Protein expression of the gp78 E3 ligase predicts poor breast cancer outcome based on race

Women of African ancestry suffer higher rates of breast cancer mortality compared with all other groups in the United States. Though the precise reasons for these disparities remain unclear, many recent studies have implicated a role for differences in tumor biology. Using an epitope-validated antibody against the endoplasmic reticulum-associated E3 ligase, gp78, we show that elevated levels of gp78 in patient breast cancer cells predict poor survival. Moreover, high levels of gp78 are associated with poor outcomes in both ER+ and ER- tumors, and breast cancers expressing elevated amounts of gp78 protein are enriched in gene expression pathways that influence cell cycle, metabolism, receptor-mediated signaling, and cell stress response pathways. In multivariate analysis adjusted for subtype and grade, gp78 protein is an independent predictor of poor outcomes in women of African ancestry. Furthermore, gene expression signatures, derived from patients stratified by gp78 protein expression, are strong predictors of recurrence and pathological complete response in retrospective clinical trial data and share many common features with gene sets previously identified to be overrepresented in breast cancers based on race. These findings implicate a prominent role for gp78 in tumor progression and offer insights into our understanding of racial differences in breast cancer outcomes.Fil: Singhal, Sandeep K.. No especifíca;Fil: Byun, Jung S.. National Institutes of Health; Estados UnidosFil: Yan, Tingfen. National Institutes of Health; Estados UnidosFil: Yancey, Ryan. Columbia University; Estados UnidosFil: Caban, Ambar. Columbia University; Estados UnidosFil: Hernandez, Sara Gil. National Institutes of Health; Estados UnidosFil: Bufford, Sediqua. No especifíca;Fil: Hewitt, Stephen M.. No especifíca;Fil: Winfield, Joy. Columbia University; Estados UnidosFil: Pradhan, Jaya. Columbia University; Estados UnidosFil: Mustkov, Vesco. Columbia University; Estados UnidosFil: McDonald, Jasmine A.. No especifíca;Fil: Pérez Stable, Eliseo J.. National Institutes of Health; Estados UnidosFil: Nápoles, Anna María. National Institutes of Health; Estados UnidosFil: Vohra, Nasreen. No especifíca;Fil: de Siervi, Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Yates, Clayton. No especifíca;Fil: Davis, Melissa B.. No especifíca;Fil: Yang, Mei. No especifíca;Fil: Tsai, Yien Che. No especifíca;Fil: Weissman, Allan M.. No especifíca;Fil: Gardner, Kevin. Columbia University; Estados Unido

Ubiquitylation in ERAD: Reversing to Go Forward?

Proteins are co-translationally inserted into the endoplasmic reticulum (ER) where they undergo maturation. Homeostasis in the ER requires a highly sensitive and selective means of quality control. This occurs through ER-associated degradation (ERAD).This complex ubiquitin-proteasome–mediated process involves ubiquitin conjugating enzymes (E2) and ubiquitin ligases (E3),lumenal and cytosolic chaperones, and other proteins, including the AAA ATPase p97 (VCP; Cdc48 in yeast). Probing of processes involving proteasomal degradation has generally depended on proteasome inhibitors or knockdown of specific E2s or E3s. In this issue of PLoS Biology, Ernst et al. demonstrate the utility of expressing the catalytic domain of a viral deubiquitylating enzyme to probe the ubiquitin system. Convincing evidence is provided that deubiquitylation is integral to dislocation of ERAD substrates from the ER membrane. The implications of this work for understanding ERAD and the potential of expressing deubiquitylating enzyme domains for studying ubiquitin-mediated processes are discussed

Differences in the Tumor Microenvironment between African-American and European-American Breast Cancer Patients

Background: African-American breast cancer patients experience higher mortality rates than European-American patients despite having a lower incidence of the disease. We tested the hypothesis that intrinsic differences in the tumor biology may contribute to this cancer health disparity. Methods and Results: Using laser capture microdissection, we examined genome-wide mRNA expression specific to tumor epithelium and tumor stroma in 18 African-American and 17 European-American patients. Numerous genes were differentially expressed between these two patient groups and a two-gene signature in the tumor epithelium distinguished between them. To identify the biological processes in tumors that are different by race/ethnicity, Gene Ontology and disease association analyses were performed. Several biological processes were identified which may contribute to enhanced disease aggressiveness in African-American patients, including angiogenesis and chemotaxis. African-American tumors also contained a prominent interferon signature. The role of angiogenesis in the tumor biology of African-American

The Unfolded Protein Response, Degradation from the Endoplasmic Reticulum, and Cancer

