Structural and biochemical basis of interdependent FANCI‐FANCD2 ubiquitination
Kimon Lemonidis, Martin L. Rennie, Connor Arkinson, Viduth K. Chaugule, Mairi Clarke, James Streetley, Helen Walden
Abstract
Abstract Di‐monoubiquitination of the FANCI‐FANCD2 (ID2) complex is a central and crucial step for the repair of DNA interstrand crosslinks via the Fanconi anaemia pathway. While FANCD2 ubiquitination precedes FANCI ubiquitination, FANCD2 is also deubiquitinated at a faster rate than FANCI, which can result in a FANCI‐ubiquitinated ID2 complex (I Ub D2). Here, we present a 4.1 Å cryo‐EM structure of I Ub D2 complex bound to double‐stranded DNA. We show that this complex, like ID2 Ub and I Ub D2 Ub , is also in the closed ID2 conformation and clamps on DNA. The target lysine of FANCD2 (K561) becomes fully exposed in the I Ub D2‐DNA structure and is thus primed for ubiquitination. Similarly, FANCI's target lysine (K523) is also primed for ubiquitination in the ID2 Ub ‐DNA complex. The I Ub D2‐DNA complex exhibits deubiquitination resistance, conferred by the presence of DNA and FANCD2. ID2 Ub ‐DNA, on the other hand, can be efficiently deubiquitinated by USP1‐UAF1, unless further ubiquitination on FANCI occurs. Therefore, FANCI ubiquitination effectively maintains FANCD2 ubiquitination in two ways: it prevents excessive FANCD2 deubiquitination within an I Ub D2 Ub ‐DNA complex, and it enables re‐ubiquitination of FANCD2 within a transient, closed‐on‐DNA, I Ub D2 complex.