共查询到20条相似文献,搜索用时 0 毫秒
1.
Antiangiogenic activity of restin, NC10 domain of human collagen XV: comparison to endostatin 总被引:13,自引:0,他引:13
Ramchandran R Dhanabal M Volk R Waterman MJ Segal M Lu H Knebelmann B Sukhatme VP 《Biochemical and biophysical research communications》1999,255(3):735-739
Based on a homology search with endostatin, the C-terminus 185 aa of collagen XVIII, we report the cloning, expression, and antiangiogenic activity of a 22 kDa human collagen XV fragment, that we have named restin. Restin was expressed in the prokaryotic pET expression system. We have shown that restin inhibits the migration of endothelial cells in vitro but has no effect on the proliferation of these cells. A polyclonal antibody raised against endostatin cross-reacted with restin. Systemic administration of restin suppressed the growth of tumors in a xenograft renal carcinoma model. 相似文献
2.
ángel M Cuesta David Sánchez-Martín Ana Blanco-Toribio Maider Villate Kelly Enciso-álvarez Ana Alvarez-Cienfuegos Noelia Sainz-Pastor Laura Sanz Francisco J Blanco Luis álvarez-Vallina 《MABS-AUSTIN》2012,4(2):226-232
We recently described the in vitro and in vivo properties of an engineered homotrimeric antibody made by fusing the N-terminal trimerization region of collagen XVIII NC1 domain to the C-terminus of a scFv fragment [trimerbody (scFv-NC1)3; 110 kDa]. Here, we demonstrated the utility of the N-terminal trimerization region of collagen XV NC1 domain in the engineering of trivalent antibodies. We constructed several scFv-based trimerbodies containing the human type XV trimerization domain and demonstrated that all the purified trimerbodies were trimeric in solution and exhibited excellent antigen binding capacity. Importantly, type XV trimerbodies demonstrated substantially greater thermal and serum stability and resistance to protease digestion than type XVIII trimerbodies. In summary, the small size, high expression level, solubility and stability of the trimerization domain of type XV collagen make it the ideal choice for engineering homotrimeric antibodies for cancer detection and therapy.Key words: antibody engineering, multivalent antibody, collagen XVIII, collagen XV, tumor targeting 相似文献
3.
《MABS-AUSTIN》2013,5(2):226-232
We recently described the in vitro and in vivo properties of an engineered homotrimeric antibody made by fusing the N-terminal trimerization region of collagen XVIII NC1 domain to the C-terminus of a scFv fragment [trimerbody (scFv-NC1)3; 110 kDa]. Here, we demonstrated the utility of the N-terminal trimerization region of collagen XV NC1 domain in the engineering of trivalent antibodies. We constructed several scFv-based trimerbodies containing the human type XV trimerization domain and demonstrated that all the purified trimerbodies were trimeric in solution and exhibited excellent antigen binding capacity. Importantly, type XV trimerbodies demonstrated substantially greater thermal and serum stability and resistance to protease digestion than type XVIII trimerbodies. In summary, the small size, high expression level, solubility and stability of the trimerization domain of type XV collagen make it the ideal choice for engineering homotrimeric antibodies for cancer detection and therapy. 相似文献
4.
Lolkema MP Gervais ML Snijckers CM Hill RP Giles RH Voest EE Ohh M 《The Journal of biological chemistry》2005,280(23):22205-22211
The tumor suppressor function of the von Hippel-Lindau protein (pVHL) has previously been linked to its role in regulating hypoxia-inducible factor levels. However, VHL gene mutations suggest a hypoxia-inducible factor-independent function for the N-terminal acidic domain in tumor suppression. Here, we report that phosphorylation of the N-terminal acidic domain of pVHL by casein kinase-2 is essential for its tumor suppressor function. This post-translational modification did not affect the levels of hypoxia-inducible factor; however, it did change the binding of pVHL to another known binding partner, fibronectin. Cells expressing phospho-defective mutants caused improper fibronectin matrix deposition and demonstrated retarded tumor formation in mice. We propose that phosphorylation of the acidic domain plays a role in the regulation of proper fibronectin matrix deposition and that this may be relevant for the development of VHL-associated malignancies. 相似文献
5.
