Replicative stress induces intragenic transcription of the ASE1 gene that negatively regulates Ase1 activity.
Intragenic transcripts initiate within the coding region of a gene, thereby producing shorter mRNAs and proteins. Although intragenic transcripts are widely expressed [1], their role in the functional regulation of genes remains largely unknown. In budding yeast, DNA replication stress activates the S phase checkpoint that stabilizes replication forks and arrests cells in S phase with a short spindle [2-4]. When yeast cells were treated with hydroxyurea (HU) to block DNA synthesis and induce replication stress, we found that Ase1, a conserved spindle midzone protein [5], appeared as two short protein isoforms in addition to the full-length protein. We further demonstrated that the short isoforms result from intragenic transcription of ASE1, which depends on the S phase checkpoint. Blocking generation of the short isoforms leads to a destabilized S phase spindle, characterized by increased spindle dynamics and frequent spindle collapse. Because the short Ase1 isoforms localize at the spindle in HU-treated cells and overexpression of the short Ase1 isoforms impairs the spindle midzone localization of full-length Ase1, it is likely that the presence of short Ase1 isoforms stabilizes the spindle by antagonizing full-length Ase1. Together, our results reveal intragenic transcription as a unique mechanism to downregulate gene functions in response to DNA replication stress.
Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL 32306, USA.
Kelly
K
McKnight
Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL 32306, USA.
Hong
H
Liu
Department of Biomedical Sciences, College of Medicine, Florida State University, 1115 West Call Street, Tallahassee, FL 32306, USA. Electronic address: yanchang.wang@med.fsu.edu.
Yanchang
Y
Wang
D004261:DNA Replication / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects*; D006918:Hydroxyurea / Q000494:pharmacology; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
C472767:Ase1 protein, S cerevisiae; D004791:Enzyme Inhibitors; D008869:Microtubule-Associated Proteins; D020033:Protein Isoforms; D029701:Saccharomyces cerevisiae Proteins; D006918:Hydroxyurea