The chemokine CCL1 triggers an AMFR-SPRY1 pathway that promotes differentiation of lung fibroblasts into myofibroblasts and drives pulmonary fibrosis.

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.

Targeting Degradation of the Transcription Factor C/EBPß Reduces Lung Fibrosis by Restoring Activity of the Ubiquitin-Editing Enzyme A20 in Macrophages.

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/EBPß 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-3ß (GSK-3ß) interacted with and phosphorylated A20 to suppress C/EBPß degradation. Ectopic expression of A20 or pharmacological restoration of A20 activity by disturbing the A20-GSK-3ß interaction accelerated C/EBPß degradation and showed potent therapeutic efficacy against experimental PF. Our study indicates that a regulatory mechanism of the GSK-3ß-A20-C/EBPß axis in AMs may be a potential target for treating PF and fibroproliferative lung diseases.

TRIB3 Interacts With ß-Catenin and TCF4 to Increase Stem Cell Features of Colorectal Cancer Stem Cells and Tumorigenesis.

BACKGROUND & AIMS: Activation of Wnt signaling to ß-catenin contributes to the development of colorectal cancer (CRC). Expression of tribbles pseudo-kinase 3 (TRIB3) is increased in some colorectal tumors and associated with poor outcome. We investigated whether increased TRIB3 expression promotes stem cell features of CRC cells and tumor progression by interacting with the Wnt signaling pathway. METHODS: We performed studies with C57BL/6J-ApcMin/J mice injected with an adeno-associated virus vector that expresses a small hairpin RNA against Trib3 mRNA (ApcMin/J-Trib3KD) or a control vector (ApcMin/J-Ctrl). We created BALB/c mice that overexpress TRIB3 from an adeno-associated virus vector and mice with small hairpin RNA-mediated knockdown of ß-catenin. The mice were given azoxymethane followed by dextran sodium sulfate to induce colitis-associated cancer. Intestinal tissues were collected and analyzed by histology, gene expression profiling, immunohistochemistry, and immunofluorescence. Leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5)-positive (LGR5Pos) and LGR5-negative (LGR5Neg) HCT-8 CRC cells, with or without knockdown or transgenic expression of TRIB3, were sorted and analyzed in sphere-formation assays. We derived organoids from human and mouse colorectal tumors to analyze the function of TRIB3 and test the effect of a peptide inhibitor. Wnt signaling to ß-catenin was analyzed in dual luciferase reporter, chromatin precipitation, immunofluorescence, and immunoblot assays. Proteins that interact with TRIB3 were identified by immunoprecipitation. CRC cell lines were grown in nude mice as xenograft tumors. RESULTS: At 10 weeks of age, more than half the ApcMin/J-Ctrl mice developed intestinal high-grade epithelial neoplasia, whereas ApcMin/J-Trib3KD mice had no intestinal polyps and normal histology. Colon tissues from ApcMin/J-Trib3KD mice expressed lower levels of genes regulated by ß-catenin and genes associated with cancer stem cells. Mice with overexpression of Trib3 developed more tumors after administration of azoxymethane and dextran sodium sulfate than BALB/c mice. Mice with knockdown of ß-catenin had a lower tumor burden after administration of azoxymethane and dextran sodium sulfate, regardless of Trib3 overexpression. Intestinal tissues from mice with overexpression of Trib3 and knockdown of ß-catenin did not have activation of Wnt signaling or expression of genes regulated by ß-catenin. LGR5Pos cells sorted from HCT-8 cells expressed higher levels of TRIB3 than LGR5Neg cells. CRC cells that overexpressed TRIB3 had higher levels of transcription by ß-catenin and formed larger spheroids than control CRC cells; knockdown of ß-catenin prevented the larger organoid size caused by TRIB3 overexpression. TRIB3 interacted physically with ß-catenin and transcription factor 4 (TCF4). TRIB3 overexpression increased, and TRIB3 knockdown decreased, recruitment of TCF4 and ß-catenin to the promoter region of genes regulated by Wnt. Activated ß-catenin increased expression of TRIB3, indicating a positive-feedback loop. A peptide (P2-T3A6) that bound ß-catenin disrupted its interaction with TRIB3 and TCF4. In primary CRC cells and HCT-8 cells, P2-T3A6 decreased expression of genes regulated by ß-catenin and genes associated with cancer stem cells and decreased cell viability and migration. Injection of C57BL/6J-ApcMin/J mice with P2-T3A6 decreased the number and size of tumor nodules and colon expression of genes regulated by ß-catenin. P2-T3A6 increased 5-fluorouracil-induced death of CRC cells and survival times of mice with xenograft tumors. CONCLUSION: TRIB3 interacts with ß-catenin and TCF4 in intestine cells to increase expression of genes associated with cancer stem cells. Knockdown of TRIB3 decreases colon neoplasia in mice, migration of CRC cells, and their growth as xenograft tumors in mice. Strategies to block TRIB3 activity might be developed for treatment of CRC.