《Cell cycle (Georgetown, Tex.)》2013,12(17):2836-2835
Comment on: Ferrer I, et al. Cell Cycle 2011; 10:1948-55. 相似文献
6.
Nuclear PTEN-mediated growth suppression is independent of Akt down-regulation 总被引:12,自引:0,他引:12 下载免费PDF全文
Liu JL Sheng X Hortobagyi ZK Mao Z Gallick GE Yung WK 《Molecular and cellular biology》2005,25(14):6211-6224
The tumor suppressor gene PTEN is a phosphoinositide phosphatase that is inactivated by deletion and/or mutation in diverse human tumors. Wild-type PTEN is expressed both in the cytoplasm and nucleus in normal cells, with a preferential nuclear localization in differentiated or resting cells. To elucidate the relationship between PTEN's subcellular localization and its biologic activities, we constructed different PTEN mutants that targeted PTEN protein into different subcellular compartments. Our data show that the subcellular localization patterns of a PTEN (deltaPDZB) mutant versus a G129R phosphatase mutant were indistinguishable from those of wild-type PTEN. In contrast, the Myr-PTEN mutant demonstrated an enhanced association with the cell membrane. We found that nuclear PTEN alone is capable of suppressing anchorage-independent growth and facilitating G1 arrest in U251MG cells without inhibiting Akt activity. Nuclear compartment-specific PTEN-induced growth suppression is dependent on possessing a functional lipid phosphatase domain. In addition, the down-regulation of p70S6K could be mediated, at least in part, through activation of AMP-activated protein kinase in an Akt-independent fashion. Introduction of a constitutively active mutant of Akt, Akt-DD, only partially rescues nuclear PTEN-mediated growth suppression. Our collective results provide the first direct evidence that PTEN can contribute to G1 growth arrest through an Akt-independent signaling pathway. 相似文献
7.
Y Ning J W Shay M Lovell L Taylor D H Ledbetter O M Pereira-Smith 《Experimental cell research》1991,192(1):220-226
Previous hybrid studies involving fusion of normal with immortal human cells indicated that the phenotype of cellular senescence is dominant and that immortality results from recessive changes in normal growth regulatory genes. We have further assigned 28 different immortal human cell lines to at least four complementation groups for indefinite division. In order to identify the chromosomes involved in regulating cell proliferation, we have introduced single human chromosomes by microcell fusion into immortal human cells representative of the different complementation groups. Our results demonstrate that the introduction of chromosome 11, implicated in tumor suppression, does not cause cellular senescence in three different immortal human cell lines tested. 相似文献
8.
Jacqueline A. Wirz Sergei P. Boudko Thomas F. LerchMichael S. Chapman Hans Peter Bächinger 《Matrix biology》2011,30(1):9-15
Correct folding of the collagen triple helix requires a self-association step which selects and binds α-chains into trimers. Here we report the crystal structure of the trimerization domain of human type XV collagen. The trimerization domain of type XV collagen contains three monomers each composed of four β-sheets and an α-helix. The hydrophobic core of the trimer is devoid of solvent molecules and is shaped by β-sheet planes from each monomer. The trimerization domain is extremely stable and forms at picomolar concentrations. It is found that the trimerization domain of type XV collagen is structurally similar to that of type XVIII, despite only 32% sequence identity. High structural conservation indicates that the multiplexin trimerization domain represents a three dimensional fold that allows for sequence variability while retaining structural integrity necessary for tight and efficient trimerization. 相似文献
9.
