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Summary: Although DELLA protein destabilization mediated by post‐translational modifications is essential for gibberellin (GA) signal transduction and GA‐regulated anthocyanin biosynthesis, the related mechanisms remain largely unknown. In this study, we report the ubiquitination and phosphorylation of an apple DELLA protein MdRGL2a in response to GA signaling and its regulatory role in anthocyanin biosynthesis.MdRGL2a could interact with MdWRKY75 to enhance the MdWRKY75‐activated transcription of anthocyanin activator MdMYB1 and interfere with the interaction between anthocyanin repressor MdMYB308 and MdbHLH3 or MdbHLH33, thereby promoting anthocyanin accumulation.A protein kinase MdCIPK20 was found to phosphorylate and protect MdRGL2a from degradation, and it was essential for MdRGL2a‐promoting anthocyanin accumulation. However, MdRGL2a and MdCIPK20 were ubiquitinated and degraded by E3 ubiquitin ligases MdSINA1 and MdSINA2, respectively, both of which were activated in the presence of GA.Our results display the integration of SINA1/2 with CIPK20 to dynamically regulate GA signaling and will be helpful toward understanding the mechanism of GA signal transduction and GA‐inhibited anthocyanin biosynthesis. The discovery of extensive interactions between DELLA and SINA and CIPK proteins in apple will provide reference for the study of ubiquitination and phosphorylation of DELLA proteins in other species. [ABSTRACT FROM AUTHOR]

PERfecting breast cancer treatment: Basal-like breast cancers are associated with a poorer prognosis, and there are no targeted therapies. Li et al. examined Faciogenital Dysplasia 5 (FGD5) in basal-like breast cancer, finding that FGD5 supported tumor initiation by reducing epidermal growth factor receptor (EGFR) ubiquitination and degradation, leading to increased invasiveness and self-renewal. The authors developed a peptide, PER3, to reduce the interaction between EGFR and FGD5. PER3 treatment suppressed growth of tumors displaying high EGFR and FGD5 abundance in mice and resensitized resistant tumors to paclitaxel. Although further studies are needed to determine the safety of PER3, FGD5 is a promising target for the treatment of basal-like breast cancers. Most basal-like breast cancers (BLBCs) are triple-negative breast cancers (TNBCs), which have the worst prognosis and distant metastasis-free survival among breast cancer subtypes. Now, no targeted therapies are available for patients with BLBC due to the lack of reliable and effective molecular targets. Here, we performed the BLBC tissue microarray–based immunohistochemical analysis and showed that Faciogenital Dysplasia 5 (FGD5) abundance is associated with poor prognosis in BLBCs. FGD5 deletion decreased the proliferation, invasion, and tumorsphere formation capacity of BLBC cells. Furthermore, genetic inhibition of Fgd5 in mouse mammary epithelial cells attenuated BLBC initiation and progression by reducing the self-renewal ability of tumor-initiating cells. In addition, FGD5 abundance was positively correlated with the abundance of epidermal growth factor receptor (EGFR) in BLBCs. FGD5 ablation decreased EGFR abundance by reducing EGFR stability in TNBC cells in 2D and 3D culture conditions. Mechanistically, FGD5 binds to EGFR and interferes with basal EGFR ubiquitination and degradation induced by the E3 ligase ITCH. Impaired EGFR degradation caused BLBC cell proliferation and promoted invasive properties and self-renewal. To verify the role of the FGD5-EGFR interaction in the regulation of EGFR stability, we screened a cell-penetrating α-helical peptide PER3 binding with FGD5 to disrupt the interaction. Treatment of BLBC patient-derived xenograft–bearing mice with the peptide PER3 disrupting the FGD5-EGFR interaction either with or without chemotherapy reduced BLBC progression. Our study identified FGD5 as a positive modulator of tumor-initiating cells and suggests a potential therapeutic option for the BLBC subtype of breast cancer. [ABSTRACT FROM AUTHOR]

