DAP3 promotes mitochondrial activity and tumour progression in hepatocellular carcinoma by regulating MT-ND5 expression.

Cancer cells often exhibit fragmented mitochondria and dysregulated mitochondrial dynamics, but the underlying mechanism remains elusive. Here, we found that the mitochondrial protein death-associated protein 3 (DAP3) is localized to mitochondria and promotes the progression of hepatocellular carcinoma (HCC) by regulating mitochondrial function. DAP3 can promote the proliferation, migration, and invasion of HCC cells in vitro and in vivo by increasing mitochondrial respiration, inducing the epithelial-mesenchymal transition (EMT), and slowing cellular senescence. Mechanistically, DAP3 can increase mitochondrial complex I activity in HCC cells by regulating the translation and expression of MT-ND5. The phosphorylation of DAP3 at Ser185 mediated by AKT is the key event mediating the mitochondrial localization and function of DAP3 in HCC cells. In addition, the DAP3 expression in HCC samples is inversely correlated with patient survival. Our results revealed a mechanism by which DAP3 promotes mitochondrial function and HCC progression by regulating MT-ND5 translation and expression, indicating that DAP3 may be a therapeutic target for HCC.

Remdesivir alleviates skin fibrosis by suppressing TGF-ß1 signaling pathway.

Fibrotic skin diseases, such as keloids, are pathological results of aberrant tissue healing and are characterized by overgrowth of dermal fibroblasts. Remdesivir (RD), an antiviral drug, has been reported to have pharmacological activities in a wide range of fibrotic diseases. However, whether RD function on skin fibrosis remains unclear. Therefore, in our study, we explored the potential effect and mechanisms of RD on skin fibrosis both in vivo and in vitro. As expected, the results demonstrated that RD alleviated BLM-induced skin fibrosis and attenuates the gross weight of keloid tissues in vivo. Further studies suggested that RD suppressed fibroblast activation and autophagy both in vivo and in vitro. In addition, mechanistic research showed that RD attenuated fibroblasts activation by the TGF-ß1/Smad signaling pathway and inhibited fibroblasts autophagy by the PI3K/Akt/mTOR signaling pathway. In summary, our results demonstrate therapeutic potential of RD for skin fibrosis in the future.

Targeted isolation, identification, and antioxidant evaluation of aromatic polyketides from a plant-derived fungus Ophiobolus cirsii LZU-1509.

There are >350 species of the Ophiobolus genus, which is not yet very well-known and lacks research reports on secondary metabolites. Three new 3,4-benzofuran polyketides 1-3, a new 3,4-benzofuran polyketide racemate 4, two new pairs of polyketide enantiomers (±)-5 and (±)-7, two new acetophenone derivatives 6 and 8, and three novel 1,4-dioxane aromatic polyketides 9-11, were isolated from a fungus Ophiobolus cirsii LZU-1509 derived from an important medicinal and economic crop Anaphalis lactea. The isolation was guided by LC-MS/MS-based GNPS molecular networking analysis. The planar structures and relative configurations were mainly elucidated by NMR and HR-ESI-MS data. Their absolute configurations were determined by using X-ray diffraction analysis and via comparing computational and experimental ECD, NMR, and specific optical rotation data. 9 possesses an unreported 5/6/6/6/5 five-ring framework with a 1,4-dioxane, and 10 and 11 feature unprecedented 6/6/6/5 and 6/6/5/6 four-ring frames containing a 1,4-dioxane. The biosynthetic pathways of 9-11 were proposed. 1-11 were nontoxic in HT-1080 and HepG2 tumor cells at a concentration of 20 µM, whereas 3 and 5 exerted higher antioxidant properties in the hydrogen peroxide-stimulated model in the neuron-like PC12 cells. They could be potential antioxidant agents for neuroprotection.

Resveratrol protects against myocardial ischemic injury in obese mice via activating SIRT3/FOXO3a signaling pathway and restoring redox homeostasis.