Henrik Wegener Sarah Leineweber Karsten Seeger 《Biochemical and biophysical research communications》2013,430(2):449-453
Type VII collagen (Col7) is the major component of anchoring fibrils and very important for skin integrity. This is emphasized by the Col7 related skin blistering diseases dystrophic epidermolysis bullosa and epidermolysis bullosa acquisita. Structural data that provides insights into the interaction network of Col7 and thus providing a basis for a better understanding of the pathogenesis of the diseases is missing.We proved that the von-Willebrand-factor A like domain 2 (vWFA2) of Col7 is responsible for type I collagen binding. The interaction has a KD value of 90 μM as determined by SPR and is enthalpy driven as derived from the van’t Hoff equation. Furthermore, a hitherto unknown interaction of this domain with type IV collagen was identified. The interaction of vWFA2 with type I collagen is sensitive to the presence of magnesium ions, however, vWFA2 does not contain a magnesium binding site thus magnesium must bind to type I collagen.A lysine residue has been identified to be crucial for type I collagen binding. This allowed localization of the binding site. Mutational analysis suggests different interaction mechanisms in different species and that these interactions might be of covalent nature. 相似文献
10.
Exton MS Elfers A Jeong WY Bull DF Westermann J Schedlowski M 《American journal of physiology. Regulatory, integrative and comparative physiology》2000,279(4):R1310-R1315
The present study investigated the role of sympathetic innervation of the spleen in conditioned suppression of a contact hypersensitivity (CHS) reaction. Behavioral conditioning was achieved by pairing saccharin drinking solution (conditioned stimulus, CS) with injection of cyclosporin A (CsA, 20 mg/kg; unconditioned stimulus, UCS). Four days after sensitization of the animals by application of a 5% 2,4-dinitrochlorobenzene (DNCB) to abdominal skin, the animals were challenged by applying a 1% DNCB solution to the ear. The CHS response was monitored by measuring the degree of ear swelling. Saccharin re-presentation reduced ear swelling to a magnitude that approached that achieved by CsA treatment. Histological examination demonstrated that the conditioned reduction of ear swelling was produced by a reduced leukocyte infiltration of the ear. Prior sympathetic denervation of the spleen did not alter the conditioned suppression of the CHS response. These data indicate that behavioral conditioning using CsA produces alterations of CHS that, unlike conditioned prolongation of heart allograft survival, are independent of sympathetically regulated conditioned alterations in the spleen. 相似文献
11.
Peter S Amenta Nicole A Scivoletti Marissa D Newman Justin P Sciancalepore Deqin Li Jeanne C Myers 《The journal of histochemistry and cytochemistry》2005,53(2):165-176
Type XV is a large collagen-proteoglycan found in all human tissues examined. By light microscopy it was localized to most epithelial and all nerve, muscle, fat and endothelial basement membrane zones except for the glomerular capillaries or hepatic/splenic sinusoids. This widespread distribution suggested that type XV may be a discrete structural component that acts to adhere basement membrane to the underlying connective tissue. To address these issues, immunogold ultrastructural analysis of type XV collagen in human kidney, placenta, and colon was conducted. Surprisingly, type XV was found almost exclusively associated with the fibrillar collagen network in very close proximity to the basement membrane. Type XV exhibited a focal appearance directly on the surface of, or extending from, the fibers in a linear or clustered array. The most common single arrangement was a bridge of type XV gold particles linking thick-banded fibers. The function of type XV in this restricted microenvironment is expected to have an intrinsic dependence upon its modification with glycosaminoglycan chains. Present biochemical characterization showed that the type XV core protein in vivo carries chains of chondroitin/dermatan sulfate alone, or chondroitin/dermatan sulfate together with heparan sulfate in a differential ratio. Thus, type XV collagen may serve as a structural organizer to maintain a porous meshwork subjacent to the basement membrane, and in this domain may play a key role in signal transduction pathways. 相似文献
12.
