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Centrosome duplication continues in cycloheximide-treated Xenopus blastulae in the absence of a detectable cell cycle
~ONITOR
A 'digest' of some recent papers of interest in the primary journals.
Deletions and a translocation interrupt a cloned gene at the Neurofi!iromatosis Type I locus D. VISKOCHIL ETAL.
Cell 62, 187-192
A major segment of the Neurofibgomatosis ~pe I gene: cDNA sequence, genomic structure, and point mutations k.M. CAWTHON ETAL.
Cell 62, 193-201
.Type 1 Neuroflbromatosis gene: identification of a large transcript disrupted in three NF1 patients M.R. WALLACEETAL.
Science 249, 181-186
Two groups have reported finding a gene that is a strong candidate for the neurofibromatosis type 1 (NF1) locus. NF1 is an autosomal dominant disorder that mainly affects neural crest-derived tissues. Within the last few years, two NFl-associated chromosomal translocations [t(l:17) and t(17:22)] have been found that placed the locus within a 600 kb region on chromosome 17q11.2, and three genes (EVI2, OMGP and RC1) have been identified between the translocation breakpoints, but NFl-specific mutations have not been detected in any of these genes. By adjusting the conditions of pulsed-fi¢.ld gel electrophoresis so as to improve resolution of small DNA fragments, Viskochil et aL were able to identify three NFl-associated deletions, one of which removed Olfly --11 i,:b of DNA in the t(17:22) breakpoint region. Once again, the strategy of using sequence conservation as an indication of the presence of a gene has proved successful: a probe from this region identified a sequence
Protein synthesis and the cell cycle: centrosome reproductionin set, urchin eggs is not under translationalcontrol G. SLUDER, F.J. MILLER, R. COLE AND C.L. RIEDER
J. Cell Biol. 110, 202.r~--2032
Centrosome duplication continues in cyclohex~!de.treated Xesopus
blastulae in the absence of a detectable cell cycle D.L. GARD, S. HAFEZI, T. ZHANG AND S.J. DOXSEY
J. Cell BioL 110, 2033-2042
The centrosome is a key component of the mitotic apparatus, but information has been scant about what controls its replication before mitosis,
from a murine cDNA library, and this cDNA in turn picked up sequences in several human cDNA libraries. Independently, Wallace et al. used the same sequence-conservation strategy, but obtained their probe to the translocation breakpoint region by chromosome jumping; another probe to screen human cDNA libraries was provided by a yeast artificial chromosome (YAC) clone from a YAC contig spanning the region. Both groups arrive at the same conclusion: that their cDNAs come from a large gene spanning both translocation breakpoints, and including within one of its introns all three of the previous candidate genes. The gene is expressed in a wide range of tissue types. Sequencing of the exons represented by the cDNAs obtained so far reveals no homology to any other known gene. Crucially, both groups have identified, in NF1 patients, point mutations in putative exons of the gene, strengthening the assertion that it is indeed the NF1 gene. Wallace et al. in Science and Cawthon et ai. in Cell point out that the dominant inheritance of NFI needs to be reconciled with the fact that each of the mutations detected so far would be expected to inactivate the NF1 gene. Perhaps, as is the case for retinoblastoma, tumours only develop when the normal NF1 allele is also inactivated by somatic mutation. The large size and complex organization of the NF1 region also raise intriguing questions about its regulation, with the possibility that some of the symptoms of NFI may be due to effects of mutations in the NF1 gene on expression of one or more of the oppositely transcribed EVI2, OMGPand RC1 genes, ah
