CUL1

PF3D7_0811000 (CUL1) is the cullin scaffold subunit of a Plasmodium SCF (Skp1–Cullin–F-box) E3 ubiquitin ligase, and in P. falciparum it assembles a functional SCF complex (PfSCF) comprising CUL1, PfRbx1, PfSkp1, PfFBXO1, and calcyclin binding protein that is competent for ubiquitination in vitro (PMID:38416790). The protein is expressed predominantly in erythrocytic trophozoite and schizont stages and shows nucleocytoplasmic localization with chromatin association (PMID:38416790). CUL1 activity is regulated by neddylation: it is a physiological substrate of PfNEDD8, identified as a cullin neddylation target by IP–mass spectrometry in P. falciparum and corroborated by heterologous complementation in S. cerevisiae cullin (cdc53) (PMID:33214620). In P. berghei, the CUL1 ortholog assembles a conserved SKP1/CUL1/RBX1 complex with FBXO1 and FBXL2 adaptors, co-purifying with Nedd8 and polyubiquitin, and this SCF^FBXO1 complex is required for nuclear segregation during schizogony, centrosome partitioning during microgametogenesis, gamete egress, and integrity of the apical and inner membrane complexes (PMID:36898988). Beyond its role as an SCF scaffold and NEDD8 target, the direct substrates ubiquitinated through CUL1 in P. falciparum have not been characterized in the available corpus.

1. 2019 Low

   Before this work it was unknown whether any Plasmodium enzyme could reverse cullin neddylation; showing that a parasite deubiquitylase acts on a neddylated cullin established that the neddylation cycle relevant to CUL1 regulation is enzymatically tractable in this system.

   Evidence In vitro deneddylation assay with recombinant PfUCHL3 acting on a reconstituted heterologous human SCF complex (Cul1–Skp1–Rbx1), read out by anti-Cul1/anti-Nedd8 Western blot

   PMID:31658303

   Open questions at the time

   • The SCF substrate used was human Cul1, not P. falciparum CUL1, so activity on the native parasite protein was not tested
   • Single in vitro assay from a single lab
   • Does not establish that PfUCHL3 regulates CUL1 in vivo
2. 2020 Medium

   It was unknown which parasite proteins are neddylated; identifying CUL1 as a PfNEDD8 substrate placed CUL1 within a regulated ubiquitin-ligase cycle and connected the NEDD8 pathway to cullin function.

   Evidence IP–mass spectrometry of tagged PfNEDD8 from P. falciparum identifying cullins as substrates, plus heterologous complementation in S. cerevisiae rub1Δ/ubc12Δ mutants confirming NEDD8 targeting of cullin (cdc53)

   PMID:33214620

   Open questions at the time

   • The specific neddylation lysine on CUL1 was not mapped in this study
   • Functional consequence of CUL1 neddylation for SCF activity not directly measured
   • IP-MS cannot distinguish direct from indirect neddylation
3. 2023 High

   The composition and biological requirement of a CUL1-based SCF in Plasmodium was undefined; reciprocal AP-MS and knockouts in P. berghei defined a SKP1/CUL1/RBX1 complex with FBXO1/FBXL2 adaptors and tied it to specific cell-division and egress events.

   Evidence Affinity purification of HA-tagged RBX1, SKP1, FBXO1, FBXL2 from synchronised gametocytes with label-free MS (CUL1, Nedd8, polyubiquitin co-purifying), plus genetic disruption with phenotypic analysis and ubiquitinomic survey

   PMID:36898988

   Open questions at the time

   • Phenotypes were assigned to the complex/FBXO1; the individual contribution of CUL1 was not isolated by CUL1-specific knockout
   • Substrates ubiquitinated during nuclear segregation or IMC assembly not identified
   • Shown in P. berghei, not P. falciparum
4. 2024 High

   Whether P. falciparum CUL1 forms an intact, catalytically active SCF was unresolved; direct reconstitution and in-parasite characterization established a functional PfSCF and defined CUL1's expression and localization.

   Evidence Tagged-protein expression in P. falciparum, immunoprecipitation, in vitro protein–protein interaction and ubiquitination assays, and subcellular localization microscopy

   PMID:38416790

   Open questions at the time

   • Physiological substrates ubiquitinated by PfSCF not identified
   • Requirement of CUL1 for parasite viability/stage progression not tested by knockdown/knockout
   • Role of calcyclin binding protein within the complex not mechanistically defined

• The direct substrates targeted for ubiquitination by the CUL1-containing SCF in P. falciparum, and how substrate selection is controlled across the erythrocytic and sexual stages, remain unknown.
• No P. falciparum SCF substrate identified
• No structural model of the parasite CUL1–Rbx1–Skp1–FBXO1 assembly
• Consequence of blocking CUL1 neddylation for parasite growth untested