ABSTRACT
Approximately 30% of human lung cancers acquire mutations in either Keap1 or Nfe2l2, resulting in the stabilization of Nrf2, the Nfe2l2 gene product, which controls oxidative homeostasis. Here, we show that heme triggers the degradation of Bach1, a pro-metastatic transcription factor, by promoting its interaction with the ubiquitin ligase Fbxo22. Nrf2 accumulation in lung cancers causes the stabilization of Bach1 by inducing Ho1, the enzyme catabolizing heme. In mouse models of lung cancers, loss of Keap1 or Fbxo22 induces metastasis in a Bach1-dependent manner. Pharmacological inhibition of Ho1 suppresses metastasis in a Fbxo22-dependent manner. Human metastatic lung cancer display high levels of Ho1 and Bach1. Bach1 transcriptional signature is associated with poor survival and metastasis in lung cancer patients. We propose that Nrf2 activates a metastatic program by inhibiting the heme- and Fbxo22-mediated degradation of Bach1, and that Ho1 inhibitors represent an effective therapeutic strategy to prevent lung cancer metastasis.
Subject(s)
Basic-Leucine Zipper Transcription Factors/metabolism , Lung Neoplasms/pathology , NF-E2-Related Factor 2/metabolism , Animals , Basic-Leucine Zipper Transcription Factors/antagonists & inhibitors , Basic-Leucine Zipper Transcription Factors/genetics , Cell Line, Tumor , Cell Movement , F-Box Proteins/antagonists & inhibitors , F-Box Proteins/genetics , F-Box Proteins/metabolism , Female , Heme Oxygenase-1/antagonists & inhibitors , Heme Oxygenase-1/genetics , Heme Oxygenase-1/metabolism , Humans , Kaplan-Meier Estimate , Kelch-Like ECH-Associated Protein 1/antagonists & inhibitors , Kelch-Like ECH-Associated Protein 1/genetics , Kelch-Like ECH-Associated Protein 1/metabolism , Lung Neoplasms/metabolism , Lung Neoplasms/mortality , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , NF-E2-Related Factor 2/genetics , Neoplasm Metastasis , RNA Interference , RNA, Small Interfering/metabolism , Receptors, Cytoplasmic and Nuclear/antagonists & inhibitors , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Cytoplasmic and Nuclear/metabolism , Transcriptional ActivationABSTRACT
Protein prenylation is believed to be catalyzed by three heterodimeric enzymes: FTase, GGTase1 and GGTase2. Here we report the identification of a previously unknown human prenyltransferase complex consisting of an orphan prenyltransferase α-subunit, PTAR1, and the catalytic ß-subunit of GGTase2, RabGGTB. This enzyme, which we named GGTase3, geranylgeranylates FBXL2 to allow its localization at cell membranes, where this ubiquitin ligase mediates the polyubiquitylation of membrane-anchored proteins. In cells, FBXL2 is specifically recognized by GGTase3 despite having a typical carboxy-terminal CaaX prenylation motif that is predicted to be recognized by GGTase1. Our crystal structure analysis of the full-length GGTase3-FBXL2-SKP1 complex reveals an extensive multivalent interface specifically formed between the leucine-rich repeat domain of FBXL2 and PTAR1, which unmasks the structural basis of the substrate-enzyme specificity. By uncovering a missing prenyltransferase and its unique mode of substrate recognition, our findings call for a revision of the 'prenylation code'.
Subject(s)
Alkyl and Aryl Transferases/metabolism , Dimethylallyltranstransferase/metabolism , F-Box Proteins/metabolism , Alkyl and Aryl Transferases/chemistry , Cell Line , Crystallography, X-Ray , Dimethylallyltranstransferase/chemistry , F-Box Proteins/chemistry , HeLa Cells , Humans , Models, Molecular , Polyubiquitin/metabolism , Protein Conformation , Protein Prenylation , Protein Subunits/chemistry , Protein Subunits/metabolismABSTRACT
The mammalian FBXL10-RNF68-RNF2 ubiquitin ligase complex (FRRUC) mono-ubiquitylates H2A at Lys119 to repress transcription in unstressed cells. We found that the FRRUC is rapidly and transiently recruited to sites of DNA damage in a PARP1- and TIMELESS-dependent manner to promote mono-ubiquitylation of H2A at Lys119, a local decrease of H2A levels, and an increase of H2A.Z incorporation. Both the FRRUC and H2A.Z promote transcriptional repression, double strand break signaling, and homologous recombination repair (HRR). All these events require both the presence and activity of the FRRUC. Moreover, the FRRUC and its activity are required for the proper recruitment of BMI1-RNF2 and MEL18-RNF2, two other ubiquitin ligases that mono-ubiquitylate Lys119 in H2A upon genotoxic stress. Notably, whereas H2A.Z is not required for H2A mono-ubiquitylation, impairment of the latter results in the inhibition of H2A.Z incorporation. We propose that the recruitment of the FRRUC represents an early and critical regulatory step in HRR.