TLR2 agonist Pam3CSK4 enhances the antibacterial functions of GM-CSF induced neutrophils to methicillin-resistant Staphylococcus aureus.

A proliferation of studies have demonstrated that the toll-like receptor 2 (TLR2) pathway affects the chemotaxis, phagocytosis, and cytokine release of neutrophils when pathogens invade. Our previous studies have demonstrated that pretreatment with high doses of Pam3CSK4 (>25 µg/ml) improves the antimicrobial activity of neutrophils, however, short-lived neutrophils limit their therapeutic functions. Here, we used granulocyte macrophage-colony stimulating factor (GM-CSF) to generate neutrophils from murine bone marrow, and assessed their effect on the immune response against methicillin-resistant Staphylococcus aureus. As comparing with classical method of generating neutrophils directly from murine bone marrow, our findings show that pretreatment with Pam3CSK4 enhanced the phagocytic and killing activities against MRSA by the GM-CSF induced neutrophils (GM-CSF neutrophils). Chemotaxis of GM-CSF induced neutrophils was significantly increased after the pretreatment with Pam3CSK4. Furthermore, Pam3CSK4 pretreatment enhanced iNOS, CRAMP, TNF-α, IL-1ß, IL-10, and IL-6 expression. Finally, we observed that p38MAPK and Akt phosphorylation kinases were increased significantly in GM-CSF neutrophils pretreatment with Pam3CSK4 in a dose- and time-dependent manner, whereas p38MAPK inhibitor (SB2021190) and PI3K inhibitor (LY294002) attenuated the antimicrobial activities including phagocytosis, killing activity, respiratory burst, and the release of lactoferrin(LTF) by the GM-CSF induced neutrophils. Together, these findings suggest that pretreatment with Pam3CSK4 enhances the antibacterial function of GM-CSF neutrophils against MRSA, and this could be related to the p38MAPK and PI3K signaling pathways.

Autophagy and the Immune Response.

Innate immunity and adaptive immunity play critical roles in maintaining normal physiological functions and the development of diseases. In innate immune responses, heterogeneous autophagy can directly remove intracellular pathogens while activating PRRs, including TLRs and NLRs, to trigger their signal transduction pathways and promote NKT cell activation, cytokine secretion, and phagocytosis. In adaptive immune responses, the autophagy reaction has an important effect on the homeostasis, function, and differentiation of T lymphocytes, the survival, and development of B lymphocytes and the survival of plasma cells. This review highlights the key role that autophagy plays in the innate immune system and the acquired immune system. Further clarifying the mechanism by which autophagy regulates the immune system is essential for elucidating the precise mechanisms of various diseases and for developing new treatment methods.

Autophagy: a new pathway for traditional Chinese medicine.