CpG islands are clusters of CpG dinucleotides that are undermethylated, compared with the high methylation status of most CpG sequences in mammalian genomes. CpG islands can be either tissue specific, when they reflect the presence of active genes, or ubiquitous. Since de novo methylation activity is high during embryogenesis, it is not clear how ubiquitous CpG islands, such as that near the promoter of the Thy-1 gene, remain hypomethylated. Szyf et al. have assumed that methylation activity in embryonic stem (ES) cells is representative of that in cells in the early embryo. They find that, when the CpG island in the promoter region of the Thy-1 gene is transfected into such cells, not only does it remain hypomethylated, but so does more than 1 kb of surrounding sequence. The region that conferred this protection was narrowed down to a 214 bp segment of the Thy-1 promoter. Szyf et al. suggest that a nucMar protein binding to the protecting sequence - which may be a member of a class of such sequences - functions to prevent methyla~ion of ubiquitous CpG islands during early embryogenesis. Once methylation has been established, the pattern of methylation would be propagated during development by maintenance methylase activity, which methylates the daughter strand of newly replicated hemimethylated DNA in the same positions as the parent DNA strand, ah
close to the time when DNA replication begins. Two possibilities are that centrosome division may be regulated by cyclins, or by the synthesis of centrosome proteins. Centrosome reproduction has been studied in the presence of protein inhibitors in fertilized sea-urchin eggs by Sluder et aL, and in Xenopus blastulae by Gard et al. In sea-urchins, the centrosome used for the f'wst mitotic division comes ,from the fertilizing sperm and replicates at the time of pronuclear fusion. Continuous treatment with a combination of drugs that completely blocks protein synthesis allows the DNA synthesis accompanying pronuclear fusion to proceed before arrest of the cell cycle, and Sluder et
ai. f'md that under these conditions there is at least one, and sometimes several, doublings of the centrosomes, almost all of which contain a normal pair of centrioles. In agreement with these results, Gard et al. find that in Xenopus blastomeres treated for extended periods with cycloheximide, the cell cycle is blocked but multiple asters are observed, each containing functional centromeres. Thus centrosome replication can proceed during the arrest of both the cell cycle and protein synthesis, presumably using centrosome components pre-existi~ig in the cell. The mechanism that normally synchronizes centrosome replication with the cell cycle remains unknown, ah
~c SEV'r~B~R 1990 VOL.6 NO. 9
A DNA signal from the Thy-I gene defines de novo methylation patterns in embryonic stem cells M. SZYF, G. TANIGAWAAND P. MCCARTHYJR
Mol. Cell. Biol. 10, 4396--4400
A 'digest' of some recent papers of interest in the primary journals.
Deletions and a translocation interrupt a cloned gene at the Neurofi!iromatosis Type I locus D. VISKOCHIL ETAL.
Cell 62, 187-192
A major segment of the Neurofibgomatosis ~pe I gene: cDNA sequence, genomic structure, and point mutations k.M. CAWTHON ETAL.
Cell 62, 193-201
.Type 1 Neuroflbromatosis gene: identification of a large transcript disrupted in three NF1 patients M.R. WALLACEETAL.
Science 249, 181-186
Two groups have reported finding a gene that is a strong candidate for the neurofibromatosis type 1 (NF1) locus. NF1 is an autosomal dominant disorder that mainly affects neural crest-derived tissues. Within the last few years, two NFl-associated chromosomal translocations [t(l:17) and t(17:22)] have been found that placed the locus within a 600 kb region on chromosome 17q11.2, and three genes (EVI2, OMGP and RC1) have been identified between the translocation breakpoints, but NFl-specific mutations have not been detected in any of these genes. By adjusting the conditions of pulsed-fi¢.ld gel electrophoresis so as to improve resolution of small DNA fragments, Viskochil et aL were able to identify three NFl-associated deletions, one of which removed Olfly --11 i,:b of DNA in the t(17:22) breakpoint region. Once again, the strategy of using sequence conservation as an indication of the presence of a gene has proved successful: a probe from this region identified a sequence
Protein synthesis and the cell cycle: centrosome reproductionin set, urchin eggs is not under translationalcontrol G. SLUDER, F.J. MILLER, R. COLE AND C.L. RIEDER
J. Cell Biol. 110, 202.r~--2032
Centrosome duplication continues in cyclohex~!de.treated Xesopus
blastulae in the absence of a detectable cell cycle D.L. GARD, S. HAFEZI, T. ZHANG AND S.J. DOXSEY
J. Cell BioL 110, 2033-2042
The centrosome is a key component of the mitotic apparatus, but information has been scant about what controls its replication before mitosis,
