GAP-43，Netrin-1，Collapsin-1和Neuropilin-1在出生后小鼠小脑中的动态表达

GAP-43,netrin-1,collapsin-1和neuropilin-1被认为在成网络分布的神经联系中发挥重要的作用．在年幼的啮齿类动物中,小脑包含5种不同的集中分布层：白质、内颗粒细胞层(IGL)、浦肯野氏细胞层(PCL)、分子层(ML)和外颗粒细胞层(EGL)．与浦肯野氏神经元在出生前产生这一点不同的是,EGL中的细胞在出生后产生,它们接受从前脑olivary核团发出的攀援纤维的主要神经投射,以及从内颗粒细胞发出的平行纤维的神经投射．这些神经投射主要在出生后的前3个星期内建立,同时还有浦肯野氏细胞的发育和成熟．而GAP-43,netrin-1,collapsin-1和neuropilin-1在出生后小脑发育的潜在作用仍然不清楚．为了更加清楚地探讨上述问题,检验了GAP-43,netrin-1,collapsin-1和neuropilin-1的mRNA与蛋白质在出生后5,10,20天和成年小鼠小脑中的表达情况．研究结果显示,这4种分子在小鼠出生后的小脑中有不同的时间和空间表达形式,这些结果与出生后发育和成年期间的轴突发生、延伸以及突触形成都有关联．通过免疫组织化学双标染色,发现小鼠出生后10天的小脑中,GAP-43阳性的浦肯野氏细胞也显示netrin-1或collapsin-1阳性,并且collapsin-1阳性的细胞也对 netrin-1 阳性．上述研究结果证明这4种分子可能参与了小脑的出生后发育．     

E3 ubiquitin ligases in regulating stress fiber,lamellipodium, and focal adhesion dynamics

Recent discoveries have unveiled the roles of a complicated network of E3 ubiquitin ligases in regulating cell migration machineries. The E3 ubiquitin ligases Smurf1 and Cul/BACURD ubiquitinate RhoA to regulate stress fiber formation and cell polarity, and ASB2α ubiquitinates filamins to modulate cytoskeletal stiffness, thus regulating cell spreading and cell migration. HACE1, XIAP, and Skp1-Cul1-F-box bind to Rac1 and cause its ubiquitination and degradation, thus suppressing lamellipodium protrusions, while PIAS3, a SUMO ligase, activates Rac1 to promote lamellipodium dynamics. Smurf1 also enhances Rac1 activation but it does not ubiquitinate Rac1. Both Smurf1 and HECTD1 regulate focal adhesion (FA) assembly and (or) disassembly through ubiquitinating the talin head domain and phosphatidylinositol 4 phosphate 5-kinase type I γ (PIPKIγ90), respectively. Thus, E3 ubiquitin ligases regulate stress fiber formation, cell polarity, lamellipodium protrusions, and FA dynamics through ubiquitinating the key proteins that control these processes.     

Nod1, a CARD protein,enhances pro-interleukin-1beta processing through the interaction with pro-caspase-1

The production of bioactive interleukin-1beta (IL-1beta), a pro-inflammatory cytokine, is mediated by activated caspase-1. One of the known molecular mechanisms underlying pro-caspase-1 processing and activation involves interaction between the caspase recruit domains (CARDs) of caspase-1 and a serine/threonine kinase RIP2. While the association of Nod1 with both caspase-1 and RIP2 is already known, the consequences of these interactions are poorly understood. Because Nod1 also binds to RIP2, we hypothesized that Nod1 plays a role in pro-caspase-1 activation and IL-1beta processing. We show here that Nod1 binds to both RIP2 and caspase-1 by CARD interactions. Nod1 enhances pro-caspase-1 oligomerization and pro-caspase-1 processing. Nod1 enhances caspase-1-induced IL-1beta secretion, as well as lipopolysaccharide (LPS)-induced IL-1beta secretion in transfected cells. Moreover, HT1080 cells stably transfected with Nod1 showed higher LPS-induced IL-1beta secretion than non-transfected cells, suggesting a role of Nod1 in LPS-induced responses. Our data indicate that Nod1 can regulate IL-1beta secretion, implying that Nod1 may play a role in inflammatory responses to bacterial LPS.     

DISC1, PDE4B, and NDE1 at the centrosome and synapse

Disrupted-In-Schizophrenia 1 (DISC1) is a risk factor for schizophrenia and other major mental illnesses. Its protein binding partners include the Nuclear Distribution Factor E Homologs (NDE1 and NDEL1), LIS1, and phosphodiesterases 4B and 4D (PDE4B and PDE4D). We demonstrate that NDE1, NDEL1 and LIS1, together with their binding partner dynein, associate with DISC1, PDE4B and PDE4D within the cell, and provide evidence that this complex is present at the centrosome. LIS1 and NDEL1 have been previously suggested to be synaptic, and we now demonstrate localisation of DISC1, NDE1, and PDE4B at synapses in cultured neurons. NDE1 is phosphorylated by cAMP-dependant Protein Kinase A (PKA), whose activity is, in turn, regulated by the cAMP hydrolysis activity of phosphodiesterases, including PDE4. We propose that DISC1 acts as an assembly scaffold for all of these proteins and that the NDE1/NDEL1/LIS1/dynein complex is modulated by cAMP levels via PKA and PDE4.     

