RFWD3 (Q6PCD5) review notes

Gene: RFWD3 / RNF201 ; Fanconi anemia complementation group W (FANCW). Human, 774 aa,
chromosome 16. Domains: degenerate RING-type zinc finger (aa 287-331), coiled-coil
(361-413), WD40 β-propeller (repeats ~495-628, crystallized in PDB 6CVZ residues 425-774).
EC 2.3.2.27 (RING-type E3 ligase). ATM/ATR phospho-substrate at Ser46 and Ser63.

Core biology (synthesis)

RFWD3 is a RING-finger E3 ubiquitin ligase. It is recruited via its C-terminal WD40 domain
to the RPA2 subunit of the RPA complex that coats ssDNA at stalled replication forks / sites
of DNA damage. Once localized, RFWD3 polyubiquitinates RPA (all three subunits) and RAD51,
promoting their VCP/p97-dependent removal/turnover from DNA damage sites so that homologous
recombination (HR) and interstrand crosslink (ICL) repair can progress. It also promotes
ubiquitination of proteins on ssDNA to drive PCNA ubiquitination and translesion synthesis
(TLS). Biallelic RFWD3 mutations cause Fanconi anemia group W (FANCW). A secondary, less
central function is stabilization of p53 in the late DNA-damage response via an RFWD3-MDM2
complex (G1/S checkpoint).

Key provenance

Catalytic activity / MF

• PMID:21504906 — first in-vivo characterization.
• PMID:26474068
• UniProt: EC=2.3.2.27, RING-type (degenerate). Catalytic Cys315 (C315A abolishes activity).

RPA / RAD51 ubiquitination, HR, fork restart

• PMID:26474068
• PMID:26474068
• PMID:28575658
• PMID:28575658
• PMID:28575658
• PMID:28575658

RPA recruitment via WD40 / ICL repair / FANCW

• PMID:28575657
• PMID:28575657
• PMID:28575657
• PMID:28575657
• PMID:28575657
• PMID:28691929
• PMID:28691929

Recruitment to stalled forks / RPA2 binding / checkpoint

• PMID:21504906
• PMID:21504906
• PMID:21504906
• [PMID:21558276 title] RFWD3 associates with RPA and facilitates RPA-mediated DNA damage response; up-regulated in S-G2; partially at PML bodies (UniProt subcell).

Translesion synthesis

• PMID:33321094
• PMID:33321094

p53 / MDM2 (secondary role)

• PMID:20173098
• PMID:20173098
• PMID:20173098

Annotation decisions summary

• Core MF = RING E3 ubiquitin ligase activity (GO:0061630); process = ICL repair (GO:0036297),
  HR (GO:0000724), protein ubiquitination (GO:0016567); locations = nucleus (GO:0005634),
  site of DNA damage (GO:0090734).
• protein binding (GO:0005515) IPI annotations: all MARK_AS_OVER_ANNOTATED (uninformative;
  the biologically meaningful partners RPA2, RAD51, MDM2, p53, UBE2N are captured by specific
  terms or the core function).
• p53-axis terms (p53 binding, MDM2/MDM4 binding, G1 checkpoint, response to IR) =
  KEEP_AS_NON_CORE (secondary, single-group evidence).
• cytoplasm / cytosol / PML body = KEEP_AS_NON_CORE (undamaged-cell pool, not the functional site).