Intrinsic and extrinsic pathway signaling during neuronal apoptosis

Abstract

Trophic factor deprivation (TFD)-induced apoptosis in sympathetic neurons requires macromolecular synthesis–dependent BAX translocation, cytochrome c (cyt c) release, and caspase activation. Here, we report the contributions of other intrinsic and extrinsic pathway signals to these processes. Sympathetic neurons expressed all antiapoptotic BCL-2 proteins examined, yet expressed only certain BH3-only and multidomain proapoptotic BCL-2 family members. All coexpressed proapoptotic proteins did not, however, exhibit functional redundancy or compensatory expression, at least in the Bax −/−, Bak −/−, Bim −/−, Bid −/−, and Bad −/− neurons examined. Although the subcellular distribution or posttranslational modification of certain BCL-2 proteins changed with TFD, neither transcriptional nor posttranslational mechanisms regulated the expression or subcellular localization of BID, BAD, or BAK in this paradigm. Despite modest induction of Fas and FasL expression, Fas-mediated signaling did not contribute to TFD-induced apoptosis in sympathetic neurons. Similar findings were obtained with K+ withdrawal–induced apoptosis in cerebellar granule neurons, a model for activity-dependent neuronal survival in the CNS. Thus, expression alone does not guarantee functional redundancy (or compensation) among BCL-2 family members, and, at least in some cells, extrinsic pathway signaling and certain BH3-only proteins (i.e., BID and BAD) do not contribute to BAX-dependent cyt c release or apoptosis caused by TFD.

BH3-only proteins: key regulators of neuronal apoptosis

“…Both sympathetic neurons and CGNs express multiple proapoptotic Bcl-2 family proteins. [15][16][17][18] Of the multidomain proapoptotic proteins (Bax and Bak), Bax is essential for the release of mitochondrial cytochrome c in sympathetic neurons and for NGF withdrawal-induced death, and for the KCl/serum deprivationinduced death of CGNs, whereas Bak is not required. [18][19][20] This is an interesting observation because in other cell types the expression of both Bax and Bak must be lost to prevent cell death induced by survival factor withdrawal.…”

“…[15][16][17][18] Of the multidomain proapoptotic proteins (Bax and Bak), Bax is essential for the release of mitochondrial cytochrome c in sympathetic neurons and for NGF withdrawal-induced death, and for the KCl/serum deprivationinduced death of CGNs, whereas Bak is not required. [18][19][20] This is an interesting observation because in other cell types the expression of both Bax and Bak must be lost to prevent cell death induced by survival factor withdrawal. 21,22 This difference may be related to the fact that postnatal sympathetic neurons, hippocampal neurons and CGNs cultured in vitro exclusively express N-Bak, a neuron-specific splice variant of Bak, which is a BH3-only protein lacking the BH1 and BH2 domains of full-length Bak.…”

“…Bad À/À knockout mice have no gross abnormalities in the nervous system and bad À/À sympathetic neurons die at the same rate as wild-type neurons after NGF withdrawal. 18,53 However, Bad knockin mice have been constructed in which serines 112, 136 and 155 in Bad have been mutated to alanines so that the endogenous Bad protein cannot be phosphorylated (Bad 3SA mice). 54 Bad 3SA mice are viable and have no gross abnormalities although there are alterations in the normal development of pro-B and pro-T cells.…”

“…In the case of sympathetic neurons cultured in vitro, Bid is not cleaved after NGF deprivation and Bid À/À neurons die at the same rate after NGF withdrawal as wild-type neurons. 15,18 However, there is some evidence that Bid does have a role in cortical neuron death induced by oxygen/ glucose deprivation in vitro and following focal cerebral ischaemia in vivo. 67,68 In C57BL/6 mouse cortical neurons cultured in vitro oxygen/glucose deprivation leads to caspase-8 activation, Bid cleavage and apoptosis, and cortical neurons isolated from Bid À/À mouse brain are somewhat resistant to death induced by oxygen/glucose deprivation compared to wild-type neurons.…”

Apoptosome dependent caspase-3 activation pathway is non-redundant and necessary for apoptosis in sympathetic neurons

“…To assess the long-term survival of sympathetic neurons from Apaf-1, caspase-9, and caspase-3 null mice in a rigorous manner, we examined their ability to recover from NGF deprivation following the release of cytochrome c. Following 24 h of NGF deprivation, approximately 90% of WT, Apaf-1, caspase-9, and caspase-3 null sympathetic neurons had released cytochrome c (Figure 1c), 25,26 and the WT neurons had already died by apoptosis (Figure 1a). To assess longterm recoverability following the release of cytochrome c, cultures were deprived of NGF for 24-30 h, following which NGF was restored to the cultures and survival was assessed following 7 days.…”