BACKGROUND: Increasing global overweight and obesity rates not only increase the prevalence of myocardial infarction (MI), but also exacerbate ischemic injury and result in worsened prognosis. Currently, there are no drugs that can reverse myocardial damage once MI has occurred, therefore discovering drugs that can potentially limit the extent of ischemic damage to the myocardium is critical. Resveratrol is a polyphenol known for its antioxidant properties, however whether prolonged daily intake of resveratrol during obesity can protect against MI-induced damage remains unexplored. METHODS: We established murine models of obesity via high-fat/high-fructose diet, along with daily administrations of resveratrol or vehicle, then performed surgical MI to examine the effects and mechanisms of resveratrol in protecting against myocardial ischemic injury. RESULTS: Daily administration of resveratrol in obese mice robustly protected against myocardial ischemic injury and improved post-MI cardiac function. Resveratrol strongly inhibited oxidative and DNA damage via activating SIRT3/FOXO3a-dependent antioxidant enzymes following MI, which were completely prevented upon administration of 3-TYP, a selective SIRT3 inhibitor. Hence, the cardioprotective effects of prolonged resveratrol intake in protecting obese mice against myocardial ischemic injury was due to reestablishment of intracellular redox homeostasis through activation of SIRT3/FOXO3a signaling pathway. CONCLUSION: Our findings provide important new evidence that supports the daily intake of resveratrol, especially in those overweight or obese, which can robustly decrease the extent of ischemic damage following MI. Our study therefore provides new mechanistic insight and suggests the therapeutic potential of resveratrol as an invaluable drug in the treatment of ischemic heart diseases.

The potential of herbal drugs to treat heart failure: The roles of Sirt1/AMPK.

Heart failure (HF) is a highly morbid syndrome that seriously affects the physical and mental health of patients and generates an enormous socio-economic burden. In addition to cardiac myocyte oxidative stress and apoptosis, which are considered mechanisms for the development of HF, alterations in cardiac energy metabolism and pathological autophagy also contribute to cardiac abnormalities and ultimately HF. Silent information regulator 1 (Sirt1) and adenosine monophosphate-activated protein kinase (AMPK) are nicotinamide adenine dinucleotide (NAD+)-dependent deacetylases and phosphorylated kinases, respectively. They play similar roles in regulating some pathological processes of the heart through regulating targets such as peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), protein 38 mitogen-activated protein kinase (p38 MAPK), peroxisome proliferator-activated receptors (PPARs), and mammalian target of rapamycin (mTOR). We summarized the synergistic effects of Sirt1 and AMPK in the heart, and listed the traditional Chinese medicine (TCM) that exhibit cardioprotective properties by modulating the Sirt1/AMPK pathway, to provide a basis for the development of Sirt1/AMPK activators or inhibitors for the treatment of HF and other cardiovascular diseases (CVDs).

Mechanistic regulation of FOXO transcription factors in the nucleus.

FOXO proteins represent evolutionarily conserved transcription factors (TFs) that play critical roles in responding to various physiological signals or pathological stimuli, either through transcription-dependent or -independent mechanisms. Dysfunction of these proteins have been implicated in numerous diseases, including cancer. Although the regulation of FOXO TFs shuttling between the cytoplasm and the nucleus has been extensively studied and reviewed, there's still a lack of a comprehensive review focusing on the intricate interactions between FOXO, DNA, and cofactors in the regulation of gene expression. In this review, we aim to summarize recent advances and provide a detailed understanding of the mechanism underlying FOXO proteins binding to target DNA. Additionally, we will discuss the challenges associated with pharmacological approaches in modulating FOXO function, and explore the dynamic association between TF, DNA, and RNA on chromatin. This review will contribute to a better understanding of mechanistic regulations of eukaryotic TFs within the nucleus.

Slik maintains tissue homeostasis by preventing JNK-mediated apoptosis.

