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no poles encodes a predicted E3 ubiquitin ligase required for early embryonic development of Drosophila.

Merkle JA, Rickmyre JL, Garg A, Loggins EB, Jodoin JN, Lee E, Wu LP, Lee LA
Development. 2009 136 (3): 449-59

PMID: 19141674 · PMCID: PMC2687590 · DOI:10.1242/dev.027599

In a screen for cell-cycle regulators, we identified a Drosophila maternal effect-lethal mutant that we named ;no poles' (nopo). Embryos from nopo females undergo mitotic arrest with barrel-shaped, acentrosomal spindles during the rapid S-M cycles of syncytial embryogenesis. We identified CG5140, which encodes a candidate RING domain-containing E3 ubiquitin ligase, as the nopo gene. A conserved residue in the RING domain is altered in our EMS-mutagenized allele of nopo, suggesting that E3 ligase activity is crucial for NOPO function. We show that mutation of a DNA checkpoint kinase, CHK2, suppresses the spindle and developmental defects of nopo-derived embryos, revealing that activation of a DNA checkpoint operational in early embryos contributes significantly to the nopo phenotype. CHK2-mediated mitotic arrest has been previously shown to occur in response to mitotic entry with DNA damage or incompletely replicated DNA. Syncytial embryos lacking NOPO exhibit a shorter interphase during cycle 11, suggesting that they may enter mitosis prior to the completion of DNA replication. We show that Bendless (BEN), an E2 ubiquitin-conjugating enzyme, interacts with NOPO in a yeast two-hybrid assay; furthermore, ben-derived embryos arrest with a nopo-like phenotype during syncytial divisions. These data support our model that an E2-E3 ubiquitination complex consisting of BEN-UEV1A (E2 heterodimer) and NOPO (E3 ligase) is required for the preservation of genomic integrity during early embryogenesis.

MeSH Terms (19)

Amino Acid Sequence Animals Checkpoint Kinase 2 DNA Damage Drosophila Drosophila Proteins Embryo, Nonmammalian Female HeLa Cells Humans Mitosis Molecular Sequence Data Mutation Protein-Serine-Threonine Kinases Protein Binding Spindle Apparatus Two-Hybrid System Techniques Ubiquitin-Conjugating Enzymes Ubiquitin-Protein Ligases

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