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The Caenorbabditis elegans genes ced-3 and ced-4 act cell autonomously to cause programmed cell death
~ONITOR The Caemorhabditis elegams genes ced.3 and ced.4 act cell autonomously to cause programmedcell death J. YUAN AND H.R. HORVITZ
Dev. Biol. 138, 33--41 Of the 1090 somatic cells generated during the development of the nematode C. elegans, 131 are programmed to die soon after their births. What marks out some cells for death, and then how do the doomed cells die? Yuan and Horvitz have begun to look for answers to these questions by genetic mosaic analysis of the genes ced-3 and ced-4. Mutations in these genes prevent almost all cell deaths, indicating that these genes act to cause cell death. Clearly, it is important to
Large restriction fragments containing poly.TG are highly polymorphic in a variety of vertebrates Y. KASHI E T A L .
Nucleic Acids Res. 18,
1129-1132 The reiterated, tandemly repeated 'minisatellite' sequences of higher eukaryotic genomes are famous as the basis for the technique of DNA fingerprinting developed by Jeffreys and colleagues. Shorter, repetitive 'micro.~atellite' sequences also exist, most notably the poly-(TG) family, which in humans accounts for about 0.04% of the total DNA. Kashi et al. were able to obtain DNA fingerprints in a number of higher vertebrates (chicken, sheep, horse, mouse, human) when they used as a probe a bovine DNA clone consisting c f seven tracts of poly-(TG) repeats separated by a short linker sequence. In chicken, the fingerprint bands v, ere inherited in a mendelian fashion. Variable fingerprints were only obtained when the genomic DNA was digested with restriction enzymes having 4 bp recognition sequences, presumably because these conditions left intact large fragments containing clusters of poly-(TG) repeats. Although most if not all poly-(TG)-containing loci in higher vertebrates show some length variation, possibly caused by replication slippage, Koshi et al. suggest that their probes pick up only gross length variation resulting from unequal crossing over. In view of the regulatory functions that have been proposed for poly-(TG) sequences, variation in this class of microsatellite sequences might have functional and perhaps evolutionary implications, a~
know whether ced-3 and ced-4 act within dying cells, or within cells that cause dying cells to die. To answer this question, strains were generated that were homozygous for either ced-3 or ced-4, but carried the corresponding wild-type allele on a free chromosomal duplication. Free duplications are often lost at cell division, and so mosaic animals result, in which the mutant phenotype is unmasked in some cells. Since the entire cell division pattern in C. elegans is known, one can pinpoint the division at which duplication loss occurred, and thus establish the genotype of every cell. If the ced genes act cell-autonomously, a given cell that would normally die in wild-type animals will only
Position-specific activity of the nox 1.1 promoter in transgenic mice A.W. POSCHEL, R. BALL1NG AND P. GRUSS
Development 108, 435-442 Homeobox genes have been assumed to be important in mouse development on the basis of their similarity to genes implicated in Drosophila development and their ordered spatial and temporal patterns of expression. Such patterns are probably generated by controlling gene expression. Transgenic mice offer a powerful tool for determining which features of the genes are important for which aspect of this regulation. The transgene used by P0schel et al. consisted of lacZ (for easy determination of expression
Serum and v.src increase the level of a CCAAT-bindingfactor required for transcription from a retroviral long terminal repeat A. DUTFA, M.Y. STOECKLE AND H. HANAFUSA
Genes Dev. 4, 243-254 Factors binding to the 'CCAAT box' found in the enhancers of a number of mammalian and viral genes may function in a novel signal transduction pathway. This is the surprising conclusion from the experiments reported by Dutta et aL, who began by examining whether serum would stimulate transcription from the long terminal repeat (LTR) of Rous sarcoma virus (RSV). It did, and stimulation was found to depend on binding of a nuclear protein to two CCAAT box sequences in the LTR. New protein synthesis was not required for stimulation. The CCAAT factor involved was identified by oligonucleot~de binding experiments as belonging to the CP1,2 class of TIG MAY 1 9 9 0 VOL. 6 NO. 5
