SUMMARY Adipose tissue is an essential regulator of metabolic homeostasis. In contrast with white adipose tissue, which stores excess energy in the form of triglycerides, brown adipose tissue is thermogenic, dissipating energy as heat via the unique expression of the mitochondrial uncoupling protein UCP1. A subset of UCP1+ adipocytes develop within white adipose tissue in response to physiological stimuli, however, the developmental origin of these “brite” or “beige” adipocytes is unclear. Here, we report the identification of a BMP7-ROCK signaling axis regulating beige adipocyte formation via control of the G-actin-regulated transcriptional coactivator myocardin related transcription factor A, MRTFA. White adipose tissue from MRTFA–/– mice contains more multilocular adipocytes and expresses enhanced levels of brown-selective proteins, including UCP1. MRTFA–/– mice also show improved metabolic profiles and protection from diet-induced obesity and insulin resistance. Our study hence unravels a central pathway driving the development of physiologically functional beige adipocytes.

No abstract

No abstract

BackgroundShark new antigen receptor variable domain (VNAR) antibodies can bind restricted epitopes that may be inaccessible to conventional antibodies.MethodsHere, we developed a library construction method based on polymerase chain reaction (PCR)-Extension Assembly and Self-Ligation (named “EASeL”) to construct a large VNAR antibody library with a size of 1.2 × 1010 from six naïve adult nurse sharks (Ginglymostoma cirratum).ResultsThe next-generation sequencing analysis of 1.19 million full-length VNARs revealed that this library is highly diversified because it covers all four classical VNAR types (Types I–IV) including 11% of classical Type I and 57% of classical Type II. About 30% of the total VNARs could not be categorized as any of the classical types. The high variability of complementarity determining region (CDR) 3 length and cysteine numbers are important for the diversity of VNARs. To validate the use of the shark VNAR library for antibody discovery, we isolated a panel of VNAR phage binders to cancer therapy-related antigens, including glypican-3, human epidermal growth factor receptor 2 (HER2), and programmed cell death-1 (PD1). Additionally, we identified binders to viral antigens that included the Middle East respiratory syndrome (MERS) and severe acute respiratory syndrome (SARS) spike proteins. The isolated shark single-domain antibodies including Type I and Type II VNARs were produced in Escherichia coli and validated for their antigen binding. A Type II VNAR (PE38-B6) has a high affinity (Kd = 10.1 nM) for its antigen.ConclusionsThe naïve nurse shark VNAR library is a useful source for isolating single-domain antibodies to a wide range of antigens. The EASeL method may be applicable to the construction of other large diversity gene expression libraries.

Polygonatum cyrtonema lectin (PCL), a mannose/sialic acid-binding lectin, has been reported to display remarkable inhibitory and cytotoxic activity toward cancer cells. However, the precise mechanism by which PCL induces tumor cell death is still only rudimentarily understood. In the present study, PCL was shown to markedly inhibit the growth of human melanoma A375 cells with concomitant low toxicity to the normal melanocytes. Subsequently, PCL was found to simultaneously induce A375 cell apoptosis and autophagy. The mechanism of apoptosis following treatment with PCL involved regulation of Bax, Bcl-x(L) and Bcl-2 proteins, which then caused collapse of the mitochondrial membrane potential, leading to cytochrome c release and caspase activation. The treatment with PCL also abrogated the glutathione antioxidant system, and induced mitochondria to generate massive ROS accumulation, which subsequently resulted in p38 and p53 activation. Further experimental data confirmed that the ROS-p38-p53 pathway could be involved in the stimulation of autophagy, suggesting that autophagy may play a death-promoting role via the above-mentioned apoptotic pathway. In conclusion, these findings indicate that PCL induces both apoptosis and autophagy in cancer cells through a mitochondria-mediated ROS-p38-p53 pathway.

No abstract

No abstract

Growing evidence suggests that a physiological activity of the cellular prion protein (PrPC) plays a crucial role in several neurodegenerative disorders, including prion and Alzheimer’s diseases. However, how the functional activity of PrPC is subverted to deliver neurotoxic signals remains uncertain. Transgenic mice expressing PrP with a deletion of residues 105–125 in the central region (referred to as ΔCR PrP) provide important insights into this problem. Tg(ΔCR) mice exhibit neonatal lethality and massive degeneration of cerebellar granule neurons, a phenotype that is dose-dependently suppressed by the presence of wild-type PrP. When expressed in cultured cells, ΔCR PrP induces large, ionic currents that can be detected by patch-clamping techniques. Here, we have tested the hypothesis that abnormal ion channel activity underlies the neuronal death seen in Tg(ΔCR) mice. We find that ΔCR PrP induces abnormal ionic currents in neurons in culture and in cerebellar slices, and that this activity sensitizes the neurons to glutamate-induced, calcium-mediated death. In combination with ultrastructural and biochemical analyses, these results demonstrate a role for glutamate-induced excitotoxicity in PrP-mediated neurodegeneration. A similar mechanism may operate in other neurodegenerative disorders due to toxic, β-rich oligomers that bind to PrPC.