The Experts below are selected from a list of 1474299 Experts worldwide ranked by ideXlab platform
Billy G Hudson - One of the best experts on this subject based on the ideXlab platform.
-
crystal structure of nc1 domains structural basis for type iv collagen assembly in basement membranes
Journal of Biological Chemistry, 2002Co-Authors: Munirathinam Sundaramoorthy, Parvin Todd, Muthuraman Meiyappan, Billy G HudsonAbstract:Type IV collagen, which is present in all metazoan, exists as a family of six homologous Alpha(IV) chains, Alpha1-Alpha6, in mammals. The six chains assemble into three different triple helical protomers and self-associate as three distinct networks. The network underlies all epithelia as a component of basement membranes, which play important roles in cell adhesion, growth, differentiation, tissue repair and molecular ultrafiltration. The specificity of both protomer and network assembly is governed by amino acid sequences of the C-terminal noncollagenous (NC1) domain of each chain. In this study, the structural basis for protomer and network assembly was investigated by determining the crystal structure of the ubiquitous [(Alpha1)(2).Alpha2](2) NC1 hexamer of bovine lens capsule basement membrane at 2.0 A resolution. The NC1 monomer folds into a novel tertiary structure. The (Alpha1)(2).Alpha2 trimer is organized through the unique three-dimensional domain swapping interactions. The differences in the primary sequences of the hypervariable region manifest in different secondary structures, which determine the chain specificity at the monomer-monomer interfaces. The trimer-trimer interface is stabilized by the extensive hydrophobic and hydrophilic interactions without a need for disulfide cross-linking.
-
the nc1 domain of collagen iv encodes a novel network composed of the Alpha 1 Alpha 2 Alpha 5 and Alpha 6 chains in smooth muscle basement membranes
Journal of Biological Chemistry, 2001Co-Authors: Dorinbogdan Borza, Yoshifumi Ninomiya, Ichiro Naito, Yoshikazu Sado, Olga Bondar, Parvin Todd, Billy G HudsonAbstract:Type IV collagen, the major component of basement membranes (BMs), is a family of six homologous chains (Alpha1-Alpha6) that have a tissue-specific distribution. The chains assemble into supramolecular networks that differ in the chain composition. In this study, a novel network was identified and characterized in the smooth muscle BMs of aorta and bladder. The noncollagenous (NC1) hexamers solubilized by collagenase digestion were fractionated by affinity chromatography using monoclonal antibodies against the Alpha5 and Alpha6 NC1 domains and then characterized by two-dimensional gel electrophoresis and Western blotting. Both BMs were found to contain a novel Alpha1.Alpha2.Alpha5.Alpha6 network besides the classical Alpha1.Alpha2 network. The Alpha1.Alpha2.Alpha5.Alpha6 network represents a new arrangement in which a protomer (triple-helical isoform) containing the Alpha5 and Alpha6 chains is linked through NC1-NC1 interactions to an adjoining protomer composed of the Alpha1 and Alpha2 chains. Re-association studies revealed that the NC1 domains contain recognition sequences sufficient to encode the assembly of both networks. These findings, together with previous ones, indicate that the six chains of type IV collagen are distributed in three major networks (Alpha1.Alpha2, Alpha3.Alpha4.Alpha5, and Alpha1.Alpha2.Alpha5.Alpha6) whose chain composition is encoded by the NC1 domains. The existence of the Alpha1.Alpha2.Alpha5.Alpha6 network provides a molecular explanation for the concomitant loss of Alpha5 and Alpha6 chains from the BMs of patients with X-linked Alport's syndrome.
-
glomerular basement membrane identification of a novel disulfide cross linked network of α3 α4 and α5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
-
glomerular basement membrane identification of a novel disulfide cross linked network of Alpha3 Alpha4 and Alpha5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
-
isoform switching of type iv collagen is developmentally arrested in x linked alport syndrome leading to increased susceptibility of renal basement membranes to endoproteolysis
Journal of Clinical Investigation, 1997Co-Authors: Raghuram Kalluri, Billy G Hudson, Parvin Todd, Charles F Shield, Eric G NeilsonAbstract:Normal glomerular capillaries filter plasma through a basement membrane (GBM) rich in Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains of type IV collagen. We now show that these latter isoforms are absent biochemically from the glomeruli in patients with X-linked Alport syndrome (XAS). Their GBM instead retain a fetal distribution of Alpha1(IV) and Alpha2(IV) isoforms because they fail to developmentally switch their Alpha-chain use. The anomalous persistence of these fetal isoforms of type IV collagen in the GBM in XAS also confers an unexpected increase in susceptibility to proteolytic attack by collagenases and cathepsins. The incorporation of cysteine-rich Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains into specialized basement membranes like the GBM may have normally evolved to protectively enhance their resistance to proteolytic degradation at the site of glomerular filtration. The relative absence of these potentially protective collagen IV isoforms in GBM from XAS may explain the progressive basement membrane splitting and increased damage as these kidneys deteriorate.
