JunD regulates genes involved in antioxidant defence. We took advantage of the chronic oxidative stress resulting from junD deletion to examine the role of reactive oxygen species (ROS) in tumour development. In a model of mammary carcinogenesis, junD inactivation increased tumour incidence and revealed an associated reactive stroma. junD‐inactivation in the stroma was sufficient to shorten tumour‐free survival rate and enhance metastatic spread. ROS promoted conversion of fibroblasts into highly migrating myofibroblasts through accumulation of the hypoxia‐inducible factor (HIF)‐1α transcription factor and the CXCL12 chemokine. Accordingly, treatment with an antioxidant reduced the levels of HIF and CXCL12 and numerous myofibroblast features. CXCL12 accumulated in the stroma of HER2‐human breast adenocarcinomas. Moreover, HER2 tumours exhibited a high proportion of myofibroblasts, which was significantly correlated to nodal metastases. Interestingly, this subset of tumours exhibited a significant nuclear exclusion of JunD and revealed an associated oxido‐reduction signature, further demonstrating the relevance of our findings in human cancers. Collectively, our data uncover a new mechanism by which oxidative stress increases the migratory properties of stromal fibroblasts, which in turn potentiate tumour dissemination.

The triple-negative breast cancer (TN BC) subtype is the most aggressive form of invasive BC. Despite intensive efforts to improve BC treatments, patients with TN BC continue to exhibit poor survival, with half developing resistance to chemotherapy. Here we identify autophagy as a key mechanism in the progression and chemoresistance of a subset of TN tumors. We demonstrate that LC3B, a protein involved in autophagosome formation, is a reliable marker of poor prognosis in TN BC, validating this prognostic value at both the mRNA and protein levels in several independent cohorts. We also show that LC3B has no prognostic value for other BC subtypes (Luminal or HER2 BC), thus revealing a specific impact of autophagy on TN tumors. Autophagy is essential for the proliferative and invasive properties in 3D of TN BC cells characterized by high LC3B levels. Interestingly, the activity of the transcriptional co-activator YAP1 (Yes-associated protein 1) is regulated by the autophagy process and we identify YAP1 as a new actor in the autophagy-dependent proliferative and invasive properties of high-LC3B TN BC. Finally, inhibiting autophagy by silencing ATG5 or ATG7 significantly impaired high-LC3B TN tumor growth in vivo. Moreover, using a patient-derived TN tumor transplanted into mice, we show that an autophagy inhibitor, chloroquine, potentiates the effects of chemotherapeutic agents. Overall, our data identify LC3B as a new prognostic marker for TN BC and the inhibition of autophagy as a promising therapeutic strategy for TN BC patients.

Background: We and others have recently demonstrated that the human retrovirus HTLV-I was producing a spliced antisense transcript, which led to the synthesis of the HBZ protein. The objective of the present study was to demonstrate the existence of antisense transcription in HIV-1 and to provide a better characterization of the transcript and its regulation.Results: Initial experiments conducted by standard RT-PCR analysis in latently infected J1.1 cell line and pNL4.3-transfected 293T cells confirmed the existence of antisense transcription in HIV-1. A more adapted RT-PCR protocol with limited RT-PCR artefacts also led to a successful detection of antisense transcripts in several infected cell lines. RACE analyses demonstrated the existence of several transcription initiation sites mapping near the 5' border of the 3'LTR (in the antisense strand). Interestingly, a new polyA signal was identified on the antisense strand and harboured the polyA signal consensus sequence. Transfection experiments in 293T and Jurkat cells with an antisense luciferase-expressing NL4.3 proviral DNA showed luciferase reporter gene expression, which was further induced by various T-cell activators. In addition, the viral Tat protein was found to be a positive modulator of antisense transcription by transient and stable transfections of this proviral DNA construct. RT-PCR analyses in 293T cells stably transfected with a pNL4.3-derived construct further confirmed these results. Infection of 293T, Jurkat, SupT1, U937 and CEMT4 cells with pseudotyped virions produced from the antisense luciferase-expressing NL4.3 DNA clone led to the production of an AZT-sensitive luciferase signal, which was however less pronounced than the signal from NL4.3Luc-infected cells.Conclusion: These results demonstrate for the first time that antisense transcription exists in HIV-1 in the context of infection. Possible translation of the predicted antisense ORF in this transcript should thus be re-examined.