Summary: The plant hormone jasmonic acid (JA) is involved in the cold stress response, and the inducer of CBF expression 1 (ICE1)‐ C‐repeat binding factor (CBF) regulatory cascade plays a key role in the regulation of cold stress tolerance. In this study, we showed that a novel B‐box (BBX) protein MdBBX37 positively regulates JA‐mediated cold‐stress resistance in apple.We found that MdBBX37 bound to the MdCBF1 and MdCBF4 promoters to activate their transcription, and also interacted with MdICE1 to enhance the transcriptional activity of MdICE1 on MdCBF1, thus promoting its cold tolerance.Two JA signaling repressors, MdJAZ1 and MdJAZ2 (JAZ, JAZMONATE ZIM‐DOMAIN), interacted with MdBBX37 to repress the transcriptional activity of MdBBX37 on MdCBF1 and MdCBF4, and also interfered with the interaction between MdBBX37 and MdICE1, thus negatively regulating JA‐mediated cold tolerance. E3 ligase MdMIEL1 (MIEL1, MYB30‐Interacting E3 Ligase1) reduced MdBBX37‐improved cold resistance by mediating ubiquitination and degradation of the MdBBX37 protein.The data reveal that MIEL1 and JAZ proteins co‐regulate JA‐mediated cold stress tolerance through the BBX37‐ICE1‐CBF module in apple. These results will aid further examination of the post‐translational modification of BBX proteins and the regulatory mechanism of JA‐mediated cold stress tolerance. [ABSTRACT FROM AUTHOR]

Jasmonic acid (JA) is shown to induce leaf senescence. However, the underlying molecular mechanism is not well understood, especially in woody plants such as fruit trees. In this study, we are interested in exploring the biological role of MdBT2 in JA‐mediated leaf senescence. We found that MdBT2 played an antagonistic role in MdMYC2‐promoted leaf senescence. Our results revealed that MdBT2 interacted with MdMYC2 and accelerated its ubiquitination degradation, thus negatively regulated MdMYC2‐promoted leaf senescence. In addition, MdBT2 acted as a stabilizing factor to improve the stability of MdJAZ2 through direct interaction, thereby inhibited JA‐mediated leaf senescence. Furthermore, our results also showed that MdBT2 interacted with a subset of JAZ proteins in apple, including MdJAZ1, MdJAZ3, MdJAZ4 and MdJAZ8. Our investigations provide new insight into molecular mechanisms of JA‐modulated leaf senescence. The dynamic JA‐MdBT2‐MdJAZ2‐MdMYC2 regulatory module plays an important role in JA‐modulated leaf senescence. [ABSTRACT FROM AUTHOR]

The transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in MycEμ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression. c-MYC is often deregulated in human cancers including lymphomas. Here, the authors show that a member of the pseudokinase family, tribbles homologue 3 (TRIB3), interacts with c-MYC to suppress c-MYC ubiquitination and degradation, leading to increased proliferation and self-renewal of lymphoma cells. [ABSTRACT FROM AUTHOR]

High expression or aberrant activation of epidermal growth factor receptor (EGFR) is related to tumor progression and therapy resistance across cancer types, including non-small cell lung cancer (NSCLC). EGFR tyrosine kinase inhibitors (TKIs) are first-line therapy for NSCLC. However, patients eventually deteriorate after inevitable acquisition of EGFR TKI-resistant mutations, highlighting the need for therapeutics with alternative mechanisms of action. Here, we report that the elevated tribbles pseudokinase 3 (TRIB3) is positively associated with EGFR stability and NSCLC progression. TRIB3 interacts with EGFR and recruits PKCα to induce a Thr654 phosphorylation and WWP1-induced Lys689 ubiquitination in the EGFR juxtamembrane region, which enhances EGFR recycling, stability, downstream activity, and NSCLC stemness. Disturbing the TRIB3-EGFR interaction with a stapled peptide attenuates NSCLC progression by accelerating EGFR degradation and sensitizes NSCLC cells to chemotherapeutic agents. These findings indicate that targeting EGFR degradation is a previously unappreciated therapeutic option in EGFR-related NSCLC. Depletion of tribbles pseudokinase 3 (TRIB3) is known to suppress the expression of several tumor-promoting factors, including EGFR. Here, the authors show that TRIB3 interacts with EGFR and regulates its stability and activity, and perturbing EGFR-TRIB3 interaction attenuates NSCLC progression by accelerating EGFR degradation. [ABSTRACT FROM AUTHOR]