John R Atack - One of the best experts on this subject based on the ideXlab platform.
-
discriminative stimulus effects of l 838 417 7 tert butyl 3 2 5 difluoro phenyl 6 2 methyl 2h 1 2 4 triazol 3 ylmethoxy 1 2 4 triazolo 4 3 b pyridazine role of gabaa receptor subtypes
Neuropharmacology, 2010Co-Authors: Stephanie C Licata, John R Atack, Donna M Platt, Daniela Ruedibettschen, Gerard R Dawson, Michael L Van Linn, James M Cook, James K RowlettAbstract:Previous reports suggest that gamma-aminobutyric acid type A (GABA(A)) receptors containing Alpha1 subunits may play a pivotal role in mediating the discriminative stimulus effects of benzodiazepines (BZs). L-838,417 (7-tert-Butyl-3-(2,5-difluoro-phenyl)-6-(2-methyl-2H-[1,2,4]triazol-3-ylmethoxy)-[1,2,4]triazolo[4,3-b]pyridazine) is a GABA(A) receptor modulator with intrinsic efficacy in vitro at Alpha2, Alpha3, and Alpha5 subunit-containing GABA(A) receptors, and little demonstrable intrinsic efficacy in vitro at Alpha1 subunit-containing GABA(A) receptors. The present study evaluated the discriminative stimulus effects of L-838,417 in order to determine the extent to which the Alpha2, Alpha3, and Alpha5 subunit-containing GABA(A) receptors contribute to the interoceptive effects of BZ-type drugs. Squirrel monkeys (Saimiri sciureus) were trained to discriminate L-838,417 (0.3 mg/kg, i.v.) from vehicle under a 5-response fixed-ratio schedule of food reinforcement. Under test conditions, L-838,417 administration resulted in dose-dependent increases in drug-lever responding that were antagonized by the BZ-site antagonist, flumazenil. Administration of non-selective BZs, compounds with 10-fold greater affinity for Alpha1 subunit-containing GABA(A) receptors compared to Alpha2, Alpha3, and Alpha5 subunit-containing GABA(A) receptors, barbiturates and ethanol (which modulate the GABA(A) receptor via a non-BZ site), all resulted in a majority of responses on the L-838,417-paired lever (65-100% drug-lever responding). betaCCT, an antagonist that binds with 20-fold greater affinity for Alpha1 subunit-containing GABA(A) receptors relative to Alpha2, Alpha3, and Alpha5-containing GABA(A) receptors, had no significant effect on the discriminative stimulus effects of L-838,417 or the L-838,417-like effects of diazepam or zolpidem. These data suggest that efficacy at Alpha2, Alpha3, and/or Alpha5 subunit-containing GABA(A) receptors likely are sufficient for engendering BZ-like discriminative stimulus effects.
