OBI1

OBI1 (RNF219/C13ORF7) is a RING-type E3 ubiquitin ligase that operates in two distinct cellular arenas: control of DNA replication origin firing and regulation of mRNA fate through the CCR4-NOT deadenylase complex (PMID:31160578, PMID:33104214). In S-phase, OBI1 associates with the ORC complex and catalyses multi-mono-ubiquitylation of chromatin-bound ORC3 and ORC5; this modification is required for efficient origin firing and CMG helicase assembly without affecting pre-replication complex establishment, and non-ubiquitylable ORC3/5 mutants impair firing, establishing them as functionally relevant substrates (PMID:31160578). Independently, OBI1 binds the CCR4-NOT complex through its C-terminus, which directly engages the CNOT1 DUF3819 domain (PMID:40598799); in this context it inhibits CCR4-NOT deadenylase activity while enhancing the complex's deadenylation-independent capacity to repress translation (PMID:35660762), and it ubiquitinates the CCR4-NOT subunit CNOT6L to drive its proteasomal degradation (PMID:40598799). The integrity of the RING finger is central to this regulation: wild-type RING suppresses liquid-liquid phase separation, whereas mutations in its C3HC4 scaffold drive condensate formation that encapsulates and further inhibits CCR4-NOT (PMID:40497348).

1. 2019 High

   Established OBI1's first molecular function by identifying it as an ORC-associated E3 ligase whose ubiquitylation of ORC3/5 is required for replication origin firing, defining how an unstudied RING protein contributes to the cell cycle.

   Evidence Pre-RC proteomic interactome, Co-IP, in vitro ubiquitylation, siRNA, CMG formation assay, and non-ubiquitylable mutant rescue in human cells

   PMID:31160578

   Open questions at the time

   • Ubiquitin chain topology and the readers that interpret ORC3/5 mono-ubiquitylation are not defined
   • Whether the same catalytic activity links to its mRNA-regulatory functions is unresolved
2. 2020 Medium

   Connected OBI1 ligase activity to oncogenic signaling by showing it degrades α-catenin to enable YAP1/β-catenin-driven epigenetic activation of LGALS3, extending its role beyond replication into transcriptional control in cancer.

   Evidence Overexpression/knockdown, ubiquitin ligase assay, promoter epigenetic analysis and Co-IP in hepatocellular carcinoma cells

   PMID:33643786

   Open questions at the time

   • Direct ubiquitylation of α-catenin not reconstituted in abstract
   • Single tumor-cell context
3. 2020 Medium

   Opened the second functional arena by demonstrating OBI1 physically associates with the CCR4-NOT deadenylase and influences mRNA fate during stem-cell differentiation, implicating it in post-transcriptional control.

   Evidence Co-IP, in vitro deadenylation assay, RNA-seq and ES cell differentiation phenotype after knockdown

   PMID:33104214

   Open questions at the time

   • Subunit contacts not mapped
   • Direct effect on deadenylation rate not yet resolved (later refined)
4. 2022 Medium

   Refined the mechanism of CCR4-NOT regulation by showing OBI1 inhibits deadenylase activity while enhancing deadenylation-independent translational repression, recasting it as a directional modulator of the complex rather than a passive partner.

   Evidence Co-purification, in vitro deadenylase assay and translation repression reporter assay

   PMID:35660762

   Open questions at the time

   • Molecular basis for the switch toward translational repression unknown
   • Physiological mRNA targets not identified
5. 2022 Low

   Tested whether OBI1 drives oncogenic phenotypes through NF-κB signaling, providing a pathway-level placement in nasopharyngeal carcinoma.

   Evidence Overexpression/knockdown, p65 translocation, NF-κB target analysis and pharmacological inhibition rescue

   PMID:36512252

   Open questions at the time

   • No direct biochemical binding or substrate linking OBI1 to NF-κB identified
   • Indirect pathway placement via reporter and inhibitor only
   • Single cell-line context
6. 2025 Medium

   Mapped the physical interface and a catalytic substrate within CCR4-NOT, showing the OBI1 C-terminus binds the CNOT1 DUF3819 domain and ubiquitinates CNOT6L for degradation, linking complex binding to substrate turnover and proliferation control.

   Evidence MS of immunoprecipitates, domain-mapping pull-down, in vitro ubiquitination and knockdown proliferation assay in HEK293T

   PMID:40598799

   Open questions at the time

   • In-cell ubiquitylation and chain type on CNOT6L not shown
   • Whether CNOT6L turnover accounts for the differentiation phenotype unclear
7. 2025 Medium

   Revealed a phase-separation mechanism whereby RING-finger integrity gates condensate formation, showing C3HC4 mutations drive LLPS that sequesters and inhibits CCR4-NOT while wild-type RING suppresses LLPS.

   Evidence RING mutagenesis, live-cell condensate imaging, in vitro deadenylation and domain deletion in cells

   PMID:40497348

   Open questions at the time

   • Physiological triggers of RING-dependent LLPS unknown
   • Relationship between LLPS regulation and ORC-substrate ubiquitylation undefined

• How OBI1's replication-origin function and its CCR4-NOT/mRNA-regulatory functions are coordinated within a single protein, and whether they share regulatory inputs, remains unresolved.
• No unifying model linking ORC ubiquitylation and post-transcriptional control
• Endogenous mRNA targets and physiological condensate regulation not defined