Onset of the DNA replication checkpoint in the early Drosophila embryo.
The Drosophila embryo is a promising model for isolating gene products that coordinate S phase and mitosis. We have reported before that increasing maternal Cyclin B dosage to up to six copies (six cycB) increases Cdk1-Cyclin B (CycB) levels and activity in the embryo, delays nuclear migration at cycle 10, and produces abnormal nuclei at cycle 14. Here we show that the level of CycB in the embryo inversely correlates with the ability to lengthen interphase as the embryo transits from preblastoderm to blastoderm stages and defines the onset of a checkpoint that regulates mitosis when DNA replication is blocked with aphidicolin. A screen for modifiers of the six cycB phenotypes identified 10 new suppressor deficiencies. In addition, heterozygote dRPA2 (a DNA replication gene) mutants suppressed only the abnormal nuclear phenotype at cycle 14. Reduction of dRPA2 also restored interphase duration and checkpoint efficacy to control levels. We propose that lowered dRPA2 levels activate Grp/Chk1 to counteract excess Cdk1-CycB activity and restore interphase duration and the ability to block mitosis in response to aphidicolin. Our results suggest an antagonistic interaction between DNA replication checkpoint activation and Cdk1-CycB activity during the transition from preblastoderm to blastoderm cycles.
Crest
Justin
J
Department of Biology, University of Washington, Seattle, Washington 98195-1800, USA. jcrest@ucsc.edu
Oxnard
Nathan
N
Ji
Jun-Yuan
JY
Schubiger
Gerold
G
D000818:Animals; D001756:Blastoderm / Q000166:cytology; D002453:Cell Cycle; D000071877:Checkpoint Kinase 1; D002874:Chromosome Mapping; D002875:Chromosomes / Q000235:genetics; D019926:Cyclin B / Q000378:metabolism; D004261:DNA Replication; D029721:Drosophila Proteins / Q000378:metabolism; D004331:Drosophila melanogaster / Q000196:embryology* / Q000235:genetics*; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000378:metabolism*; D016153:Genes, Suppressor; D005820:Genetic Testing; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D018613:Microscopy, Confocal; D008857:Microscopy, Interference; D010641:Phenotype; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism
D019926:Cyclin B; D029721:Drosophila Proteins; D006657:Histones; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D011494:Protein Kinases; D000071877:Checkpoint Kinase 1; C000605877:grp protein, Drosophila