BACKGROUND: The c-Jun N-terminal kinase (JNK) pathway is an evolutionarily conserved regulator of cell death, which is essential for coordinating tissue homeostasis. In this study, we have characterized the Drosophila Ste20-like kinase Slik as a novel modulator of JNK pathway-mediated apoptotic cell death. RESULTS: First, ectopic JNK signaling-triggered cell death is enhanced by slik depletion whereas suppressed by Slik overexpression. Second, loss of slik activates JNK signaling, which results in enhanced apoptosis and impaired tissue homeostasis. In addition, genetic epistasis analysis suggests that Slik acts upstream of or in parallel to Hep to regulate JNK-mediated apoptotic cell death. Moreover, Slik is necessary and sufficient for preventing physiologic JNK signaling-mediated cell death in development. Furthermore, introduction of STK10, the human ortholog of Slik, into Drosophila restores slik depletion-induced cell death and compromised tissue homeostasis. Lastly, knockdown of STK10 in human cancer cells also leads to JNK activation, which is cancelled by expression of Slik. CONCLUSIONS: This study has uncovered an evolutionarily conserved role of Slik/STK10 in blocking JNK signaling, which is required for cell death inhibition and tissue homeostasis maintenance in development.

Rotationplasty type BIIIb as an effective alternative to limb salvage procedure in adults: Two case reports.

BACKGROUND: Rotationplasty is often performed for malignant tumors, but type BIIIb rotationplasty is rarely reported, and there needs to be more evidence of the procedure and treatment. The purpose of this case study was to report a new direction in the use of type BIIIb rotationplasty in treating patients with limb salvage and long-term non-healing infections. CASE SUMMARY: Case 1: A 47-year-old man underwent radiotherapy for hemangioendothelioma in his left thigh, resulting in a femoral fracture. Despite the use of plates, intramedullary nailing, and external fixators, the femoral bone failed to unite due to infectious nonunion. Multiple operations were unable to control the infection, leaving the patient immobile. We performed a modified tibia-pelvic-constrained hip rotationplasty, utilizing a constrained prosthetic hip between the tibia and pelvis following a femur resection. Two years post-surgery, the patient was able to walk with the prosthetic device without any signs of recurring infection. The corresponding functional scores were 72 points for the Musculoskeletal Tumor Society (MSTS), 53 for the Functional Mobility Assessment (FMA), 93 for the Toronto Extremity Salvage Score (TESS), and 56 for the MOS 36-item short form health survey (SF-36). Case 2: A 59-year-old woman presented with liposarcoma in her left thigh. Magnetic resonance imaging revealed tumors in the medial, anterior, and posterior femur muscles, encircling the femoral vessels and nerves. Fortunately, there were no symptoms of sciatic dysfunction, and the tumor had not invaded the sciatic nucleus. After one year of follow-up, the patient expressed satisfaction with limb preservation post-type BIIIb rotationplasty. The corresponding functional scores were 63 points for the MSTS, 47 for the FMA, 88 for the TESS, and 52 for the SF-36. CONCLUSION: Our study suggests that type BIIIb rotationplasty may be an alternative to amputation in patients with incurable infections. For malignant tumors of the lower extremities without invasion of the sciatic nerve, type BIIIb rotationplasty remains an excellent alternative to amputation. This surgical method may prevent amputation, improve functional outcomes, and facilitate biological reconstruction.

Transcription factor Zhx2 is a checkpoint that programs macrophage polarization and antitumor response.

Macrophages are usually educated to tumor-associated macrophages (TAMs) in cancer with pro-tumor functions by tumor microenvironment (TME) and TAM reprogramming has been proposed as a potential tumor immunotherapy strategy. We recently demonstrated the critical role of Zinc-fingers and homeoboxes 2 (Zhx2) in macrophages' metabolic programming. However, whether Zhx2 is responsible for macrophage polarization and TAMs reprogramming is largely unknown. Here, we show that Zhx2 controls macrophage polarization under the inflammatory stimulus and TME. Myeloid-specific deletion of Zhx2 suppresses LPS-induced proinflammatory polarization but promotes IL-4 and TME-induced anti-inflammatory and pro-tumoral phenotypes in murine liver tumor models. Factors in TME, especially lactate, markedly decrease the expression of Zhx2 in TAMs, leading to the switch of TAMs to pro-tumor phenotype and consequent cancer progression. Notably, reduced ZHX2 expression in TAM correlates with poor survival of HCC patients. Mechanistic studies reveal that Zhx2 associates with NF-κB p65 and binds to the Irf1 promoter, leading to transcriptional activation of Irf1 in macrophages. Zhx2 functions in maintaining macrophage polarization by regulating Irf1 transcription, which may be a potential target for macrophage-based cancer immunotherapy.

