_TINS July 2000 [final corr.] 12/6/00 10:39 am Page 291 R EVIEW Mechanisms of programmed cell death in the developing brain Chia-Yi Kuan, Kevin A. Roth, Richard A. Flavell and Pasko Rakic Programmed cell death (apoptosis) is an important mechanism that determines the size and shape of the vertebrate nervous system. Recent gene-targeting studies have indicated that homologs of the cell-death pathway in the nematode Caenorhabditis elegans have analogous functions in apoptosis in the developing mammalian brain.However,epistatic genetic analysis has revealed that the apoptosis of progenitor cells during early embryonic development and apoptosis of postmitotic neurons at later stage of brain development have distinct roles and mechanisms.These results provide new insight on the significance and mechanism of neural cell death in mammalian brain development. Trends Neurosci. (2000) 23, 291–297 ELL DEATH has long been recognized to occur in homologs of ced-3 comprise a family of cysteine- Cmost neuronal populations during normal devel- containing, aspartate-specific proteases called caspases5. opment of the vertebrate nervous system (reviewed in The ced-4 homolog is identified as one of the apoptosis Ref. 1). Traditionally, the investigation of neural death protease-activating factors (APAFs)6. The mammalian in development focused on the role of target-derived homologs of ced-9 belong to a growing family of Bcl2 survival factors such as NGF and related neurotrophins proteins, which share the Bcl2-homology (BH) domain (see Ref. 2 for a review). However, in the past few years, and are either pro- or anti-apoptotic7. The cloning of the genetic analysis of programmed cell death in the egl-1 indicates that it is similar to the BH3-domain- nematode Caenorhabditis elegans has inspired new ap- containing, pro-apoptotic subfamily of Bcl2 proteins4. proaches to study this phenomenon (see Box 1 for the What determines the fate of cell-autonomous apop- developmental functions of programmed cell death). tosis? Molecular cloning in C. elegans reveals that It is through such gene-targeting studies that recent ces-2 encodes a basic-leucine-zipper (bZIP) transcription- insights into the molecular regulation of mammalian factor motif, suggesting that programmed cell death programmed cell death have been obtained. could be regulated by differential gene expression8. In- deed, recent study suggests that the sex-determination The integration of specification and execution protein, TRA-1A, represses egl-1 transcription in the phases of cell death in C. elegans hermaphrodite-specific neurons (HSNs) and prevents During the development of an adult C. elegans her- apoptosis of these neurons, which would normally die maphrodite, 131 out of the total 1090 cells undergo in a male worm9. One intriguing implication of this programmed cell death in a lineage-specific and mostly study is that, although the anti-apoptotic (for example, Chia-Yi Kuan and cell-autonomous manner. Three groups of genes in- CED-9) and pro-apoptotic (for example, CED-3 and Pasko Rakic are at volved in this process have been identified by genetic CED-4) molecules are always present in living cells, the the Section of screening3. The first group of genes includes ces-1 and most-upstream molecule of the pathway (for example, Neurobiology, ces-2 (ces, cell-death specification) and affects the death EGL-1) could integrate various regulation signals to de- Yale University of specific types of cells. The second group of genes termine cell fate. Presumably, there are similar mecha- School of Medicine, affects most, if not all, of the 131 cells undergoing cell nisms in mammalian cells to integrate the regulatory New Haven, death, and is therefore involved in the execution phase signals at the initiation point of the apoptosis cas- CT 06510, USA, of cell death. These global regulators, which include cade. Such an integration point might reside at the tran- Kevin A. Roth is egl-1 (egl, egg-laying defective), ced-9 (ced, cell-death ab- scription level of the pro-apoptotic Bcl2-family genes. at the Dept of normal), ced-4 and ced-3, form an obligate cell-death Alternatively, the integration mechanisms might in- Pathology, pathway (Fig. 1a). The last group of genes, which in- clude posttranslational modifications or translocation of Washington cludes ced-1, ced-6, ced-7, ced-2, ced-5, ced-10 and nuc-1 the Bcl2-family proteins between cellular organelles. University School of (nuc, nuclease abnormal), is involved in the degradation Medicine, St. Louis, Caspase 9 and caspase 3 in the developing of DNA and phagocytosis of the cell corpses. Among MO 63110, USA, mammalian brain the three groups of cell-death genes, those involved in and Richard A. the execution phase of apoptosis have been the most The mammalian homologs of ced-3 comprise a fam- Flavell is at the extensively studied. Cumulative biochemical studies ily of cysteine-containing, aspartate-specific proteases Howard Hughes suggest that EGL-1 triggers programmed cell death by called