from a murine cDNA library, and this cDNA in turn picked up sequences in several human cDNA libraries. Independently, Wallace et al. used the same sequence-conservation strategy, but obtained their probe to the translocation breakpoint region by chromosome jumping; another probe to screen human cDNA libraries was provided by a yeast artificial chromosome (YAC) clone from a YAC contig spanning the region. Both groups arrive at the same conclusion: that their cDNAs come from a large gene spanning both translocation breakpoints, and including within one of its introns all three of the previous candidate genes. The gene is expressed in a wide range of tissue types. Sequencing of the exons represented by the cDNAs obtained so far reveals no homology to any other known gene. Crucially, both groups have identified, in NF1 patients, point mutations in putative exons of the gene, strengthening the assertion that it is indeed the NF1 gene. Wallace et al. in Science and Cawthon et ai. in Cell point out that the dominant inheritance of NFI needs to be reconciled with the fact that each of the mutations detected so far would be expected to inactivate the NF1 gene. Perhaps, as is the case for retinoblastoma, tumours only develop when the normal NF1 allele is also inactivated by somatic mutation. The large size and complex organization of the NF1 region also raise intriguing questions about its regulation, with the possibility that some of the symptoms of NFI may be due to effects of mutations in the NF1 gene on expression of one or more of the oppositely transcribed EVI2, OMGPand RC1 genes, ah
CpG islands are clusters of CpG dinucleotides that are undermethylated, compared with the high methylation status of most CpG sequences in mammalian genomes. CpG islands can be either tissue specific, when they reflect the presence of active genes, or ubiquitous. Since de novo methylation activity is high during embryogenesis, it is not clear how ubiquitous CpG islands, such as that near the promoter of the Thy-1 gene, remain hypomethylated. Szyf et al. have assumed that methylation activity in embryonic stem (ES) cells is representative of that in cells in the early embryo. They find that, when the CpG island in the promoter region of the Thy-1 gene is transfected into such cells, not only does it remain hypomethylated, but so does more than 1 kb of surrounding sequence. The region that conferred this protection was narrowed down to a 214 bp segment of the Thy-1 promoter. Szyf et al. suggest that a nucMar protein binding to the protecting sequence - which may be a member of a class of such sequences - functions to prevent methyla~ion of ubiquitous CpG islands during early embryogenesis. Once methylation has been established, the pattern of methylation would be propagated during development by maintenance methylase activity, which methylates the daughter strand of newly replicated hemimethylated DNA in the same positions as the parent DNA strand, ah
close to the time when DNA replication begins. Two possibilities are that centrosome division may be regulated by cyclins, or by the synthesis of centrosome proteins. Centrosome reproduction has been studied in the presence of protein inhibitors in fertilized sea-urchin eggs by Sluder et aL, and in Xenopus blastulae by Gard et al. In sea-urchins, the centrosome used for the f'wst mitotic division comes ,from the fertilizing sperm and replicates at the time of pronuclear fusion. Continuous treatment with a combination of drugs that completely blocks protein synthesis allows the DNA synthesis accompanying pronuclear fusion to proceed before arrest of the cell cycle, and Sluder et
ai. f'md that under these conditions there is at least one, and sometimes several, doublings of the centrosomes, almost all of which contain a normal pair of centrioles. In agreement with these results, Gard et al. find that in Xenopus blastomeres treated for extended periods with cycloheximide, the cell cycle is blocked but multiple asters are observed, each containing functional centromeres. Thus centrosome replication can proceed during the arrest of both the cell cycle and protein synthesis, presumably using centrosome components pre-existi~ig in the cell. The mechanism that normally synchronizes centrosome replication with the cell cycle remains unknown, ah
~c SEV'r~B~R 1990 VOL.6 NO. 9
A DNA signal from the Thy-I gene defines de novo methylation patterns in embryonic stem cells M. SZYF, G. TANIGAWAAND P. MCCARTHYJR
Mol. Cell. Biol. 10, 4396--4400