PVA Electrospun Fibers Coated with PPy Nanoparticles for Wearable Strain Sensors.

Current conductive polymers win wide applications in smart strain-stress sensors, bioinspired actuators, and wearable electronics. This work investigates a novel strain sensor by using conductive polypyrrole (PPy) nanoparticles coated polyvinyl alcohol (PVA) fibers as matrix. The flexible, water-resistant PVA fibers are initially prepared by combined electrospinning and annealing techniques, and then are coated with PPy nanoparticles through in situ polymerization. The resultant PPy@PVA fibers exhibit stable, favorable electrical conductivities due to the uniform point-to-point connections among PPy nanoparticles, e.g. after three-time' polymerizations, the PPy@PVA3 fiber film presents a sheet resistance of ≈840 Ω sq-1 and a bulk conductivity of ≈32.1 mS cm-1 . Cyclic sensing tests reveal that, PPy@PVA sensors show linear relationships between the relative resistance variations and the applied strains, e.g. the linear deviation of PPy@PVA3 is only 0.9 % within 33 % strain. After long-term stretching/releasing cycles, the PPy@PVA sensor exhibits stable, durable, and reversible sensing behaviors, no evident "drift" is observed over 1,000 cycles (5,000 seconds).

Comparison the treatment of anterior inferior tibiofibular ligament anatomical repair and syndesmosis screw fixation for syndesmotic injuries in ankle fracture.

BACKGROUND: The fixation method of syndesmotic injuries in ankle fractures remains controversial. The goal of the study was to compare radiographic and clinical outcomes between anterior inferior tibiofibular ligament (AITFL) anatomical repair with syndesmosis screw fixation in syndesmotic injuries. METHODS: We analyzed 62 patients who were treated with AITFL anatomical repair or syndesmosis screw fixation for syndesmotic injuries in an advanced teaching hospital between March 2016 and March 2019. Fixation was performed with AITFL anatomical repair in 30 patients (AAR group) and syndesmosis screw in 32 patients (SS group). Radiographic evaluations were the differences in mean anterior and posterior (A difference and P difference) tibiofibular distance between injured and uninjured ankle computed tomography (CT) scan at 6 months postoperatively. Clinical evaluation of patients was done using the American Orthopaedic Foot & Ankle Society (AOFAS) Ankle Hindfoot Score, the Olerud-Molander Ankle (OMA) score and visual analogue scale (VAS) score at 1, 3, 6 months and 1, 2 years postoperatively. RESULTS: The A difference and P difference on CT was no differences (1.6 ± 0.8 mm, 1.3 ± 0.7 mm vs. 1.5 ± 0.7 mm, 1.2 ± 0.7 mm) between the two groups (All of P > 0.05). The AAR group had higher mean AOFAS score (65.6 ± 5.9, 82.3 ± 4.2, 87.6 ± 5.6 vs. 61.8 ± 5.2, 79.1 ± 4.0, 83.8 ± 4.9; P = 0.008, 0.003, 0.007) and higher mean OMA score (45.7 ± 8.7, 79.2 ± 6.5, 84.1 ± 5.3 vs. 40.4 ± 7.3, 74.8 ± 6.3, 80.3 ± 5.8; P = 0.012, 0.009, 0.010)) at 1, 3 and 6 months postoperatively. The AAR group had lower mean VAS scores (2.6 ± 1.2, 1.7 ± 0.7 vs. 3.4 ± 1.2, 2.2 ± 1.1; P = 0.018, 0.038) at 1 and 3 months postoperatively. CONCLUSIONS: The results of this study suggest that the AITFL anatomical repair technique could effectively improve ankle function during daily activity. Therefore, AITFL anatomical repair technique is expected to become a better fixation method for syndesmotic injuries.