The CXCR4 receptor and its ligand CXCL12 (also named stromal cell-derived factor 1, SDF1) have a critical role in chemotaxis and homing, key steps in cancer metastasis. Although myofibroblasts expressing CXCL12 are associated with the presence of axillary metastases in HER2 breast cancers (BC), the therapeutic interest of targeting CXCR4/CXCL12 axis in the different BC subtypes remains unclear. Here, we investigate this question by testing antitumor activity of CXCR4 inhibitors in patient-derived xenografts (PDX), which faithfully reproduce human tumor properties. We observed that two CXCR4 inhibitors, AMD3100 and TN14003, efficiently impair tumor growth and metastasis dissemination in both Herceptin-sensitive and Herceptin-resistant HER2 BC. Conversely, blocking CXCR4/CXCL12 pathway in triple-negative (TN) BC does not reduce tumor growth, and can even increase metastatic spread. Moreover, although CXCR4 inhibitors significantly reduce myofibroblast content in all BC subtypes, they decrease angiogenesis only in HER2 BC. Thus, our findings suggest that targeting CXCR4 could provide some therapeutic interest for HER2 BC patients, whereas it has no impact or could even be detrimental for TN BC patients.

Kaposi's sarcoma-associated herpesvirus encodes numerous regulatory proteins capable of modulating viral and cellular gene expression and affecting host cell functions. K-bZIP, a leucine zipper-containing transcription factor encoded by ORFK8, is one such protein. During infection, transcription of the ORFK8 early gene is turned on by the immediate-early replication and transcription factor activator (RTA). One described function of the K-bZIP nuclear protein is to interact with and repress RTA-mediated transactivation of viral promoters, including that of the K8 gene. In the present work, we provide evidence that the expression of K-bZIP results in the activation of the ifn -β gene. Of interest, ifn -β gene activation by K-bZIP is independent of interferon (IFN)-responsive factor 3 (IRF-3) and nuclear factor κB (NF-κB) activation. Using a DNA binding affinity assay and electromobility shift assay, we report that K-bZIP binds efficiently to the PRDIII-I region of the beta IFN (IFN-β) promoter, and, in doing so, it prevents the attachment of activated IRF-3 but not that of NF-κB or ATF2/c-Jun to the IFN-β promoter sequence. As a consequence, ifn -β gene activation in response to IFN inducers such as Sendai virus infection or expression of retinoic acid-inducible gene I, mitochondrial antiviral signaling protein, or TANK-binding kinase 1 (TBK-1) is severely impaired (>90%) by the presence of K-bZIP. K-bZIP also prevents the activation of RANTES and CXCL11, whose promoters are also regulated by IRF-3. Lysine 158 (target for SUMO conjugation), threonine 111, and serine 167 (targets for phosphorylation) mutants of K-bZIP were equally effective as wild-type K-bZIP in mediating the repression of TBK-1-activated ifn -β gene expression. Lastly, the overexpression of CREB binding protein could not reverse the K-bZIP repression of TBK-1-activated ifn -β gene expression. In all, our results indicate that K-bZIP binds directly to the PRDIII-I region of the IFN-β promoter and, as a consequence, causes a low level of ifn -β gene transcription. In doing so, K-bZIP prevents IRF-3 from binding to the IFN-β promoter and precludes the formation of the enhanceosome, which is required for maximal ifn -β gene transcription. A new role for K-bZIP as a protein involved in immune evasion is therefore uncovered.