Ligand Perception,Activation, and Early Signaling of Plant Steroid Receptor Brassinosteroid Insensitive

Leucine-rich repeat receptor-like kinases （LRR-RLKs） belong to a large group of cell surface proteins involved in many aspects of plant development and environmental responses in both monocots and dicots. Brassinosteroid insensitive 1 （BRI1）, a member of the LRR X subfamily, was first identified through several forward genetic screenings for mutants insensitive to brassinosteroids （BRs）, which are a class of plant-specific steroid hormones. Since its identification, BRI1 and its homologs had been proved as receptors perceiving BRs and initiating BR signaling. The co-receptor BRIl-associated kinase 1 and its homologs, and other BRI1 interacting proteins such as its inhibitor BRI1 kinase inhibitor I （BKI1） were identified by genetic andbiochemical approaches. The detailed mechanisms of BR perception by BRI1 and the activation of BRI1 receptor complex have also been elucidated. Moreover, several mechanisms for termination of the activated BRI1 signaling were also discovered. In this review, we will focus on the recent advances on the mechanism of BRI1 phosphorylation and activation, the regulation of its receptor complex, the structure basis of BRI1 ectodomain and BR recognition, its direct substrates, and the termination of the activated BRI1 receptor complex.     

Conservation of a chemokine system, XCR1 and its ligand, XCL1, between human and mice

Understanding dendritic cell (DC) subset functions should lead to the development of novel types of vaccine. Here we characterized expression of XC chemokine receptor 1 (XCR1) and its ligand, XCL1. Murine XCR1 was the only chemokine receptor selectively expressed in CD8α⁺ conventional DCs. XCL1 was constitutively expressed in NK cells, which contribute to serum XCL1 levels. NK and CD8⁺ T cells increased XCL1 production upon activation. These expression patterns were conserved in human blood cells, including the BDCA3⁺ DC subset. Thus, in human and mice, certain DC subsets should be chemotactic towards NK or activated CD8⁺ T cells through XCR1.     

HIFs,angiogenesis, and cancer

Tumor hypoxia was first described in the 1950s by radiation oncologists as a frequent cause of failure to radiotherapy in solid tumors. Today, it is evident that tumor hypoxia is a common feature of many cancers and the master regulator of hypoxia, hypoxia‐inducible factor‐1 (HIF‐1), regulates multiple aspects of tumorigenesis, including angiogenesis, proliferation, metabolism, metastasis, differentiation, and response to radiation therapy. Although the tumor hypoxia response mechanism leads to a multitude of downstream effects, it is angiogenesis that is most crucial and also most susceptible to molecular manipulation. The delineation of molecular mechanisms of angiogenesis has revealed a critical role for HIF‐1 in the regulation of angiogenic growth factors. In this article, we review what has been described about HIF‐1: its structure, its regulation, and its implication for cancer therapy and we focus on its role in angiogenesis and cancer. J. Cell. Biochem. 114: 967–974, 2013. © 2012 Wiley Periodicals, Inc.     

A comparison of aflatoxin B1-induced cytotoxicity,mutagenicity and prophage induction in Salmonella typhimurium mutagen tester strains TA1535, TA1538, TA98 and TA100

Treatment of Ames mutagen tester strains with aflatoxin B1 (AFB1) and S9 mix results not only in the production of a poten mutagen, but induces a pathway that leads to the induction of prophages present in all Ames tester strains.Characterization of the prophage induction and mutagenic response following AFB1 treatment showed that plasmid pKM101 dramatically enhances mutagenesis, but suppressed prophage induction. Spontaneous release of phage by TA98 and TA100 was also lower than in TA1535 and TA1538.In addition to mutagenesis and prophage induction, survival of all 4 tester strains was quantitated after AFB1 treatment. The data show that the frameshift tester strains (TA1538 and TA98) are more sensitive to the bactericidal action of AFB1 than the base-pair tester strains (TA1535 and TA100), survival being significantly affected above 100 ng. One of several hypotheses examined was the difference in the number and types of prophages present in base-pair tester strains that are not detectable in the frame-shift tester strains.These data suggest that prophage induction can detect DNA damage that is non-mutagenic; and that it is important to characterize the lysogenic nature of the Ames strains since it may influence the observed histidine revertant rate and the survival of the tester strain.     