The tricarboxylic acid cycle enzyme aconitase in yeast is a single translation product, which is dual targeted and distributed between the mitochondria and the cytosol by a unique mechanism involving reverse translocation. There is limited understanding regarding the precise mechanism of reverse translocation across the mitochondrial membranes. Here, we examined the contribution of the mature part of aconitase to its dual targeting. We created a set of aconitase mutants harboring two kinds of alterations: (1) point mutations or very small deletions in conserved sites and (2) systematic large deletions. These mutants were screened for their localization by a α-complementation assay, which revealed that the aconitase fourth domain that is at the C-terminus (amino acids 517-778) is required for aconitase distribution. Moreover, fusion of this C-terminal domain to mitochondria-targeted passenger proteins such as dihydrofolate reductase and orotidine-5′-phosphate decarboxylase, conferred dual localization on them. These results indicate that the aconitase C-terminal domain is both necessary and sufficient for dual targeting, thereby functioning as an “independent signal”. In addition, the same C-terminal domain was shown to be necessary for aconitase efficient posttranslational import into mitochondria. 相似文献
13.
Chaperone suppression of cellular toxicity of huntingtin is independent of polyglutamine aggregation 总被引:12,自引:0,他引:12
Polyglutamine protein aggregation is associated with eight inherited neurodegenerative disorders. In Huntington's disease, N-terminal fragments of mutant huntingtin form intracellular aggregates and mediate cellular toxicity. Recent studies have shown that chaperones inhibit polyglutamine-mediated aggregation and cellular toxicity. Because chaperones also inhibit caspase activation to protect cells from death, it remains unclear whether the protective effect of chaperones on polyglutamine-mediated cellular toxicity is dependent on their inhibition of protein aggregation. In this study, we show that several chaperones including HSP 40, HSP 70, and N-ethylmaleimide-sensitive factor can inhibit cellular toxicity caused by N-terminal mutant huntingtin fragments. However, only HSP 40 is able to inhibit huntingtin aggregation. Furthermore, time-course study suggests that the protection of chaperones against huntingtin toxicity is not the result of their suppression of huntingtin aggregation. Chaperones inhibit caspase-3 and caspase-9 activation mediated by mutant huntingtin, and this inhibition is independent of huntingtin aggregation. We propose that the inhibition of caspase activity by chaperones is involved in their suppression of polyglutamine toxicity. 相似文献
14.
Jillian N. Pearring Eric C. Lieu Joan R. Winter Sheila A. Baker Vadim Y. Arshavsky 《Molecular biology of the cell》2014,25(17):2644-2649
In vertebrate photoreceptor cells, rapid recovery from light excitation is dependent on the RGS9⋅Gβ5 GTPase-activating complex located in the light-sensitive outer segment organelle. RGS9⋅Gβ5 is tethered to the outer segment membranes by its membrane anchor, R9AP. Recent studies indicated that RGS9⋅Gβ5 possesses targeting information that excludes it from the outer segment and that this information is overridden by association with R9AP, which allows outer segment targeting of the entire complex. It was also proposed that R9AP itself does not contain specific targeting information and instead is delivered to the outer segment in the same post-Golgi vesicles as rhodopsin, because they are the most abundant transport vesicles in photoreceptor cells. In this study, we revisited this concept by analyzing R9AP targeting in rods of wild-type and rhodopsin-knockout mice. We found that the R9AP targeting mechanism does not require the presence of rhodopsin and further demonstrated that R9AP is actively targeted in rods by its SNARE homology domain. 相似文献
15.
Tumor necrosis factor receptor 1 is an ATPase regulated by silencer of death domain 总被引:3,自引:0,他引:3 下载免费PDF全文
Self-aggregation of tumor necrosis factor receptor type 1 (TNFR1) induces spontaneous downstream signaling and results in cell death. It has been suggested that silencer of death domain (SODD) binds TNFR1 monomers to prevent self-aggregation. We found that SODD binds through its BAG domain to the ATPase domain of Hsp70. We also determined that SODD binds through its BAG domain to TNFR1. ATP, but not nonhydrolyzable ATP-gamma S, regulates the SODD binding by Hsp70 or TNFR1. ATP binding by TNFR1 was abolished when a point mutation was introduced into a phosphate-binding loop motif characteristic of ATP-binding proteins, suggesting that TNFR1 functions as an ATPase. Furthermore, TNFR1 was present in aggregates in ATP-depleted cells and SODD disassembled aggregates in vitro only in the presence of ATP. These data suggest that SODD functions as a cofactor analogous to the nucleotide exchange factor BAG-1, which modulates the ATPase cycle of Hsp70 proteins. We propose a new model in which a nucleotide-dependent conformational change in TNFR1 has a key role in regulating TNF signaling. 相似文献
16.