Summary: MYB transcription factors (TFs) have been demonstrated to play diverse roles in plant growth and development through interaction with basic helix‐loop‐helix (bHLH) TFs. MdbHLH33, an apple bHLH TF, has been identified as a positive regulator in cold tolerance and anthocyanin accumulation by activating the expressions of MdCBF2 and MdDFR. In the present study, a MYB TF MdMYB308L was found to also positively regulate cold tolerance and anthocyanin accumulation in apple. We found that MdMYB308L interacted with MdbHLH33 and enhanced its binding to the promoters of MdCBF2 and MdDFR. In addition, an apple RING E3 ubiquitin ligase MYB30‐INTERACTING E3 LIGASE 1 (MdMIEL1) was identified to be an MdMYB308L‐interacting protein and promoted the ubiquitination degradation of MdMYB308L, thus negatively regulated cold tolerance and anthocyanin accumulation in apple. These results suggest that MdMYB308L acts as a positive regulator in cold tolerance and anthocyanin accumulation in apple by interacting with MdbHLH33 and undergoes MdMIEL1‐mediated protein degradation. The dynamic change in MYB‐bHLH protein complex seems to play a key role in the regulation of plant growth and development. [ABSTRACT FROM AUTHOR]

Summary: Wounding stress leads to anthocyanin accumulation. However, the underlying molecular mechanism remains elusive. In this study, MdWRKY40 was found to promote wounding‐induced anthocyanin biosynthesis in association with MdMYB1 and undergo MdBT2‐mediated degradation in apple.We found that MdMYB1, a positive regulator of anthocyanin biosynthesis, was essential for the wounding‐induced anthocyanin biosynthesis in apple. MdWRKY40 was identified as an MdMYB1‐interacting protein, and enhanced the binding of MdMYB1 to its target genes in response to wounding.We found that MdBT2 interacted physically with MdWRKY40 and was involved in its degradation through the 26S proteasome pathway.Our results demonstrate that MdWRKY40 is a key modulator in the wounding‐induced anthocyanin biosynthesis, which provides new insights into the regulation of wounding‐induced anthocyanin biosynthesis at both the transcriptional and post‐translational levels in apple. [ABSTRACT FROM AUTHOR]

Summary: The molecular mechanism of leaf senescence in apple (Malus domestica) is still not fully understood. We used gene expression analysis and protein–protein interactions to decipher the relationships of abscisic acid (ABA) and two proteins, MdbHLH93 and MdBT2, in the senescence process.We found that MdbHLH93 promoted leaf senescence and the expression of senescence‐related genes, which exhibited similar effects to ABA on leaf senescence. MdbHLH93 activated directly the transcription of MdSAG18.We also found that an ABA‐responsive protein, MdBT2, interacted directly with MdbHLH93, and induced the ubiquitination and degradation of the MdbHLH93 protein, and thus delayed leaf senescence.Our findings provide new insights into the regulatory network of leaf senescence through the functional interactions among ABA, MdbHLH93 and MdBT2. [ABSTRACT FROM AUTHOR]

Aims: Pulmonary fibrosis (PF) is characterized by myofibroblast activation through oxidative stress. However, the precise regulation of myofibroblast transdifferentiation remains largely uncharacterized. Results: In this study, we found that tanshinone IIA (Tan-IIA), an active component in the root of Salvia miltiorrhiza Bunge, can suppress reactive oxygen species (ROS)-mediated activation of myofibroblast and reduce extracellular matrix deposition in bleomycin (BLM)-challenged mice through the regulation of nuclear factor-erythroid 2-related factor 2 (Nrf2). Additionally, Tan-IIA restored redox homeostasis by upregulating Nrf2 with NADPH oxidase 4 suppression and effectively prevented myofibroblast activation by blocking ROS-mediated protein kinase C delta (PKCδ)/Smad3 signaling. Nrf2 knockdown in the fibroblasts and the lungs of BLM-treated mice reduced the inhibitory effects of Tan-IIA, indicating the essential role of Nrf2 in the Tan-IIA activity. Tan-IIA impaired the binding of kelch-like ECH-associated protein 1 (Keap1) to Nrf2 by promoting the degradation of Keap1 and thereby increasing Nrf2 induction by protecting Nrf2 stability against ubiquitination and proteasomal degradation. Importantly, we also found that the glutamate anaplerotic pathway was involved in energy generation and biosynthesis in activated myofibroblasts and their proliferation. Tan-IIA shunted glutaminolysis into glutathione (GSH) production by activating Nrf2, resulting in the reduction of glutamate availability for tricarboxylic acid cycle. Ultimately, myofibroblast activation was prevented by impairing cell proliferation. Innovation and Conclusion: In addition to the regulation of redox homeostasis, our work showed that Tan-IIA activated Nrf2/GSH signaling pathway to limit glutaminolysis in myofibroblast proliferation, which provided further insight into the critical function of Nrf2 in PF. [ABSTRACT FROM AUTHOR]

