The UBXN1 Protein Associates with Autoubiquitinated Forms of the BRCA1 Tumor Suppressor and Inhibits Its Enzymatic Function

Abstract

Although the BRCA1 tumor suppressor has been implicated in many cellular processes, the biochemical mechanisms by which it influences these diverse pathways are poorly understood. The only known enzymatic function of BRCA1 is the E3 ubiquitin ligase activity mediated by its highly conserved RING domain. In vivo, BRCA1 associates with the BARD1 polypeptide to form a heterodimeric BRCA1/BARD1 complex that catalyzes autoubiquitination of BRCA1 and trans ubiquitination of other protein substrates. In most cases, BRCA1-dependent ubiquitination generates polyubiquitin chains bearing an unconventional K6 linkage that does not appear to target proteins for proteasomal degradation. Since ubiquitin-dependent processes are usually mediated by cellular receptors with ubiquitin-binding motifs, we screened for proteins that specifically bind autoubiquitinated BRCA1. Here we report that the UBXN1 polypeptide, which contains a ubiquitin-associated (UBA) motif, recognizes autoubiquitinated BRCA1. This occurs through a bipartite interaction in which the UBA domain of UBXN1 binds K6-linked polyubiquitin chains conjugated to BRCA1 while the C-terminal sequences of UBXN1 bind the BRCA1/BARD1 heterodimer in a ubiquitin-independent fashion. Significantly, the E3 ligase activity of BRCA1/BARD1 is dramatically reduced in the presence of UBXN1, suggesting that UBXN1 regulates the enzymatic function of BRCA1 in a manner that is dependent on its ubiquitination status.Germ line mutations of the BRCA1 tumor suppressor are responsible for many cases of hereditary breast and ovarian cancer (49). However, the mechanisms by which these mutations promote tumorigenesis have been difficult to ascertain, in part because BRCA1 has been implicated in a broad spectrum of cellular processes that includes-but is not limited to-cell cycle checkpoint control, DNA repair, mitotic spindle assembly, centrosome function, and transcriptional regulation (33,34,38,51). The major isoform of BRCA1 is a large protein that harbors an N-terminal RING domain and two tandem C-terminal BRCT motifs (32). In vivo, BRCA1 associates with the structurally similar BARD1 protein to form a heterodimer that has E3 ubiquitin ligase activity (16,50). Mice bearing null mutations of either Brca1 or Bard1 undergo embryonic lethality in indistinguishable manners, suggesting that the early developmental functions of both proteins are determined by the Brca1/Bard1 heterodimer (30). In addition, conditional inactivation of either gene in mouse mammary epithelial cells induces breast carcinomas that are phenotypically interchangeable and closely resemble the human breast tumors of BRCA1 mutation carriers (44). Thus, the BRCA1/BARD1 heterodimer is likely to mediate many aspects of BRCA1 function, including its critical role as a tumor suppressor in breast and ovarian tissues (1, 44).The E3 ubiquitin ligase activity of BRCA1/BARD1 is the only known enzymatic function of BRCA1 (4,16,29,54). E3 ligases catalyze the formation of an isopeptide linkage between the C-terminal carboxyl group of ubi...

Ubiquitin-associated Domain-containing Ubiquitin Regulatory X (UBX) Protein UBXN1 Is a Negative Regulator of Nuclear Factor κB (NF-κB) Signaling

“…Knockdown of VCP/p97 in the reporter assay indicated that VCP/p97 is simply not involved in UBXN1-mediated NF-B inhibition. Another study (24) revealed that UBXN1 may block the function of BRCA1 by directly binding the ubiquitin chain. In our data, UBXN1 interacts with cIAPs through the UBA domain and prevents the interaction between TRAF2 and cIAPs as the negative regulator of NF-B signaling pathway.…”

UBXD Proteins: A Family of Proteins with Diverse Functions in Cancer

“…BRCA1 participates in tumor suppression and its ubiquitin ligase activity has a significant role in this tumor suppression [127]. Therefore, UBXN1 may facilitate tumor progression by negative regulation of BRCA1 [128]. UBXN1-dependent inhibition of pro-survival protein cIAP1 and tumor suppressor protein BRCA1 suggests a controversial role for UBXN1 cancer.…”

Linkage via K27 Bestows Ubiquitin Chains with Unique Properties among Polyubiquitins

“…Ubiquitination with K6-linked chains occurs on the BRCA1-BARD1 Ub ligase and its associated substrates, suggesting a link to DNA repair processes (Wu-Baer et al, 2010). K11-polyUb may interact with the Npl4 adaptor protein during Drosophila development, among other roles associated with the mitotic phase in the cell cycle and ERAD (Meyer and Rape, 2014; Zhang et al, 2013).…”