Isolation and Identification of Novel Antioxidant Polyketides from an Endophytic Fungus Ophiobolus cirsii LZU-1509.

Sixteen new polyketides, ophicirsins A-P (1-16), including four novel carbon skeletons (5-9, 14, 15, and 16), were isolated from the extract of an endophytic fungus Ophiobolus cirsii LZU-1509. The unique frameworks of ophicirsin N (14) and O (15) feature a different cyclic ether connected with an aromatic ring system. Ophicirsin P (16) is characterized by the unprecedented heterozygote of a polyketide and an alkaloid. The absolute stereochemistries of those polyketides were characterized via single-crystal X-ray diffraction analysis and the experimental and computational electric circular dichroism spectra comparison. Theoretical reaction pathways in the fermentation to generate different novel skeletons starting from acetyl CoA and malonyl CoA helped to assign their structures. Compounds 1-16 appear almost nontoxic in HepG2 and HT-1080 tumor cells. Their antioxidant effects were further evaluated, and 15 exhibits an excellent protection activity in hydrogen peroxide-stimulated oxidative damage in neuron-like PC12 cells via screening all compounds. Moreover, 15 displays a greater ability to scavenge the 2,2-diphenyl-1-picrylhydrazyl free radicals than resveratrol. Taken together, these findings suggest that the novel polyketides could serve as potential antioxidant agents for neuroprotection.

NAD + salvage governs mitochondrial metabolism, invigorating natural killer cell antitumor immunity.

BACKGROUND AND AIMS: Natural killer (NK) cells are key players in tumor immunosurveillance, and metabolic adaptation manipulates their fate and functional state. The nicotinamide adenine dinucleotide (NAD + ) has emerged as a vital factor to link cellular metabolism and signaling transduction. Here, we identified NAD + metabolism as a central hub to determine the homeostasis and function of NK cells. APPROACH AND RESULTS: NAD + level was elevated in activated NK cells. NAD + supplementation not only enhanced cytokine production and cytotoxicity but also improved the proliferation and viability of NK cells. Intriguingly, the salvage pathway was involved in maintaining NAD + homeostasis in activated NK cells. Genetic ablation or pharmacological blockade of nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the NAD + salvage pathway, markedly destroyed the viability and function of NK cells. Mechanistically, NAD + salvage dictated the mitochondrial homeostasis and oxidative phosphorylation activity to support the optimal function of NK cells. However, in human HCC tissues, NAMPT expression and NAD + level were significantly down-regulated in tumor-infiltrating NK cells, which negatively correlated with patient survival. And lactate accumulation in the tumor microenvironment was at least partially responsible for the transcriptional repression of NAMPT in NK cells. Further, deficiency of Nampt in NK cells accelerated the growth of HCC and melanoma. Supplementation of the NAD + precursor nicotinamide mononucleotide (NMN) significantly improved NK antitumor response in both mouse and human cell-derived xenografts. CONCLUSIONS: These findings reveal NAD + salvage as an essential factor for NK-cell homeostasis and function, suggesting a potential strategy for invigorating NK cell-based immunotherapy.

Ptp61F integrates Hippo, TOR, and actomyosin pathways to control three-dimensional organ size.

Precise organ size control is fundamental for all metazoans, but how organ size is controlled in a three-dimensional (3D) way remains largely unexplored at the molecular level. Here, we screen and identify Drosophila Ptp61F as a pivotal regulator of organ size that integrates the Hippo pathway, TOR pathway, and actomyosin machinery. Pathologically, Ptp61F loss synergizes with RasV12 to induce tumorigenesis. Physiologically, Ptp61F depletion increases body size and drives neoplastic intestinal tumor formation and stem cell proliferation. Ptp61F also regulates cell contractility and myosin activation and controls 3D cell shape by reducing cell height and horizontal cell size. Mechanistically, Ptp61F forms a complex with Expanded (Ex) and increases endosomal localization of Ex and Yki. Furthermore, we demonstrate that PTPN2, the conserved human ortholog of Ptp61F, can functionally substitute for Ptp61F in Drosophila. Our work defines Ptp61F as an essential determinant that controls 3D organ size under both physiological and pathological conditions.