m
survive in a mosaic if the cell is ced in genotype, regardless of the genotype of surround,.'ng cells. This was indeed found to be the case, both for ced-3 and ced-4. Since this result applied to cell deaths in a number of different lineages, it seems likely that ced-3 + and ced-4 + themselves act to kill either the cell in which they are expressed or perhaps one of its daughter cells. This result is of broad interest, since recent observations in a variety of experimental systems have shown that cell death is an active process depending on de novo gene expression in the cells that die. Understanding how ced-3 and ced-4 work may prove relevant to cell death in mammals and, perhaps, in human disease, a~ patterns) flanked by 3.6 kb of 5' and 1.7 kb of 3' Hox 1.1 sequences. Endogenous Hox 1.1 is expressed in mouse embryogenesis between days 7.5 and 14; the transgenic promoter sequences faithfully reproduced the expression pattern between days 7.5 and 8.5, but subsequent expression differed. In this phase of expression, although some features of Hox 1.1 are mimicked (e.g. the anterior boundary is the same), others are not (e.g. the posterior boundary). Regulation in this second phase, during which expression becomes restricted to a specific subregion of the original area, could be mediated through sequences farther upstream/downstream, or possibly in the Hox 1.1 intron. factors, which includes the factors that bind to the CCAAT boxes in the human Hsp70 and c-Ha-ms genes. The fact that the CCAAT factor seems to be directly responsible for mediating the signal from serum suggests that it is activated in some way by serum; in view of the large number of genes whose transcription is stimulated by serum, it is tempting to suggest that the CCAAT factor, with its binding sites in many cellular promoters, may be widely implicated. The dual role of the CCAAT factor in both constitutive transcription and serum induction may also hint at a mechanism for the wide-ranging effects of a number of oncogenes. Indeed, Dutta et aL found that stimulation of transcription from the RSV LTR occurred in the absence of serum in v-src-transformed cells; stimulation depended on the tyrosine kinase activity of pp60 v-src and was associated with increased amounts of the CCAAT factor. ,~
Dev. Biol. 138, 33--41 Of the 1090 somatic cells generated during the development of the nematode C. elegans, 131 are programmed to die soon after their births. What marks out some cells for death, and then how do the doomed cells die? Yuan and Horvitz have begun to look for answers to these questions by genetic mosaic analysis of the genes ced-3 and ced-4. Mutations in these genes prevent almost all cell deaths, indicating that these genes act to cause cell death. Clearly, it is important to
Large restriction fragments containing poly.TG are highly polymorphic in a variety of vertebrates Y. KASHI E T A L .
Nucleic Acids Res. 18,
1129-1132 The reiterated, tandemly repeated 'minisatellite' sequences of higher eukaryotic genomes are famous as the basis for the technique of DNA fingerprinting developed by Jeffreys and colleagues. Shorter, repetitive 'micro.~atellite' sequences also exist, most notably the poly-(TG) family, which in humans accounts for about 0.04% of the total DNA. Kashi et al. were able to obtain DNA fingerprints in a number of higher vertebrates (chicken, sheep, horse, mouse, human) when they used as a probe a bovine DNA clone consisting c f seven tracts of poly-(TG) repeats separated by a short linker sequence. In chicken, the fingerprint bands v, ere inherited in a mendelian fashion. Variable fingerprints were only obtained when the genomic DNA was digested with restriction enzymes having 4 bp recognition sequences, presumably because these conditions left intact large fragments containing clusters of poly-(TG) repeats. Although most if not all poly-(TG)-containing loci in higher vertebrates show some length variation, possibly caused by replication slippage, Koshi et al. suggest that their probes pick up only gross length variation resulting from unequal crossing over. In view of the regulatory functions that have been proposed for poly-(TG) sequences, variation in this class of microsatellite sequences might have functional and perhaps evolutionary implications, a~
