PF3D7_0619700

FBXO1 (PF3D7_0619700) is the substrate-recognition F-box subunit of a conserved SCF (SKP1–Cullin1–FBXO1) E3 ubiquitin ligase in Plasmodium that governs inner membrane complex (IMC) biogenesis and cell division across asexual and sexual life-cycle stages (PMID:36898988, PMID:38416790). In P. falciparum the protein assembles into a functional SCF complex (PfCullin-1, PfRbx1, PfSkp1, PfFBXO1, and calcyclin-binding protein) with demonstrated in vitro ubiquitination activity, with PfSKP1 as its stable interacting partner (PMID:38416790, PMID:39915671). FBXO1 localizes to the apical region of dividing merozoites and transitions to an IMC-like distribution during segmentation, and occupies a unilateral IMC region in gametocytes, tying its position to IMC assembly (PMID:39915671). Inducible depletion in P. falciparum causes abnormal segmentation and karyokinesis yielding inviable merozoites, and malformed gametocytes that fail to mature (PMID:39915671); in P. berghei, loss of the SCF^FBXO1 complex disrupts nuclear segregation during schizogony, centrosome partitioning during microgametogenesis, gamete egress, and apical/IMC integrity (PMID:36898988). Its ubiquitination activity intersects with CDPK1-mediated phosphorylation signaling (PMID:36898988). The identity of FBXO1's ubiquitination substrates and the biochemical basis of the FBXO1–CDPK1 link have not been resolved in the available corpus.

1. 2020 Low

   Before mechanism could be probed, a tool was needed to deplete FBXO1 in the stages where it acts; tagging the fbxo1 product with an auxin-inducible degron established that the protein is expressed in schizonts and gametocytes and can be conditionally degraded.

   Evidence CRISPR/Cas9 AID tagging with auxin-induced degradation confirmed by western blot in P. yoelii

   PMID:33307135

   Open questions at the time

   • No downstream FBXO1-specific phenotype reported
   • Does not establish complex membership or enzymatic activity
   • Single-method demonstration
2. 2023 High

   It was unknown whether FBXO1 functions as an E3 ligase in vivo and what cellular processes it controls; genetic depletion combined with ubiquitinomics showed it forms a functional SCF^FBXO1 complex essential for nuclear segregation, centrosome partitioning, gamete egress, and apical/IMC integrity.

   Evidence Conditional knockdown, ubiquitinomic mass spectrometry, and stage-resolved immunofluorescence in P. berghei

   PMID:36898988

   Open questions at the time

   • Specific ubiquitination substrates not defined
   • No structural model of the complex
   • Mechanism linking ubiquitination to IMC integrity unresolved
3. 2023 Medium

   How FBXO1 activity integrates with other regulatory signals was unknown; comparing ubiquitinome and phosphoproteome upon FBXO1 depletion revealed an interplay between SCF^FBXO1 ubiquitination and CDPK1-mediated phosphorylation.

   Evidence Ubiquitinomic profiling in FBXO1-depleted parasites combined with phosphoproteomics in P. berghei

   PMID:36898988

   Open questions at the time

   • No direct biochemical reconstitution of the FBXO1–CDPK1 link
   • Directionality of the ubiquitination/phosphorylation crosstalk unclear
   • Shared substrates not defined
4. 2024 High

   Whether the human-pathogen species retains the same complex and enzymatic activity was untested; reconstitution and interaction assays confirmed PfFBXO1 is part of a functional PfSCF complex with in vitro ubiquitination activity.

   Evidence Co-immunoprecipitation, in vitro protein–protein interaction assay, and in vitro ubiquitination assay in P. falciparum

   PMID:38416790

   Open questions at the time

   • Physiological substrates of the ligase not identified
   • Role of calcyclin-binding protein in substrate selection unknown
5. 2025 High

   The subcellular basis of FBXO1 function was unresolved; imaging showed it localizes apically in dividing merozoites and transitions to an IMC-like distribution and a unilateral IMC region in gametocytes, and inducible knockout produced abnormal segmentation/karyokinesis and immature gametocytes, tying FBXO1 to IMC-dependent division in both asexual and sexual stages.

   Evidence Epitope-tagged live/immunofluorescence imaging, inducible knockout with morphological/viability assays and Co-IP/MS in P. falciparum

   PMID:39915671

   Open questions at the time

   • Mechanism recruiting FBXO1 to the apical/IMC scaffold unknown
   • Co-purifying IMC proteins not validated as ubiquitination substrates
   • PfSKP1 interaction not reciprocally validated in this study

• The direct ubiquitination substrates of FBXO1 at the IMC scaffold and the biochemical mechanism coupling its ligase activity to CDPK1 phosphorylation signaling remain to be defined.
• No validated in vivo ubiquitination substrate
• No structural model of substrate recognition
• Mechanistic basis of ubiquitination/phosphorylation crosstalk unresolved