Autophagy is a major intracellular degradation pathway that sequesters multiple cytoplasmic components, including accumulated proteins, damaged organelles, or invading micro-organisms and delivers them to lysosomes for degradation. Autophagy dysregulation is implicated in the pathogenesis of multiple diseases, such as aging, cancers, diabetes. The latest insights into molecular mechanisms of autophagy lead to the discovery of potential drug targets. Traditional drugs with new clinical applications are not only commonly found in western medicines, but also highlighted in traditional Chinese medicines (TCMs). Recent research findings shed light on the potential novel applications and formulation of TCMs via regulation of autophagy, indicating autophagy modulation may be an important mechanism underlying the therapeutic effect of TCMs in treating diseases. Here, we summarize the roles of autophagy in the pharmacological actions of TCMs and discuss to discover ideal autophagy modulators from TCMs with considerably higher selectivity for various human disease treatment.

P7, a novel antagonist of corticotropin releasing factor receptor type 1 (CRFR1) screened from phage display library.

The corticotropin releasing factor (CRF) plays a central role in regulating the activities of hypothalamic-pituitary-adrenal (HPA) axis in the presence of a variety of stressful stimuli via binding to its type 1 receptors (CRFR1). Despite that many peptidic or non-peptidic antagonists of CRFR1 have been developed to serve as therapeutic tools to CRF-related pathologies, none of them have been utilized clinically. Targeting the extracellular domain 1 (EC1) of CRFR1, the CRF-binding site, represents a new strategy to inhibit the function of the receptor. However, no such agents have been identified up to now. Herein, by using an 87-amino acid fragment corresponding to the EC1 region as the bait, we screened the binding polypeptides from a phage display (Ph.D.-12) peptide library. After 3-round biopanning, positive clones were selected and the polypeptides carried by them were identified. 5 polypeptides were found to bind with the target specifically. Among them, the P7 exhibited the highest affinity. By evaluating the cAMP accumulation in the CRFR1 or CRFR2-expressing HEK293 cells, we demonstrated that P7 blocking the function of CRFR1, but not CRFR2. In addition, we also found that P7 and CRF act on CRFR1 competitively. Taken together, we reveal that P7, a novel polypeptide identified from phage display library, inhibits the function of CRFR1 effectively and specifically by binding at its EC1 domain. The new polypeptide might provide a promising agent for diagnostic or therapeutic utilities in CRF-related disorders.

Case Report: A case report of myocardial fibrosis activation assessment after unstable angina using 68Ga-FAPI-04 PET/CT.

Myocardial ischemia may induce myocardial fibrosis, a condition that progressively leads to ventricular remodeling, heightening the risk of heart failure. The timely detection of myocardial fibrosis is crucial for intervention and improved outcomes. 68Ga-FAPI-04 PET/CT shows promise in assessing fibroblast activation in patients with early myocardial infarction characterized by prolonged myocardial ischemia. However, there is a notable absence of data regarding patients with short-term myocardial ischemia, such as those experiencing unstable angina (UA). In this report, we evaluated a 49-year-old male with UA and severe stenosis in multiple coronary arteries using 68Ga-FAPI-04 PET/CT. The results demonstrated tracer-specific uptake (SUVmax = 4.6) in the left anterior descending artery (LAD) territory, consistent with myocardial anterior wall ischemia indicated by the electrocardiogram. Following vascular recanalization therapy and regular medication treatment, the patient remained free of angina recurrence. A subsequent review at 2 months revealed a significant reduction in myocardial tracer uptake (SUVmax = 1.8). This case illustrates the validity of 68Ga-FAPI-04 PET/CT in assessing the extent of early myocardial fibroblast activation in patients with UA. This approach offers valuable insights for early detection and visual evidence, providing information on disease progression and treatment response.

FGD5 in basal cells induces CXCL14 secretion that initiates a feedback loop to promote murine mammary epithelial growth and differentiation.

