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Euphorbia kansui, a toxic Chinese medicine used for more than 2000 years, has the effect of “purging water to promote drinking” and “reducing swelling and dispersing modules”. Diterpenes and triterpenes are the main bioactive components of E. kansui. Among them, ingenane-type diterpenes have multiple biological activities as a protein kinase C δ (PKC-δ) activator, which have previously been shown to promote anti-proliferative and pro-apoptotic effects in several human cancer cell lines. However, the activation of PKC subsequently promoted the survival of macrophages. Recently, we found that 13-hydroxyingenol-3-(2,3-dimethylbutanoate)-13-dodecanoate (compound A) from E. kansui showed dual bioactivity, including the inhibition of tumor-cell-line proliferation and regulation of macrophage polarization. This study identifies the possible mechanism of compound A in regulating the polarization state of macrophages, by regulating PKC-δ-extracellular signal regulated kinases (ERK) signaling pathways to exert anti-tumor immunity effects in vitro, which might provide a new treatment method from the perspective of immune cell regulation.

Although it is well recognized that mycosporine-like amino acids (MAAs) are ultraviolet (UV) protective agents that can reduce UV damage, the specific biological mechanism of its role in the skin remains unclear. In this study, we investigated the effect of MAAs extracted from Antarctic diatom Phaeodactylum tricornutum ICE-H on UVB-induced skin damage using a mice model. The MAAs components identified by liquid chromatography-tandem mass spectrometry included 4-deoxygadusol, shinorine, and porphyra-334, which were purified using a Supledean Carboxen1000 solid phase extraction column. The antioxidant activities of these MAA compounds were tested in vitro. For UVB-induced skin photodamage in mice, MAAs alleviated skin swelling and epidermal thickening in this study. We detected the content of reactive oxygen species (ROS), malondialdehyde, and collagen in skin tissue. In addition, quantitative real-time polymerase chain reaction was used to detect nuclear factor-κB (NF-κB), tumor necrosis factor α, interleukin-1β, cyclooxygenase-2, mitogen activated protein kinase (MAPK) family (extracellular signal-regulated kinase, c-Jun amino-terminal kinase, and p38 kinase), and matrix metalloproteinases. The expression of these cytokines and enzymes is related to inflammatory responses and collagen degradation. In comparison to the model group without MAA treatment, the MAA component decreased the concentration of ROS, the degree of oxidative stress in the skin tissue, and the expression of genes involved in the NF-κB and MAPK pathways. In summary, these MAA components extracted from Phaeodactylum tricornutum ICE-H protected against UVB-induced skin damage by inhibiting ROS generation, relieving skin inflammation, and slowing down collagen degradation, suggesting that these MAA components are effective cosmetic candidate molecules for the protection and therapy of UVB damage.

MORN proteins play a key role in the cytoskeletal structure of eukaryotes and are essential for the close arrangement of the endoplasmic reticulum and plasma membrane. A gene with nine MORN motifs (TGGT1_292120, named TgMORN2) was identified in the Toxoplasma gondii genome; it was presumed to belong to the MORN protein family and to have the function of forming the cytoskeleton, which affects the survival of T. gondii. However, the genetic deletion of MORN2 did not noticeably affect parasite growth and virulence. Using adjacent protein labeling techniques, we identified a network of TgMORN2 interactions, which mainly included endoplasmic reticulum stress (ER stress)-related proteins. In exploring these data, we found that the pathogenicity of the KO-TgMORN2 strain was significantly reduced in the case of tunicamycin-induced ER stress. Reticulon TgRTN (TGGT1_226430) and tubulin β-Tubulin were identified as interaction proteins of TgMORN2. Collectively, TgMORN2 plays a role in ER stress, which lays a foundation for further research on the function of the MORN protein in T. gondii.

Improvement of low nitrogen (LN) tolerance or nitrogen use efficiency (NUE) in crops is imperative for environment-friendly agriculture development. The basic helix-loop-helix (bHLH) transcription factors are involved in multiple abiotic stresses and are suitable as candidate genes for improving LN tolerance. Few studies were performed on the characterization of the HvbHLH gene family and their function in response to LN stress in barley. In this study, 103 HvbHLH genes were identified through genome-wide analysis. HvbHLH proteins were classified into 20 subfamilies based on phylogenetic analysis in barley, which was supported by conserved motifs and gene structure analysis. The stress-related cis-element analysis in the promoters showed that HvbHLHs are probably involved in multiple stress responses. By phylogenetic analysis of HvbHLHs and bHLHs in other plants, some HvbHLHs were predicted to play roles in response to nutrition deficiency stress. Furthermore, at least 16 HvbHLHs were differentially expressed in two barley genotypes differing in LN tolerance under LN stress. Finally, overexpression of HvbHLH56 enhanced LN stress tolerance in transgenic Arabidopsis, suggesting it is an important regulator in LN stress response. The differentially expressed HvbHLHs identified herein may be valuable for the breeding of barley cultivars with LN tolerance.