The endoplasmic reticulum (ER) is an essential organelle involved in many cellular functions including protein folding and secretion, lipid biosynthesis, and calcium homeostasis. Proteins destined for the cell surface or for secretion are made in the ER, where they are folded and assembled into multi-subunit complexes. The ER plays a vital role in cellular protein quality control by extracting and degrading proteins that are not correctly folded or assembled into native complexes. This process, known as ER-associated degradation (ERAD), ensures that only properly folded and assembled proteins are transported to their final destinations. Besides its role in protein folding and transport in the secretory pathway, the ER regulates the biosynthesis of cholesterol and other membrane lipids. ERAD is an important means to ensure that levels of the responsible enzymes are appropriately maintained. The ER is also a major organelle for oxygen and nutrient sensing as cells adapt to their microenvironment. Stresses that disrupt ER function lead to accumulation of unfolded proteins in the ER, a condition known as ER stress. Cells adapt to ER stress by activating an integrated signal transduction pathway called the unfolded protein response (UPR). The UPR represents a survival response by the cells to restore ER homeostasis. If ER stress persists, cells activate mechanisms that result in cell death. Chronic ER stress is increasingly being recognized as a factor in many human diseases such as diabetes, neurodegenerative disorders, and cancer. In this review, we discuss the roles of the UPR and ERAD in cancer and suggest directions for future research

Models for ERAD.

<p>(a–c) Classical view of ERAD. (a) ERAD substrate (black) is recognized by ER chaperones and partially translocated through a protein conducting channel complex/retrotranslocon (brown). The substrate is conjugated with chains of ubiquitin by an ER-resident ubiquitin ligase (E3) and its cognate ubiquitin conjugating enzyme (E2) on the cytosolic face of the ER membrane. (b) The p97 complex, comprising a hexamer of the AAA ATPase p97 and accessory proteins such as Ufd1 and Npl4 (not depicted), associates with the retrotranslocation complex, recognizes the polyubiquitin chain and extracts the ubiquitylated substrate to the cytosol. (c) Polyubiquitin chains target the dislocated substrate to the 26S proteasome (magenta) for degradation, in some cases assisted by shuttle proteins (pink) that bind both to ubiquitin chains and the proteasome. (d–f) Model based on results in Ernst et al. <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001038#pbio.1001038-Ernst1" target="_blank">[33]</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001038#pbio.1001038-Ernst2" target="_blank">[36]</a>. The exact mechanism by which the p97 complex extracts the substrate is not well understood. These new findings suggest that (d) the p97 complex recognizes polyubiquitin chains on the substrate as it moves through the protein-conducting channel. (e) DUBs associated with the p97 complex (purple) or potentially free in the cytosol (green) trim off the polyubiquitin chain on the substrate, allowing it to be threaded into the narrow channel of the p97 complex. (f) The dislocated substrate is ubiquitylated a second time by either ER-resident or cytosolic E3s for targeting to the proteasome (depicted in [c]).</p

Roles of ubiquitylation in cellular regulation.

<p>Conjugation of ubiquitin onto protein substrates requires at least three enzymes. One of two ubiquitin activating enzymes (E1) activates ubiquitin through an ATP-dependent step, forming a thioester linkage between the active site cysteine of E1 and the C-terminal carboxylate of ubiquitin. E1 then transfers the ubiquitin to the active site cysteine of one of approximately 40 mammalian ubiquitin conjugating enzymes (E2). The ubiquitin can be transferred to the active site of a HECT domain ubiquitin ligase (E3), which binds the substrate and mediates the conjugation of ubiquitin. RING finger and related E3s function as allosteric activators of E2, promoting the transfer of ubiquitin directly from the E2 to the substrate. The combination of E2/E3 determines the length and type of polyubiquitin chains assembled on the substrate, which can lead to diverse cellular effects, some of which are noted. Ubiquitylation is best characterized as modifying primary amines (lysines and the N-termini of proteins) <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001038#pbio.1001038-Ciechanover1" target="_blank">[44]</a>. More recently, there has been evidence that other nucleophillic amino acids including serine, threonine, and cysteine can also be modified with polyubiquitin chains <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001038#pbio.1001038-Wang1" target="_blank">[45]</a>–<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001038#pbio.1001038-Ishikura1" target="_blank">[48]</a>. DUBs perform a number of cellular roles, including removing ubiquitin from specific substrates. There are at least five classes of DUBs, and many DUBs show strong preferences for specific polyubiquitin chain linkages.</p

Stress-Induced Phosphorylation and Proteasomal Degradation of Mitofusin 2 Facilitates Mitochondrial Fragmentation and Apoptosis

Mitochondria play central roles in integrating pro- and antiapoptotic stimuli, and JNK is well known to have roles in activating apoptotic pathways. We establish a critical link between stress-induced JNK activation, mitofusin 2, which is an essential component of the mitochondrial outer membrane fusion apparatus, and the ubiquitin-proteasome system (UPS). JNK phosphorylation of mitofusin 2 in response to cellular stress leads to recruitment of the ubiquitin ligase (E3) Huwe1/Mule/ARF-BP1/HectH9/E3Histone/Lasu1 to mitofusin 2, with the BH3 domain of Huwe1 implicated in this interaction. This results in ubiquitin-mediated proteasomal degradation of mitofusin 2, leading to mitochondrial fragmentation and enhanced apoptotic cell death. The stability of a nonphosphorylatable mitofusin 2 mutant is unaffected by stress and protective against apoptosis. Conversely, a mitofusin 2 phosphomimic is more rapidly degraded without cellular stress. These findings demonstrate how proximal signaling events can influence both mitochondrial dynamics and apoptosis through phosphorylation-stimulated degradation of the mitochondrial fusion machinery