The interactions of environmental compartments with epithelial cells are essential for mammary gland development and homeostasis. Currently, the direct crosstalk between the endothelial niche and mammary epithelial cells remains poorly understood. Here, we show that faciogenital dysplasia 5 (FGD5) is enriched in mammary basal cells (BCs) and mediates critical interactions between basal and endothelial cells (ECs) in the mammary gland. Conditional deletion of Fgd5 reduced, whereas conditional knockin of Fgd5 increased, the engraftment and expansion of BCs, regulating ductal morphogenesis in the mammary gland. Mechanistically, murine mammary BC-expressed FGD5 inhibited the transcriptional activity of activating transcription factor 3 (ATF3), leading to subsequent transcriptional activation and secretion of CXCL14. Furthermore, activation of CXCL14/CXCR4/ERK signaling in primary murine mammary stromal ECs enhanced the expression of HIF-1α-regulated hedgehog ligands, which initiated a positive feedback loop to promote the function of BCs. Collectively, these findings identify functionally important interactions between BCs and the endothelial niche that occur through the FGD5/CXCL14/hedgehog axis.

Targeting IL-17A enhances imatinib efficacy in Philadelphia chromosome-positive B-cell acute lymphoblastic leukemia.

Dysregulated hematopoietic niches remodeled by leukemia cells lead to imbalances in immunological mediators that support leukemogenesis and drug resistance. Targeting immune niches may ameliorate disease progression and tyrosine kinase inhibitor (TKI) resistance in Philadelphia chromosome-positive B-ALL (Ph+ B-ALL). Here, we show that T helper type 17 (Th17) cells and IL-17A expression are distinctively elevated in Ph+ B-ALL patients. IL-17A promotes the progression of Ph+ B-ALL. Mechanistically, IL-17A activates BCR-ABL, IL6/JAK/STAT3, and NF-kB signalling pathways in Ph+ B-ALL cells, resulting in robust cell proliferation and survival. In addition, IL-17A-activated Ph+ B-ALL cells secrete the chemokine CXCL16, which in turn promotes Th17 differentiation, attracts Th17 cells and forms a positive feedback loop supporting leukemia progression. These data demonstrate an involvement of Th17 cells in Ph+ B-ALL progression and suggest potential therapeutic options for Ph+ B-ALL with Th17-enriched niches.

TRIB3 promotes pulmonary fibrosis through inhibiting SLUG degradation by physically interacting with MDM2.

Pulmonary fibrosis (PF) is the pathological structure of incurable fibroproliferative lung diseases that are attributed to the repeated lung injury-caused failure of lung alveolar regeneration (LAR). Here, we report that repetitive lung damage results in a progressive accumulation of the transcriptional repressor SLUG in alveolar epithelial type II cells (AEC2s). The abnormal increased SLUG inhibits AEC2s from self-renewal and differentiation into alveolar epithelial type I cells (AEC1s). We found that the elevated SLUG represses the expression of the phosphate transporter SLC34A2 in AEC2s, which reduces intracellular phosphate and represses the phosphorylation of JNK and P38 MAPK, two critical kinases supporting LAR, leading to LAR failure. TRIB3, a stress sensor, interacts with the E3 ligase MDM2 to suppress SLUG degradation in AEC2s by impeding MDM2-catalyzed SLUG ubiquitination. Targeting SLUG degradation by disturbing the TRIB3/MDM2 interaction using a new synthetic staple peptide restores LAR capacity and exhibits potent therapeutic efficacy against experimental PF. Our study reveals a mechanism of the TRIB3-MDM2-SLUG-SLC34A2 axis causing the LAR failure in PF, which confers a potential strategy for treating patients with fibroproliferative lung diseases.

Mesaconine alleviates doxorubicin-triggered cardiotoxicity and heart failure by activating PINK1-dependent cardiac mitophagy.

