Proteomic analysis unveils a FANCA-modulated neddylation pathway involved in CXCR5 membrane targeting and cell mobility

Abstract

The aim of this study was to identify novel substrates of the FANCcore complex, the inactivation of which leads to the genetic disorder Fanconi anemia, which is associated with bone marrow failure, developmental abnormalities and a predisposition to cancer. Eight FANC proteins participate in the nuclear FANCcore complex, which functions as an E3 ubiquitin-ligase that monoubiquitylates FANCD2 and FANCI in response to replicative stress. Here, we use mass spectrometry to compare proteins from FANCcore-complexdeficient cells to those of rescued control cells after treatment with hydroxyurea, an inducer of FANCD2 monoubiquitylation. FANCD2 and FANCI appear to be the only targets of the FANCcore complex. We identify other proteins that are post-translationally modified in a FANCA-or FANCC-dependent manner. The majority of these potential targets localize to the cell membrane. Finally, we demonstrate that (a) the chemokine receptor CXCR5 is neddylated; (b) FANCA but not FANCC appears to modulate CXCR5 neddylation through an unknown mechanism; (c) CXCR5 neddylation is involved in targeting the receptor to the cell membrane; and (d) CXCR5 neddylation stimulates cell migration and motility. Our work has uncovered a pathway involving FANCA in neddylation and cell motility.

FANCD2 functions as a critical factor downstream of MiTF to maintain the proliferation and survival of melanoma cells

“…Probably, both migration and invasiveness are independent from the DDR activity of the pathway, but, more realistically, linked to the recently reported FANCA loss-of-function-associated defect in CXCR5 neddylation, which decreases the membrane localisation of this receptor involved in cell motility, invasion and homing26. Clearly, our data indicate that the FANC proteins have a central role in melanoma development, favouring growth, motility, invasiveness and DNA damage resistance of melanoma cells.…”

Regulation of cancer-related pathways by protein NEDDylation and strategies for the use of NEDD8 inhibitors in the clinic

“…Further analysis showed that CXCR5 is NEDDylated but Printed in Great Britain not ubiquitinated on lysine 339. The role of CXCR5 NEDDylation is to promote the membrane localisation of the receptor and is required for cell motility, a process controlled by CXCR5 in B-lymphocytes upon its ligand binding (Renaudin et al 2014).…”

Ubiquitin and Fanconi Anemia

“…In DT40, cell line mutations of FANC complex proteins lead to inactivating of FANCC and USP1 without affecting FANCD2 ubiquitylation. These results suggesting FANC complex proteins may have a significant role in the DNA damage response and other cellular functions interpedently of FANCD2/FANCI ubiquitylation [13,14].…”

“…This process is crucial for recycling of the ID molecules while the ubiquitylated and nonubiquitylated forms are necessary for normal cellular function. USP1 seems to be enzyme involved in deubiquitylating procedure [13,14].…”

“…Therefore, when the cell stress processes occur and the cells are in S/G2 phase monoubiquitinated FANCD2 and FANCI identified in DNA repair area. These two monoubiquitinated proteins form a heterodimer complex [13]. The binding of FANCD2 with histone H2B reveals that ubiquitination is crucial of complex identification to chromatin and DNA repair process [26,27].…”