Inhibiting proteasome activity causes overreplication of DNA and blocks entry into mitosis in sea urchin embryos.
The proteasome has been shown to be involved in exit from mitosis by bringing about destruction of mitotic cyclins. Here, we present evidence that the proteasome is also required for proper completion of S phase and for entry into mitosis in the sea urchin embryonic cleavage cycle. A series of structurally related peptide-aldehydes prevent nuclear envelope breakdown in their order of inhibitory efficacies against the proteasome. Their efficacies in blocking exit from S phase and exit from mitosis correlate well, indicating that the proteasome is involved at both these steps. Mitotic histone HI kinase activation and tyrosine dephosphorylation of p34(cdc2) kinase are blocked by inhibition of the proteasome, indicating that the proteasome plays an important role in the pathway that leads to embryonic p34(cdc2 )kinase activation. Arrested embryos continued to incorporate [(3)H]thymidine and characteristically developed large nuclei. Pre-mitotic arrest can be overcome by treatment with caffeine, a manoeuvre that is known to override the DNA replication checkpoint. These data demonstrate that the proteasome is involved in the control of termination of S phase and consequently in the initiation of M phase of the first embryonic cell cycle.
Kawahara
H
H
Department of Physiological Sciences, Medical School, University of Newcastle, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK.
Philipova
R
R
Yokosawa
H
H
Patel
R
R
Tanaka
K
K
Whitaker
M
M
D000818:Animals; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism; D002110:Caffeine / Q000494:pharmacology; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002467:Cell Nucleus / Q000502:physiology; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects / Q000502:physiology*; D004625:Embryo, Nonmammalian / Q000502:physiology; D004640:Emetine / Q000494:pharmacology; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D007976:Leupeptins / Q000494:pharmacology; D008938:Mitosis / Q000187:drug effects / Q000235:genetics*; D009097:Multienzyme Complexes / Q000378:metabolism*; D009685:Nuclear Envelope / Q000502:physiology; D010641:Phenotype; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D016196:S Phase / Q000187:drug effects / Q000235:genetics; D012617:Sea Urchins / Q000502:physiology*; D013936:Thymidine / Q000493:pharmacokinetics; D014316:Tritium
D007976:Leupeptins; D009097:Multienzyme Complexes; D010726:Phosphodiesterase Inhibitors; D011480:Protease Inhibitors; D011500:Protein Synthesis Inhibitors; C089486:carbobenzoxy-leucyl-leucyl-norvalinal; D014316:Tritium; D019000:Phosphotyrosine; D002110:Caffeine; D011494:Protein Kinases; C102333:histone H1 kinase; D016203:CDC2 Protein Kinase; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D013936:Thymidine; D004640:Emetine