Aberrant mitophagy has been identified as a driver for energy metabolism disorder in most cardiac pathological processes. However, finding effective targeted agents and uncovering their precise modulatory mechanisms remain unconquered. Fuzi, the lateral roots of Aconitum carmichaelii, shows unique efficacy in reviving Yang for resuscitation, which has been widely used in clinics. As a main cardiotonic component of Fuzi, mesaconine has been proven effective in various cardiomyopathy models. Here, we aimed to define a previously unrevealed cardioprotective mechanism of mesaconine-mediated restoration of obstructive mitophagy. The functional implications of mesaconine were evaluated in doxorubicin (DOX)-induced heart failure models. DOX-treated mice showed characteristic cardiac dysfunction, ectopic myocardial energy disorder, and impaired mitophagy in cardiomyocytes, which could be remarkably reversed by mesaconine. The cardioprotective effect of mesaconine was primarily attributed to its ability to promote the restoration of mitophagy in cardiomyocytes, as evidenced by elevated expression of PINK1, a key mediator of mitophagy induction. Silencing PINK1 or deactivating mitophagy could completely abolish the protective effects of mesaconine. Together, our findings suggest that the cardioprotective effects of mesaconine appear to be dependent on the activation of PINK1-induced mitophagy and that mesaconine may constitute a promising therapeutic agent for the treatment of heart failure.

External Ba2+ block of the two-pore domain potassium channel TREK-1 defines conformational transition in its selectivity filter.

TREK-1 is a member of the two-pore domain potassium channel family that is known as a leak channel and plays a key role in many physiological and pathological processes. The conformational transition of the selectivity filter is considered as an effective strategy for potassium channels to control the course of potassium efflux. It is well known that TREK-1 is regulated by a large volume of extracellular and intracellular signals. However, until now, little was known about the selectivity filter gating mechanism of the channel. In this research, it was found that Ba(2+) blocked the TREK-1 channel in a concentration- and time-dependent manner. A mutagenesis analysis showed that overlapped binding of Ba(2+) at the assumed K(+) binding site 4 (S4) within the selectivity filter was responsible for the inhibitory effects on TREK-1. Then, Ba(2+) was used as a probe to explore the conformational transition in the selectivity filter of the channel. It was confirmed that collapsed conformations were induced by extracellular K(+)-free and acidification at the selectivity filters, leading to nonconductive to permeable ions. Further detailed characterization demonstrated that the two conformations presented different properties. Additionally, the N-terminal truncated isoform (ΔN41), a product derived from alternative translation initiation, was identified as a constitutively nonconductive variant. Together, these results illustrate the important role of selectivity filter gating in the regulation of TREK-1 by the extracellular K(+) and proton.

The cell cycle inhibitor P21 promotes the development of pulmonary fibrosis by suppressing lung alveolar regeneration.

The cell cycle inhibitor P21 has been implicated in cell senescence and plays an important role in the injury-repair process following lung injury. Pulmonary fibrosis (PF) is a fibrotic lung disorder characterized by cell senescence in lung alveolar epithelial cells. In this study, we report that P21 expression was increased in alveolar epithelial type 2 cells (AEC2s) in a time-dependent manner following multiple bleomycin-induced PF. Repeated injury of AEC2s resulted in telomere shortening and triggered P21-dependent cell senescence. AEC2s with elevated expression of P21 lost their self-renewal and differentiation abilities. In particular, elevated P21 not only induced cell cycle arrest in AEC2s but also bound to P300 and ß-catenin and inhibited AEC2 differentiation by disturbing the P300-ß-catenin interaction. Meanwhile, senescent AEC2s triggered myofibroblast activation by releasing profibrotic cytokines. Knockdown of P21 restored AEC2-mediated lung alveolar regeneration in mice with chronic PF. The results of our study reveal a mechanism of P21-mediated lung regeneration failure during PF development, which suggests a potential strategy for the treatment of fibrotic lung diseases.

Faciogenital Dysplasia 5 supports cancer stem cell traits in basal-like breast cancer by enhancing EGFR stability.

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.

TRIB3-EGFR interaction promotes lung cancer progression and defines a therapeutic target.

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.

Disruption of the EGFR-SQSTM1 interaction by a stapled peptide suppresses lung cancer via activating autophagy and inhibiting EGFR signaling.