Bip-Yorkie interaction determines oncogenic and tumor-suppressive roles of Ire1/Xbp1s activation.

Unfolded protein response (UPR) is the mechanism by which cells control endoplasmic reticulum (ER) protein homeostasis. ER proteostasis is essential to adapt to cell proliferation and regeneration in development and tumorigenesis, but mechanisms linking UPR, growth control, and cancer progression remain unclear. Here, we report that the Ire1/Xbp1s pathway has surprisingly oncogenic and tumor-suppressive roles in a context-dependent manner. Activation of Ire1/Xbp1s up-regulates their downstream target Bip, which sequesters Yorkie (Yki), a Hippo pathway transducer, in the cytoplasm to restrict Yki transcriptional output. This regulation provides an endogenous defensive mechanism in organ size control, intestinal homeostasis, and regeneration. Unexpectedly, Xbp1 ablation promotes tumor overgrowth but suppresses invasiveness in a Drosophila cancer model. Mechanistically, hyperactivated Ire1/Xbp1s signaling in turn induces JNK-dependent developmental and oncogenic cell migration and epithelial-mesenchymal transition (EMT) via repression of Yki. In humans, a negative correlation between XBP1 and YAP (Yki ortholog) target gene expression specifically exists in triple-negative breast cancers (TNBCs), and those with high XBP1 or HSPA5 (Bip ortholog) expression have better clinical outcomes. In human TNBC cell lines and xenograft models, ectopic XBP1s or HSPA5 expression alleviates tumor growth but aggravates cell migration and invasion. These findings uncover a conserved crosstalk between the Ire1/Xbp1s and Hippo signaling pathways under physiological settings, as well as a crucial role of Bip-Yki interaction in tumorigenesis that is shared from Drosophila to humans.

A conserved interplay between FOXO and SNAI/snail in autophagy.

Dysfunction of macroautophagy/autophagy has been implicated in homeostasis maintenance and contributes to various diseases. Yet the mechanisms that regulate autophagy have not been fully understood. In a recent study, we uncovered a coherent FOXO3-SNAI2 feed-forward regulatory loop in mammals that reinforces autophagy gene induction upon energy stress. Strikingly, a foxo-sna (snail) feed-forward circuit also exists in Drosophila, suggesting this regulating loop is evolutionarily conserved. Moreover, our results highlight that binding of FOXO3 to the DNA appears to be both necessary and sufficient to antagonize CRM1-dependent nuclear export, illustrating a critical role of DNA in regulating protein nuclear localization.

A coherent FOXO3-SNAI2 feed-forward loop in autophagy.

SignificanceUnderstanding autophagy regulation is instrumental in developing therapeutic interventions for autophagy-associated disease. Here, we identified SNAI2 as a regulator of autophagy from a genome-wide screen in HeLa cells. Upon energy stress, SNAI2 is transcriptionally activated by FOXO3 and interacts with FOXO3 to form a feed-forward regulatory loop to reinforce the expression of autophagy genes. Of note, SNAI2-increased FOXO3-DNA binding abrogates CRM1-dependent FOXO3 nuclear export, illuminating a pivotal role of DNA in the nuclear retention of nucleocytoplasmic shuttling proteins. Moreover, a dFoxO-Snail feed-forward loop regulates both autophagy and cell size in Drosophila, suggesting this evolutionarily conserved regulatory loop is engaged in more physiological activities.

Phosphoinositide-Binding Protein TIPE1 Promotes Alternative Activation of Macrophages and Tumor Progression via PIP3/Akt/TGFß Axis.