Interaction of myelin basic protein, melittin and bovine serum albumin with gangliosides, sulphatide and neutral glycosphingolipids in mixed monolayers

Some parameters that may regulate the miscibility and stability of mixed lipid-protein monolayers at the air-145 mM NaCl interface were studied employing six glycosphingolipids (acidic or neutral), three different types of proteins (soluble, extrinsic or highly amphipathic) and some phospholipids. The results obtained show that the percentage of the total area occupied by the protein at the interface is an important parameter leading to lateral phase separations; the amount and area contribution of the protein accepted in the film before the components become immiscible increase with the complexity of the polar head group of the glycosphingolipids. The interactions occur with progressive reductions of the intermolecular packing as the polar head group of the glycosphingolipid becomes more complex and this is accompanied by more negative values of the excess free energy of mixing. The lipid component seems to be the major responsible for the reduction in mean molecular area.     

Death domain complex of the TNFR-1, TRADD,and RIP1 proteins for death-inducing signaling

Apoptosis can be induced by an extrinsic pathway involving the ligand-mediated activation of death receptors such as tumor necrosis factor receptor-1 (TNFR-1). TNFR-1-associated death domain (TRADD) protein is an adapter molecule that bridges the interaction between TNFR-1 and receptor-interacting serine/threonine-protein kinase 1 (RIP1). However, the molecular mechanism of the complex formation of these proteins has not yet been identified. Here, the binding among TNFR-1, TRADD, and RIP1 was identified using a GST pull-down assay and Biacore biosensor experiment. This study showed that structural characterization and formation of the death-signaling complex could be predicted using TNFR-1, TRADD, and RIP1. In addition, we found that the structure-based mutations of TNFR-1 (P367A and P368A), TRADD (F266A), and RIP1 (M637A and R638A) disrupted formation of the death domain (DD) complex and prevented stable interactions among those DDs.     

Neuraminidase 1 deficiency attenuates cardiac dysfunction,oxidative stress,fibrosis, inflammatory via AMPK-SIRT3 pathway in diabetic cardiomyopathy mice

Diabetic cardiomyopathy (DCM) is associated with oxidative stress and augmented inflammation in the heart. Neuraminidases (NEU) 1 has initially been described as a lysosomal protein which plays a role in the catabolism of glycosylated proteins. We investigated the role of NEU1 in the myocardium in diabetic heart. Streptozotocin (STZ) was injected intraperitoneally to induce diabetes in mice. Neonatal rat ventricular myocytes (NRVMs) were used to verify the effect of shNEU1 in vitro. NEU1 is up-regulated in cardiomyocytes under diabetic conditions. NEU1 inhibition alleviated oxidative stress, inflammation and apoptosis, and improved cardiac function in STZ-induced diabetic mice. Furthermore, NEU1 inhibition also attenuated the high glucose-induced increased reactive oxygen species generation, inflammation and, cell death in vitro. ShNEU1 activated Sirtuin 3 (SIRT3) signaling pathway, and SIRT3 deficiency blocked shNEU1-mediated cardioprotective effects in vitro. More importantly, we found AMPKα was responsible for the elevation of SIRT3 expression via AMPKα-deficiency studies in vitro and in vivo. Knockdown of LKB1 reversed the effect elicited by shNEU1 in vitro. In conclusion, NEU1 inhibition activates AMPKα via LKB1, and subsequently activates sirt3, thereby regulating fibrosis, inflammation, apoptosis and oxidative stress in diabetic myocardial tissue.     

Preclinical pharmacokinetics,pharmacodynamics, tissue distribution,and tumor penetration of anti-PD-L1 monoclonal antibody,an immune checkpoint inhibitor

MPDL3280A is a human monoclonal antibody that targets programmed cell death-1 ligand 1 (PD-L1), and exerts anti-tumor activity mainly by blocking PD-L1 interaction with programmed cell death-1 (PD-1) and B7.1. It is being investigated as a potential therapy for locally advanced or metastatic malignancies. The purpose of the study reported here was to characterize the pharmacokinetics, pharmacodynamics, tissue distribution and tumor penetration of MPDL3280A and/or a chimeric anti-PD-L1 antibody PRO304397 to help further clinical development.

The pharmacokinetics of MPDL3280A in monkeys at 0.5, 5 and 20 mg·kg^?1 and the pharmacokinetics / pharmacodynamics of PRO304397 in mice at 1, 3 10 mg·kg^?1 were determined after a single intravenous dose. Tissue distribution and tumor penetration for radiolabeled PRO304397 in tumor-bearing mouse models were determined.

The pharmacokinetics of MPDL3280A and PRO304397 were nonlinear in monkeys and mice, respectively. Complete saturation of PD-L1 in blood in mice was achieved at serum concentrations of PRO304397 above ～0.5 µg·mL^?1. Tissue distribution and tumor penetration studies of PRO304397 in tumor-bearing mice indicated that the minimum tumor interstitial to plasma radioactivity ratio was ～0.3; saturation of target-mediated uptake in non–tumor tissues and desirable exposure in tumors were achieved at higher serum concentrations, and the distribution into tumors was dose-and time-dependent.

The biodistribution data indicated that the efficacious dose is mostly likely higher than that estimated based on simple pharmacokinetics/pharmacodynamics in blood. These data also allowed for estimation of the target clinical dose for further development of MPDL3280A.