caspases. These are present in living cells as Medical Institute, binding to CED-9 and thus releasing the cell-death acti- proenzymes that contain three domains: an N-termi- Section of vator CED-4 from a CED-9–CED-4 protein complex, nal domain, a large subunit and a small subunit. Immunobiology, which leads to activation of CED-3 (Ref. 4). Activation of caspases involves proteolytic processing Yale University Remarkably, structural homologs of all the genes in- between domains followed by association of the large School of Medicine, volved in the execution phase of cell death in C. elegans and small subunits to form an active heterodimer or New Haven, have been identified in mammals. The mammalian tetramer5. Once activated, caspases cleave other caspases CT 06510, USA. 0166-2236/00/$ – see front matter © 2000 Elsevier Science Ltd. All rights reserved. PII: S0166-2236(00)01581-2 TINS Vol. 23, No. 7, 2000 291 _TINS July 2000 [final corr.] 12/6/00 10:39 am Page 292 R EVIEW C-Y. Kuan et al. – PCD and development Box 1. Multiple functions of programmed cell death in brain development The biological functions of cell death during normal devel- the pro-apoptotic gene Bax have been generated, both of opment have captured the imagination of embryologists which exhibit increases in selected neuronal subpopulations since its discovery in the 19th centurya. In 1926, Ernst without gross malformations of the nervous systemg,h. These summarized three main types of developmental cell death: consequences of reduced developmental cell death, consis- the first occurring during regression of vestigial organs; the tent with the idea of social control of cell survival or death, second during morphogenesis of organ anlage; and the seemingly exclude a significant role of cell death in the third during remodeling of tissues. These were later named morphogenesis of the nervous system. phylogenetic, morphogenetic and histiogenetic degener- It was therefore surprising that mutant mice deficient in ation by Glucksmann in a scheme designed to explain the pro-apoptotic genes Casp3, Casp9 and Apaf1 all showed the biological functions of developmental cell deathb. severe malformations of the nervous system because of a Phylogenetic degeneration, exemplified by regression of reduction of developmental cell deathi–m. Moreover, mice pronephros and mesonephros in higher vertebrates, rarely with deficiency of the protein kinases Jnk1 and Jnk2 exhib- occurs in the developing nervous system. By contrast, cell ited pronounced neurulation defects, which were preceded death in many places of the embryonic brain, such as at by reduction of cell death prior to the closure of the hind- the edge of the neural plate and within the optic stalk was brain neural tube (Fig. I)n,o. Together, these findings strongly considered to be related to the morphogenetic process. The implicate a role of cell death during morphogenesis of the prime example of histiogenetic degeneration was described developing brain. The apparently discrepant consequences in the spinal ganglia of chick embryos. By quantitative and of reduced developmental cell death could be reconciled experimental methods, Hamburger and Levi-Montalcini by the demonstration of caspase-3-mediated, and Bax- showed that the spinal ganglia corresponding to the limbs and Bcl-XL-independent apoptosis of neural founder cells, are larger than the adjacent ganglia, and many ganglial followed by Bax- and Bcl-XL-regulated and caspase-3- neuroblasts degenerate if the limb bud is removed earlyc. dependent death of postmitotic neuronsp. These results suggested that neurons compete for a limited In summary, cell death shall no longer be considered supply of peripherally derived surviving ‘trophic’ factors merely a mechanism to match the neuron population to and their death is the consequence of an initially surplus its target fields. Rather, cell death has an addditional neuronal population. important role in adjusting the initial progenitor pool The subsequent discovery of NGF by Levi-Montalcini and needed for proper morphogenesis of the nervous system, colleagues consolidated the trophic theory, which became the mechanism of which remains to be investigated. one of the most-influential concepts of neural development of this centuryd. Consequently, the predominant view is References that cell death occurs mainly to match the size of each a Jacobson, M. (1991) Developmental Neurobiology (3rd edn), neuronal population to the magnitude of its target fields Plenum Press b Glucksmann, A. (1951) Cell deaths in normal vertebrate and to eliminate neurons with erroneous or inadequate ontogeny. Biol. Rev. 26, 59–86 projectionse,f. By contrast, the possibility that cell death c Hamburger, V. and Levi-Montalcini, R. (1949) Proliferation, can be a mechanism of morphogenesis of the nervous sys- differentiation and degeneration in the spinal ganglia of the tem has been relatively unexplored. In recent years, mutant chick embryo under normal and experimental conditions. J. Exp. Zool.