Macrophages perform key and distinct functions in maintaining tissue homeostasis by finely tuning their activation state. Within the tumor microenvironment, macrophages are reshaped to drive tumor progression. Here we report that tumor necrosis factor α-induced protein 8-like 1 (TIPE1) is highly expressed in macrophages and that depletion of TIPE1 impedes alternative activation of macrophages. TIPE1 enhanced activation of the PI3K/Akt pathway in macrophages by directly binding with and regulating the metabolism of phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 3,4,5-trisphosphate (PIP3). Accordingly, inhibition of the PI3K/Akt pathway significantly attenuated the effect of TIPE1 on macrophage alternative activation. Tumor-associated macrophages (TAM) in human liver cancer and melanoma tissues showed significantly upregulated TIPE1 expression that negatively correlated with patient survival. In vitro and in vivo, TIPE1 knockdown in macrophages retarded the growth and metastasis of liver cancer and melanoma. Furthermore, blockade or depletion of TGFß signaling in macrophages abrogated the effects of TIPE1 on tumor cell growth and migration. Together, these results highlight that the phosphoinositide-related signaling pathway is involved in reprogramming TAMs to optimize the microenvironment for cancer progression. SIGNIFICANCE: This work provides insight into the fine tuning of macrophage polarization and identifies a potential target for macrophage-based antitumor therapy.

Myc suppresses male-male courtship in Drosophila.

Despite strong natural selection on species, same-sex sexual attraction is widespread across animals, yet the underlying mechanisms remain elusive. Here, we report that the proto-oncogene Myc is required in dopaminergic neurons to inhibit Drosophila male-male courtship. Loss of Myc, either by mutation or neuro-specific knockdown, induced males' courtship propensity toward other males. Our genetic screen identified DOPA decarboxylase (Ddc) as a downstream target of Myc. While loss of Ddc abrogated Myc depletion-induced male-male courtship, Ddc overexpression sufficed to trigger such behavior. Furthermore, Myc-depleted males exhibited elevated dopamine level in a Ddc-dependent manner, and their male-male courtship was blocked by depleting the dopamine receptor DopR1. Moreover, Myc directly inhibits Ddc transcription by binding to a target site in the Ddc promoter, and deletion of this site by genome editing was sufficient to trigger male-male courtship. Finally, drug-mediated Myc depletion in adult neurons by GeneSwitch technique sufficed to elicit male-male courtship. Thus, this study uncovered a novel function of Myc in preventing Drosophila male-male courtship, and supports the crucial roles of genetic factors in inter-male sexual behavior.

Spatial distribution and functional analysis define the action pathway of Tim-3/Tim-3 ligands in tumor development.

The spatial organization of immune cells within the tumor microenvironment (TME) largely determines the anti-tumor immunity and also highly predicts tumor progression and therapeutic response. Tim-3 is a well-accepted immune checkpoint and plays multifaceted immunoregulatory roles via interaction with distinct Tim-3 ligands (Tim-3L), showing great potential as an immunotherapy target. However, the cell sociology mediated by Tim-3/Tim-3L and their contribution to tumor development remains elusive. Here, we analyzed the spatial distribution of Tim-3/Tim-3L in TME using multiplex fluorescence staining and revealed that despite the increased Tim-3 expression in various tumor-infiltrated lymphocytes, Tim-3+CD4+ cells were more accumulated in parenchymal/tumor region compared with stromal region and exhibited more close association with patient survival. Strikingly, CD4 T cells surrounding Tim-3L+ cells expressed higher Tim-3 than other cells in cancerous tissues. In vivo studies confirmed that depletion of CD4 T cells completely abrogated the inhibition of tumor growth and metastasis, as well as the functional improvement of CD8 T and NK, mediated by Tim-3 blockade, which was further validated in peripheral lymphocytes from patients with hepatocellular carcinoma. In conclusion, our findings unravel the importance of CD4 T cells in Tim-3/Tim-3L-mediated immunosuppression and provide new thoughts for Tim-3 targeted cancer immunotherapy.