We had shown in our previous study that TgUrm1 (ubiquitin-related Modifier 1) was involved in the regulation of anti-oxidant stress in Toxoplasma gondii by conjugating with TgAhp1. It is generally believed that Urm1 binds to target proteins through a mechanism involving Uba (ubiquitin-like activator protein). Here, we identified the TgUrm1-exclusive ubiquitin-like activator-TgUba1, which was located in the cytoplasm of Toxoplasma. TgUba1 contained three domains, including the atrophin-1 domain (ANT1), the E1-like domain (AD), and the rhodanese homology domain (RHD). We explored the interaction of TgUba1 with TgUrm1, and the AD domain was essential for the interaction of the two proteins. The TgUba1 knockout and complementary mutants were obtained based on CRISPR/Cas9 gene editing technology. The knockout of TgUba1 attenuated parasite proliferation and virulence in mice, but not invasion and egress processes, revealing the pivotal role played by TgUba1 in T. gondii survival. Meanwhile, the conjugate band of TgUrm1 was significantly reduced under oxidative stress stimulation without TgUba1, indicating that TgUba1 enhanced the targeted conjugation ability of TgUrm1 in response to oxidative stress, especially under diamide (Dia) stimulation. Furthermore, eleven TgUba1-interacting proteins were identified by proximity-based protein labeling techniques, relating them to ubiquitin-like modifications, anti-oxidative stress and metabolic regulation processes. In conclusion, TgUba1 was essential for T. gondii survival and might be a potential ubiquitin-like activator protein for TgUrm1.

Dexmedetomidine (DEX), a selective α2 adrenergic receptor (AR) agonist, is commonly used as a sedative drug during critical illness. In the present study, we explored a novel accelerative effect of DEX on cardiac fibroblast (CF) differentiation mediated by LPS and clarified its potential mechanism. LPS apparently increased the expression of α-SMA and collagen I/III and the phosphorylation of p38 and Smad-3 in the CFs of mice. These effects were significantly enhanced by DEX through increasing α2A-AR expression in CFs after LPS stimulation. The CFs from α2A-AR knockout mice were markedly less sensitive to DEX treatment than those of wild-type mice. Inhibition of protein kinase C (PKC) abolished the enhanced effects of DEX on LPS-induced differentiation of CFs. We also found that the α-SMA level in the second-passage CFs was much higher than that in the nonpassage and first-passage CFs. However, after LPS stimulation, the TNF-α released from the nonpassage CFs was much higher than that in the first- and second-passage CFs. DEX had no effect on LPS-induced release of TNF-α and IL-6 from CFs. Further investigation indicated that DEX promoted cardiac fibrosis and collagen I/III synthesis in mice exposed to LPS for four weeks. Our results demonstrated that DEX effectively accelerated LPS-induced differentiation of CFs to myofibroblasts through the PKC-p38-Smad2/3 signaling pathway by activating α2A-AR.

Aspartate kinase (AK) is the key enzyme in the biosynthesis of aspartate-derived amino acids. Recombinant AK was efficiently purified and systematically characterized through analysis under optimal conditions combined with steady-state kinetics study. Homogeneous AK was predicted as a decamer with a molecular weight of ~48 kDa and a half-life of 4.5 h. The enzymatic activity was enhanced by ethanol and Ni2+. Moreover, steady-state kinetic study confirmed that AK is an allosteric enzyme, and its activity was inhibited by allosteric inhibitors, such as Lys, Met, and Thr. Theoretical results indicated the binding mode of AK and showed that Arg169 is an important residue in substrate binding, catalytic domain, and inhibitor binding. The values of the kinetic parameter Vmax of R169 mutants, namely, R169Y, R169P, R169D, and R169H AK, with l-aspartate as the substrate, were 4.71-, 2.25-, 2.57-, and 2.13-fold higher, respectively, than that of the wild-type AK. Furthermore, experimental and theoretical data showed that Arg169 formed a hydrogen bond with Glu92, which functions as the entrance gate. This study provides a basis to develop new enzymes and elucidate the corresponding amino acid production.

The growth hormone receptor (GHR) gene is correlated with many phenotypic and physiological alternations in chicken, such as shorter shanks, lower body weight and muscle mass loss. However, the role of the GHR gene in mitochondrial function remains unknown in poultry. In this study, we assessed the function of mitochondria in sex-linked dwarf (SLD) chicken skeletal muscle and interfered with the expression of GHR in DF-1 cells to investigate the role of the GHR gene in chicken mitochondrial function both in vivo and in vitro. We found that the expression of key regulators of mitochondrial biogenesis and mitochondrial DNA (mtDNA)-encoded oxidative phosphorylation (OXPHOS) genes were downregulated and accompanied by reduced enzymatic activity of OXPHOS complexes in SLD chicken skeletal muscle and GHR knockdown cells. Then, we assessed mitochondrial function by measuring mitochondrial membrane potential (ΔΨm), mitochondrial swelling, reactive oxygen species (ROS) production, malondialdehyde (MDA) levels, ATP levels and the mitochondrial respiratory control ratio (RCR), and found that mitochondrial function was impaired in SLD chicken skeletal muscle and GHR knockdown cells. In addition, we also studied the morphology and structure of mitochondria in GHR knockdown cells by transmission electron microscopy (TEM) and MitoTracker staining. We found that knockdown of GHR could reduce mitochondrial number and alter mitochondrial structure in DF-1 cells. Above all, we demonstrated for the first time that the GHR gene is essential for chicken mitochondrial function in vivo and in vitro.

As the most common cause of cancer death in women, the pathogenesis of breast cancer still remains unclear. Here, we reported a long non-coding RNA (lncRNA), HOTTIP (HOXA transcript at the distal tip), that may play an important role in the pathogenesis of breast cancer. Using gain-and-loss-of experiments in vitro and in vivo, we observed the marked upregulation of HOTTIP/HOXA11 in the breast cancer cell line, MCF-7, and the downregulation of HOTTIP or HOXA11, which might inhibit cell proliferation and migration but promote cell apoptosis in breast cancer MCF-7 cells. In addition, by further rescue experiments with HOXA11 overexpression, we uncovered a novel potential regulatory mechanism between HOTTIP and one of its physical HOXA clusters, HOXA11. Hence, HOTTIP may mediate, at least partly, HOXA11 expression involved in cell growth, migration, and apoptosis of breast cancer MCF-7 cells.