Recruitment of immune cells to the site of inflammation by the chemokine CCL1 is important in the pathology of inflammatory diseases. Here, we examined the role of CCL1 in pulmonary fibrosis (PF). Bronchoalveolar lavage fluid from PF mouse models contained high amounts of CCL1, as did lung biopsies from PF patients. Immunofluorescence analyses revealed that alveolar macrophages and CD4+ T cells were major producers of CCL1 and targeted deletion of Ccl1 in these cells blunted pathology. Deletion of the CCL1 receptor Ccr8 in fibroblasts limited migration, but not activation, in response to CCL1. Mass spectrometry analyses of CCL1 complexes identified AMFR as a CCL1 receptor, and deletion of Amfr impaired fibroblast activation. Mechanistically, CCL1 binding triggered ubiquitination of the ERK inhibitor Spry1 by AMFR, thus activating Ras-mediated profibrotic protein synthesis. Antibody blockade of CCL1 ameliorated PF pathology, supporting the therapeutic potential of targeting this pathway for treating fibroproliferative lung diseases. [Display omitted] • CCL1 expression positively correlates with the development of lung fibrosis • CCL1 exerts its fibrogenic activity by signaling through AMFR on fibroblasts • CCL1-AMFR interaction relieves Spry1-repressed Ras-ERK-p70S6K activity • Suppression of the CCL1-AMFR-Ras-ERK-p70S6K axis reduces lung fibrosis Pulmonary fibrosis therapies require improved understanding of the underlying inflammatory mechanisms of disease. Liu et al. demonstrate that immune-derived CCL1 promotes lung fibrosis by two pathways: (1) recruiting CCR8-expressing fibroblasts to the lung and (2) activating lung fibroblasts into pathological myofibroblasts by binding AMFR on fibroblasts, which enhances Ras-ERK signaling and synthesis of profibrotic proteins. [ABSTRACT FROM AUTHOR]

Although recent progress provides mechanistic insights into the pathogenesis of pulmonary fibrosis (PF), rare anti-PF therapeutics show definitive promise for treating this disease. Repeated lung epithelial injury results in injury-repairing response and inflammation, which drive the development of PF. Here, we report that chronic lung injury inactivated the ubiquitin-editing enzyme A20, causing progressive accumulation of the transcription factor C/EBPβ in alveolar macrophages (AMs) from PF patients and mice, which upregulated a number of immunosuppressive and profibrotic factors promoting PF development. In response to chronic lung injury, elevated glycogen synthase kinase-3β (GSK-3β) interacted with and phosphorylated A20 to suppress C/EBPβ degradation. Ectopic expression of A20 or pharmacological restoration of A20 activity by disturbing the A20-GSK-3β interaction accelerated C/EBPβ degradation and showed potent therapeutic efficacy against experimental PF. Our study indicates that a regulatory mechanism of the GSK-3β-A20-C/EBPβ axis in AMs may be a potential target for treating PF and fibroproliferative lung diseases. • A20 activity in alveolar macrophages negatively correlates with lung fibrosis • A20 exerts its anti-fibrotic activities by destabilizing C/EBPβ in AMs • GSK-3β interacts with and phosphorylates A20 to impede its activity • Targeting GSK-3β/A20 interaction reduces lung fibrosis by degrading C/EBPβ Dysregulation of the ubiquitin-editing enzyme A20 contributes to the development of several human inflammatory diseases. Liu et al. demonstrate that suppression of A20 enzymatic activity in alveolar macrophages promotes lung fibrosis by reducing transcriptional factor C/EBPβ degradation and enhancing targeted gene expression, which directs the profibrotic phenotype of macrophages. [ABSTRACT FROM AUTHOR]