-
benzodiazepine binding site occupancy by the novel gabaa receptor subtype selective drug 7 1 1 dimethylethyl 6 2 ethyl 2h 1 2 4 triazol 3 ylmethoxy 3 2 fluorophenyl 1 2 4 triazolo 4 3 b pyridazine tpa023 in rats primates and humans
Journal of Pharmacology and Experimental Therapeutics, 2010Co-Authors: John R Atack, Dean Foster Wong, Tim D Fryer, Christine Ryan, Sandra Sanabria, Yun Zhou, Robert F Dannals, Raymond E Gibson, Donald H BurnsAbstract:The GABA(A) receptor Alpha2/Alpha3 subtype-selective compound 7-(1,1-dimethylethyl)-6-(2-ethyl-2H-1,2,4-triazol-3-ylmethoxy)-3-(2-fluorophenyl)-1,2,4-triazolo[4,3-b]pyridazine (TPA023; also known as MK-0777) is a triazolopyridazine that has similar, subnanomolar affinity for the benzodiazepine binding site of Alpha1-, Alpha2-, Alpha3-, and Alpha5-containing GABA(A) receptors and has partial agonist efficacy at the Alpha2 and Alpha3 but not the Alpha1 or Alpha5 subtypes. The purpose of the present study was to define the relationship between plasma TPA023 concentrations and benzodiazepine binding site occupancy across species measured using various methods. Thus, occupancy was measured using either in vivo [(3)H]flumazenil binding or [(11)C]flumazenil small-animal positron emission tomography (microPET) in rats, [(123)I]iomazenil gamma-scintigraphy in rhesus monkeys, and [(11)C]flumazenil PET in baboons and humans. For each study, plasma-occupancy curves were derived, and the plasma concentration of TPA023 required to produce 50% occupancy (EC(50)) was calculated. The EC(50) values for rats, rhesus monkeys, and baboons were all similar and ranged from 19 to 30 ng/ml, although in humans, the EC(50) was slightly lower at 9 ng/ml. In humans, a single 2-mg dose of TPA023 produced in the region of 50 to 60% occupancy in the absence of overt sedative-like effects. Considering that nonselective full agonists are associated with sedation at occupancies of less than 30%, these data emphasize the relatively nonsedating nature of TPA023.
Yoshikazu Sado - One of the best experts on this subject based on the ideXlab platform.
-
basement membrane type iv collagen molecules in the choroid plexus pia mater and capillaries in the mouse brain
Archives of Histology and Cytology, 2002Co-Authors: Norio Urabe, Ichiro Naito, Keriji Saito, Tomoko Yonezawa, Yoshikazu Sado, Hidekatsu Yoshioka, Shozo Kusachi, Takao Tsuji, Aiji OhtsukaAbstract:We investigated the differential distribution of basement membrane type IV collagen a chains in the mouse brain by immunohistochemistry using a chain-specific monoclonal antibodies. Subendothelial basement membranes were found to contain Alpha1 and Alpha2 chains. Basement membranes surrounding smooth muscle cells on blood vascular walls were immunoreactive for Alpha1 and Alpha2 chains but not for Alpha5 and Alpha6 chains. Interestingly, the pia mater contained a thin basement membrane which was positive for Alpha1, Alpha2, Alpha5, and Alpha6 chains, suggesting that glia limitans superficialis coheres basement membranes containing [Alpha1(IV)]2Alpha2(IV) and [Alpha5(IV)]2Alpha6(IV) molecules. In contrast, capillaries always possessed thin basement membranes of [Alpha1(IV)]2Alpha2(IV) molecules. Cerebrospinal fluid is produced through filtration of blood at the choroid plexus, where two distinct basement membranes were detected by anti-al and anti-Alpha2 antibodies. The subendothelial basement membrane appeared to consist of [Alpha1(IV)]2Alpha2(IV) molecules, whereas the subependymal basement membrane in the choroid plexus was strongly positive for Alpha3, Alpha4, and Alpha5 chains, indicating that the filtering unit was composed of Alpha3(IV)Alpha4(IV)Alpha5(IV) molecules. That the specific localizations of these molecules are shared by renal glomeruli and the choroid plexus leads us to hypothesize that the supramolecular network containing Alpha3(IV) Alpha4(IV)Alpha5(IV) molecules may function as a permeability selective barrier.
-
the nc1 domain of collagen iv encodes a novel network composed of the Alpha 1 Alpha 2 Alpha 5 and Alpha 6 chains in smooth muscle basement membranes
Journal of Biological Chemistry, 2001Co-Authors: Dorinbogdan Borza, Yoshifumi Ninomiya, Ichiro Naito, Yoshikazu Sado, Olga Bondar, Parvin Todd, Billy G HudsonAbstract:Type IV collagen, the major component of basement membranes (BMs), is a family of six homologous chains (Alpha1-Alpha6) that have a tissue-specific distribution. The chains assemble into supramolecular networks that differ in the chain composition. In this study, a novel network was identified and characterized in the smooth muscle BMs of aorta and bladder. The noncollagenous (NC1) hexamers solubilized by collagenase digestion were fractionated by affinity chromatography using monoclonal antibodies against the Alpha5 and Alpha6 NC1 domains and then characterized by two-dimensional gel electrophoresis and Western blotting. Both BMs were found to contain a novel Alpha1.Alpha2.Alpha5.Alpha6 network besides the classical Alpha1.Alpha2 network. The Alpha1.Alpha2.Alpha5.Alpha6 network represents a new arrangement in which a protomer (triple-helical isoform) containing the Alpha5 and Alpha6 chains is linked through NC1-NC1 interactions to an adjoining protomer composed of the Alpha1 and Alpha2 chains. Re-association studies revealed that the NC1 domains contain recognition sequences sufficient to encode the assembly of both networks. These findings, together with previous ones, indicate that the six chains of type IV collagen are distributed in three major networks (Alpha1.Alpha2, Alpha3.Alpha4.Alpha5, and Alpha1.Alpha2.Alpha5.Alpha6) whose chain composition is encoded by the NC1 domains. The existence of the Alpha1.Alpha2.Alpha5.Alpha6 network provides a molecular explanation for the concomitant loss of Alpha5 and Alpha6 chains from the BMs of patients with X-linked Alport's syndrome.