Tumor suppression by p53 in the absence of Atm 总被引:1,自引:0,他引:1
Bailey SL Gurley KE Hoon-Kim K Kelly-Spratt KS Kemp CJ 《Molecular cancer research : MCR》2008,6(7):1185-1192
Oncogenes can induce p53 through a signaling pathway involving p19/Arf. It was recently proposed that oncogenes can also induce DNA damage, and this can induce p53 through the Atm DNA damage pathway. To assess the relative roles of Atm, Arf, and p53 in the suppression of Ras-driven tumors, we examined susceptibility to skin carcinogenesis in 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate (TPA)-treated Atm- and p53-deficient mice and compared these results to previous studies on Arf-deficient mice. Mice with epidermal-specific deletion of p53 showed increased papilloma number and progression to malignant invasive carcinomas compared with wild-type littermates. In contrast, Atm-deficient mice showed no increase in papilloma number, growth, or malignant progression. gamma-H2AX and p53 levels were increased in both Atm(+/+) and Atm(-/-) papillomas, whereas Arf(-/-) papillomas showed much lower p53 expression. Thus, although there is evidence of DNA damage, signaling through Arf seems to regulate p53 in these Ras-driven tumors. In spontaneous and radiation-induced lymphoma models, tumor latency was accelerated in Atm(-/-)p53(-/-) compound mutant mice compared with the single mutant Atm(-/-) or p53(-/-) mice, indicating cooperation between loss of Atm and loss of p53. Although p53-mediated apoptosis was impaired in irradiated Atm(-/-) lymphocytes, p53 loss was still selected for during lymphomagenesis in Atm(-/-) mice. In conclusion, in these models of oncogene- or DNA damage-induced tumors, p53 retains tumor suppressor activity in the absence of Atm. 相似文献
17.
《Autophagy》2013,9(5):563-566
Autophagy plays a critical protective role maintaining energy homeostasis and protein and organelle quality control. These functions are particularly important in times of metabolic stress and in cells with high energy demand such as cancer cells. In emerging cancer cells, autophagy defect may cause failure of energy homeostasis and protein and organelle quality control, leading to the accumulation of cellular damage in metabolic stress. Some manifestations of this damage, such as activation of the DNA damage response and generation of genome instability may promote tumor initiation and drive cell-autonomous tumor progression. In addition, in solid tumors, autophagy localizes to regions that are metabolically stressed. Defects in autophagy impair the survival of tumor cells in these areas, which is associated with increased cell death and inflammation. The cytokine response from inflammation may promote tumor growth and accelerate cell non-autonomous tumor progression. The overreaching theme is that autophagy protects cells from damage accumulation under conditions of metabolic stress allowing efficient tolerance and recovery from stress, and that this is a critical and novel tumor suppression mechanism. The challenge now is to define the precise aspects of autophagy, including energy homeostasis, and protein and organelle turnover, that are required for the proper management of metabolic stress that suppress tumorigenesis. Furthermore, we need to be able to identify human tumors with deficient autophagy, and to develop rational cancer therapies that take advantage of the altered metabolic state and stress responses inherent to this autophagy defect. 相似文献
18.
