phosphorylated at several phosphoacceptor sites by specific kinases. 12 Hence, the immediate effect of Rb phosphorylation is to remove Rb from its specific promoters, leading to inhibition of Rb-dependent gene repression. Rb phosphorylation results in enhanced E2F1 transcriptional activity culminating in death of postmitotic neurons.11 Alterations of Rb/E2F pathway were reported in various neuropathologies, such as Parkinson's and Alzhei...

Neuronal ferritin heavy chain and drug abuse affect HIV-associated cognitive dysfunction

“…Selected experiments were performed using glia-free neuronal cultures as indicated, using previously described protocols (27). Neurons were transfected at the time of plating by Nucleofection (Amaxa), or after 8 days in culture by lipofectamine (Invitrogen), as described previously (4,27). Unless otherwise indicated, all experiments were conducted at 21 DIV.…”

“…The chemokine CXCL12 and its cognate receptor CXCR4 perform multiple functions essential for CNS development and function, including guiding migration and differentiation of neuronal precursor cells (1), regulating cell cycle proteins (2)(3)(4), and modulating NMDA receptor signaling (5). These homeostatic functions take on added significance within the context of neuroinflammatory disease, as the CXCL12/CXCR4 signaling axis aids recovery by promoting survival of mature neurons, recruitment of neural and glial progenitor cells (6)(7)(8), and neuronal-glial communication (9).…”

Interactions between Chemokines

“…Neurons were harvested from brain cortices of E17/18 Holtzman rat embryos as previously described (27,29,30). Cells were plated as reported below.…”

“…Furthermore, data from different groups have shown that SDF-1 and fractalkine are both involved in neuroprotection and regulate excitatory neurotransmission (6,24,25). However, while the neuroprotective effect of fractalkine seems to be related to its ability to counteract microglia-induced neurotoxicity, SDF-1 can directly protect neurons via the activation of its specific receptor CXCR4, which is coupled to major survival pathways, such as the phosphoinositide 3-kinase (PI3K)-Akt pathway (26,27). Interestingly, intracellular pathways stimulated by SDF-1 control transcription factors that up-regulate expression of the fractalkine gene, such as NFB and p53 (28).…”

Bilaminar Co-culture of Primary Rat Cortical Neurons and Glia

“…Neurons can be easily separated from the glial layer at any time during culture and used for different experimental applications ranging from electrophysiology 4 , cellular and molecular biology 5-8 , biochemistry 5 , imaging and microscopy 4,6,7,9,10 . The primary neurons extend axons and dendrites to form functional synapses 11 , a process which is not observed in neuronal cell lines, although some cell lines do extend processes.…”