Ubiquitylation of FACT by the cullin-E3 ligase Rtt101 connects FACT to DNA replication.
FACT plays important roles in both gene transcription and DNA replication. However, how this protein complex is targeted to these two distinct cellular processes remains largely unknown. Here we show that ubiquitylation of the Spt16 subunit of FACT by Rtt101, the cullin subunit of an E3 ubiquitin ligase in Saccharomyces cerevisiae, links FACT to DNA replication. We find Rtt101 interacts with and ubiquitylates Spt16 in vitro and in vivo. Deletion of RTT101 leads to reduced association of both FACT and the replicative helicase MCM with replication origins. Loss of Rtt101 also reduces binding of FACT to MCM, but not the association of FACT with Leo1 and Spt5, two proteins involved in transcription. Origin function is compromised in cells lacking Rtt101 or with an Spt16 mutation. These findings identify Spt16 as an Rtt101 substrate, and suggest that Spt16 ubiquitylation is important for FACT to function during DNA replication.
Han
Junhong
J
Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, Minnesota 55905, USA.
Li
Qing
Q
McCullough
Laura
L
Kettelkamp
Charisse
C
Formosa
Tim
T
Zhang
Zhiguo
Z
D044842:Cullin Proteins / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006609:High Mobility Group Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D035602:Transcriptional Elongation Factors / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D054875:Ubiquitination
D044842:Cullin Proteins; D004268:DNA-Binding Proteins; C541535:FACT protein, S cerevisiae; D006609:High Mobility Group Proteins; C476451:RTT101 protein, S cerevisiae; C071238:SPT16 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035602:Transcriptional Elongation Factors; D044767:Ubiquitin-Protein Ligases