Wolberg AS Kon RH Monroe DM Ezban M Roberts HR Hoffman M 《Biochemical and biophysical research communications》2000,272(2):332-336
Tissue factor (TF) is a transmembrane molecule that, when exposed to plasma, is the key initiator of coagulation. Cellular TF activity is normally "encrypted", but treating cells with calcium ionophore (i.e. , ionomycin or A23187) increases ("deencrypts") TF activity without increasing TF mRNA or antigen expression. Deencryption results from both plasma membrane phosphatidylserine (PS)-dependent and -independent mechanisms; however, the nature of the PS-independent component is unclear. Since deencryption has been suggested to result from release of TF dimers on the cell surface, and since TF's cytoplasmic domain binds to actin-binding protein 280 and interacts with the cytoskeleton, we hypothesized that interactions with the cytoskeleton, through the cytoplasmic domain, play a role in mediating encryption/deencryption. We examined TF deencryption and the role of the cytoplasmic domain in the PS-independent component using baby hamster kidney (BHK) cells expressing full length TF (BHK-TF) or TF lacking its cytoplasmic domain (BHK-descyt) (Sorensen et al. (1999) J. Biol. Chem. 274, 21349). Both BHK-TF and BHK-descyt cells exhibited a dose-dependent, 1.5- to 10-fold increase in TF activity upon treatment with calcium ionophore, and this increase in activity was only partially blocked by annexin V. These results indicate that deencryption is not restricted to cells which naturally express TF and that the PS-independent component of deencryption is intact on cells transfected with either full length or truncated TF. Our results clearly indicate that deencryption is not dependent on an intact cytoplasmic domain in transfected BHK cells. 相似文献
19.
Broberg A Nissinen L Potila M Heino J 《Biochemical and biophysical research communications》2001,280(1):328-333
Integrin alpha1beta1, one of the cellular collagen receptors, can participate in the regulation of collagen accumulation by acting as a negative feedback regulator. The molecular mechanism behind this phenomenon has been unknown. We have plated cells inside three-dimensional collagen and analyzed a set of chemical inhibitors for various signal transduction pathways. Only two wide-spectrum serine/threonine kinase inhibitors, H-7 and iso-H-7 could prevent the down-regulation of alpha1(I) collagen mRNA levels in cells exposed to three-dimensional collagen. In monolayer iso-H-7 slightly down-regulated collagen gene expression, indicating that inside collagen it affected integrin signaling rather than having a direct stimulatory effect on collagen mRNA levels. The effect of iso-H-7 was not dependent on its ability to inhibit protein kinases A, C, or G. H-7 and iso-H-7 could also inhibit collagen gel contraction, but this mechanism was independent of collagen gene regulation. Three-dimensional collagen could also up-regulate the mRNA levels of several matrix metalloproteinases (MMPs) but H-7 and iso-H-7 had no effect on the regulation of MMP genes. Our data indicate that three-dimensional collagenous matrix regulates distinct cellular signaling pathways and that collagen gene regulation is independent of the other effects of the matrix. 相似文献
20.
Autophagy plays a critical protective role maintaining energy homeostasis and protein and organelle quality control. These functions are particularly important in times of metabolic stress and in cells with high energy demand such as cancer cells. In emerging cancer cells, autophagy defect may cause failure of energy homeostasis and protein and organelle quality control, leading to the accumulation of cellular damage in metabolic stress. Some manifestations of this damage, such as activation of the DNA damage response and generation of genome instability may promote tumor initiation and drive cell-autonomous tumor progression. In addition, in solid tumors, autophagy localizes to regions that are metabolically stressed. Defects in autophagy impair the survival of tumor cells in these areas, which is associated with increased cell death and inflammation. The cytokine response from inflammation may promote tumor growth and accelerate cell non-autonomous tumor progression. The overreaching theme is that autophagy protects cells from damage accumulation under conditions of metabolic stress allowing efficient tolerance and recovery from stress, and that this is a critical and novel tumor suppression mechanism. The challenge now is to define the precise aspects of autophagy, including energy homeostasis and protein and organelle turnover, that are required for the proper management of metabolic stress that suppress tumorigenesis. Furthermore, we need to be able to identify human tumors with deficient autophagy, and to develop rational cancer therapies that take advantage of the altered metabolic state and stress responses inherent to this autophagy defect. 相似文献