Despite the success of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in the treatment of non-small cell lung cancer (NSCLC) harboring EGFR-activating mutations, intrinsic or acquired resistance remains the major obstacle to long-term disease remission. Defective autophagy has been reported as an EGFR-TKI resistance mechanism. However, how EGFR regulate autophagic flux are still not fully understood. Here we found that EGFR-stimulated phosphorylation of SQSTM1 at tyrosine 433 induces dimerization of its UBA domain, which disturbs the sequestration function of SQSTM1 and causes autophagic flux blocking. SAH-EJ2, a staple optimized EGFR-derived peptide, showed enhanced in vitro and in vivo antitumor activity against NSCLC than the prototype regardless of EGFR mutation status. Mechanistically, SAH-EJ2 disrupts the EGFR-SQSTM1 interaction and protects against EGFR-induced SQSTM1 phosphorylation, which hinders the dimerization of the SQSTM1 UBA domains and restores SQSTM1 cargo function. Moreover, SAH-EJ2 suppresses EGFR activity by blocking its dimerization and reducing its protein stability, which reciprocally activates the core autophagy machinery. Our observations reveal that disturbing the EGFR-SQSTM1 interaction by SAH-EJ2 confers a potential strategy in the treatment of NSCLC through suppressing EGFR signalling and activating autophagy simultaneously.

TRIB3 promotes MYC-associated lymphoma development through suppression of UBE3B-mediated MYC degradation.

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.

Disrupting the TRIB3-SQSTM1 interaction reduces liver fibrosis by restoring autophagy and suppressing exosome-mediated HSC activation.

Impaired macroautophagy/autophagy is involved in the pathogenesis of hepatic fibrosis. However, how aberrant autophagy promotes fibrosis is far from understood. Here, we aimed to define a previously unrevealed pro-fibrotic mechanism for the stress protein TRIB3 (tribbles pseudokinase 3)-mediated autophagy dysfunction. Human fibrotic liver tissues were obtained from patients with cirrhosis who underwent an open surgical repair process. The functional implications of TRIB3 were evaluated in mouse models of hepatic fibrosis induced by bile duct ligation (BDL) or thioacetamide (TAA) injection. Human fibrotic liver tissues expressed higher levels of TRIB3 and selective autophagic receptor SQSTM1/p62 (sequestosome 1) than nonfibrotic tissues and the elevated expression of TRIB3 and SQSTM1 was positively correlated in the fibrotic tissues. Silencing Trib3 protected against experimentally induced hepatic fibrosis, accompanied by restored autophagy activity in fibrotic liver tissues. Furthermore, TRIB3 interacted with SQSTM1 and hindered its binding to MAP1LC3/LC3, which caused the accumulation of SQSTM1 aggregates and obstructed autophagic flux. The TRIB3-mediated autophagy impairment not only suppressed autophagic degradation of late endosomes but also promoted hepatocellular secretion of INHBA/Activin A-enriched exosomes which caused migration, proliferation and activation of hepatic stellate cells (HSCs), the effector cells of liver fibrosis. Disrupting the TRIB3-SQSTM1 interaction with a specific helical peptide exerted potent protective effects against hepatic fibrosis by restoring autophagic flux in hepatocytes and HSCs. Together, stress-elevated TRIB3 expression promotes hepatic fibrosis by interacting with SQSTM1 and interfering with its functions in liver-parenchymal cells and activating HSCs. Targeting this interaction is a promising strategy for treating fibroproliferative liver diseases.Abbreviations: 3-MA: 3-methyladenine; AAV: adeno-associated virus; ACTA2/α-SMA: actin, alpha 2, smooth muscle, aorta; BDL: bile duct ligation; BECN1/Beclin 1: beclin 1, autophagy related; CHX: cycloheximide; CQ: chloroquine; Edu: 5-ethynyl-2-deoxyuridine; ESCRT: endosomal sorting complexes required for transport; HSC: hepatic stellate cell; ILV: intralumenal vesicle; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MVB: multivesicular body; PIK3C3: phosphatidylinositol 3-kinase, catalytic subunit type 3; PPI: protein-protein interaction; SQSTM1/p62: sequestosome 1; TAA: thioacetamide; TEM: transmission electron microscopy; TGFB1/TGFß1: transforming growth factor, beta 1; TLR2: toll-like receptor 2; TRIB3: tribbles pseudokinase 3.