know whether ced-3 and ced-4 act within dying cells, or within cells that cause dying cells to die. To answer this question, strains were generated that were homozygous for either ced-3 or ced-4, but carried the corresponding wild-type allele on a free chromosomal duplication. Free duplications are often lost at cell division, and so mosaic animals result, in which the mutant phenotype is unmasked in some cells. Since the entire cell division pattern in C. elegans is known, one can pinpoint the division at which duplication loss occurred, and thus establish the genotype of every cell. If the ced genes act cell-autonomously, a given cell that would normally die in wild-type animals will only
Position-specific activity of the nox 1.1 promoter in transgenic mice A.W. POSCHEL, R. BALL1NG AND P. GRUSS
Development 108, 435-442 Homeobox genes have been assumed to be important in mouse development on the basis of their similarity to genes implicated in Drosophila development and their ordered spatial and temporal patterns of expression. Such patterns are probably generated by controlling gene expression. Transgenic mice offer a powerful tool for determining which features of the genes are important for which aspect of this regulation. The transgene used by P0schel et al. consisted of lacZ (for easy determination of expression
Serum and v.src increase the level of a CCAAT-bindingfactor required for transcription from a retroviral long terminal repeat A. DUTFA, M.Y. STOECKLE AND H. HANAFUSA
Genes Dev. 4, 243-254 Factors binding to the 'CCAAT box' found in the enhancers of a number of mammalian and viral genes may function in a novel signal transduction pathway. This is the surprising conclusion from the experiments reported by Dutta et aL, who began by examining whether serum would stimulate transcription from the long terminal repeat (LTR) of Rous sarcoma virus (RSV). It did, and stimulation was found to depend on binding of a nuclear protein to two CCAAT box sequences in the LTR. New protein synthesis was not required for stimulation. The CCAAT factor involved was identified by oligonucleot~de binding experiments as belonging to the CP1,2 class of TIG MAY 1 9 9 0 VOL. 6 NO. 5
m
survive in a mosaic if the cell is ced in genotype, regardless of the genotype of surround,.'ng cells. This was indeed found to be the case, both for ced-3 and ced-4. Since this result applied to cell deaths in a number of different lineages, it seems likely that ced-3 + and ced-4 + themselves act to kill either the cell in which they are expressed or perhaps one of its daughter cells. This result is of broad interest, since recent observations in a variety of experimental systems have shown that cell death is an active process depending on de novo gene expression in the cells that die. Understanding how ced-3 and ced-4 work may prove relevant to cell death in mammals and, perhaps, in human disease, a~ patterns) flanked by 3.6 kb of 5' and 1.7 kb of 3' Hox 1.1 sequences. Endogenous Hox 1.1 is expressed in mouse embryogenesis between days 7.5 and 14; the transgenic promoter sequences faithfully reproduced the expression pattern between days 7.5 and 8.5, but subsequent expression differed. In this phase of expression, although some features of Hox 1.1 are mimicked (e.g. the anterior boundary is the same), others are not (e.g. the posterior boundary). Regulation in this second phase, during which expression becomes restricted to a specific subregion of the original area, could be mediated through sequences farther upstream/downstream, or possibly in the Hox 1.1 intron. factors, which includes the factors that bind to the CCAAT boxes in the human Hsp70 and c-Ha-ms genes. The fact that the CCAAT factor seems to be directly responsible for mediating the signal from serum suggests that it is activated in some way by serum; in view of the large number of genes whose transcription is stimulated by serum, it is tempting to suggest that the CCAAT factor, with its binding sites in many cellular promoters, may be widely implicated. The dual role of the CCAAT factor in both constitutive transcription and serum induction may also hint at a mechanism for the wide-ranging effects of a number of oncogenes. Indeed, Dutta et aL found that stimulation of transcription from the RSV LTR occurred in the absence of serum in v-src-transformed cells; stimulation depended on the tyrosine kinase activity of pp60 v-src and was associated with increased amounts of the CCAAT factor. ,~