-
type iv collagen of the glomerular basement membrane evidence that the chain specificity of network assembly is encoded by the noncollagenous nc1 domains
Journal of Biological Chemistry, 2000Co-Authors: Ariel Boutaud, Yoshifumi Ninomiya, Dorinbogdan Borza, Olga Bondar, Kaiolaf Netzer, Narinder Singh, Yoshikazu SadoAbstract:The ultrafiltration function of the glomerular basement membrane (GBM) of the kidney is impaired in genetic and acquired diseases that affect type IV collagen. The GBM is composed of five (Alpha1 to Alpha5) of the six chains of type IV collagen, organized into an Alpha1.Alpha2(IV) and an Alpha3.Alpha4.Alpha5(IV) network. In Alport syndrome, mutations in any of the genes encoding the Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains cause the absence of the Alpha3. Alpha4.Alpha5 network, which leads to progressive renal failure. In the present study, the molecular mechanism underlying the network defect was explored by further characterization of the chain organization and elucidation of the discriminatory interactions that govern network assembly. The existence of the two networks was further established by analysis of the hexameric complex of the noncollagenous (NC1) domains, and the Alpha5 chain was shown to be linked to the Alpha3 and Alpha4 chains by interaction through their respective NC1 domains. The potential recognition function of the NC1 domains in network assembly was investigated by comparing the composition of native NC1 hexamers with hexamers that were dissociated and reconstituted in vitro and with hexamers assembled in vitro from purified Alpha1-Alpha5(IV) NC1 monomers. The results showed that NC1 monomers associate to form native-like hexamers characterized by two distinct populations, an Alpha1.Alpha2 and Alpha3.Alpha4.Alpha5 heterohexamer. These findings indicate that the NC1 monomers contain recognition sequences for selection of chains and protomers that are sufficient to encode the assembly of the Alpha1.Alpha2 and Alpha3.Alpha4.Alpha5 networks of GBM. Moreover, hexamer formation from the Alpha3, Alpha4, and Alpha5 NC1 monomers required co-assembly of all three monomers, suggesting that mutations in the NC1 domain in Alport syndrome may disrupt the assembly of the Alpha3.Alpha4.Alpha5 network by interfering with the assembly of the Alpha3.Alpha4.Alpha5 NC1 hexamer.
-
glomerular basement membrane identification of a novel disulfide cross linked network of α3 α4 and α5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
-
glomerular basement membrane identification of a novel disulfide cross linked network of Alpha3 Alpha4 and Alpha5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
Yoshifumi Ninomiya - One of the best experts on this subject based on the ideXlab platform.
-
the nc1 domain of collagen iv encodes a novel network composed of the Alpha 1 Alpha 2 Alpha 5 and Alpha 6 chains in smooth muscle basement membranes
Journal of Biological Chemistry, 2001Co-Authors: Dorinbogdan Borza, Yoshifumi Ninomiya, Ichiro Naito, Yoshikazu Sado, Olga Bondar, Parvin Todd, Billy G HudsonAbstract:Type IV collagen, the major component of basement membranes (BMs), is a family of six homologous chains (Alpha1-Alpha6) that have a tissue-specific distribution. The chains assemble into supramolecular networks that differ in the chain composition. In this study, a novel network was identified and characterized in the smooth muscle BMs of aorta and bladder. The noncollagenous (NC1) hexamers solubilized by collagenase digestion were fractionated by affinity chromatography using monoclonal antibodies against the Alpha5 and Alpha6 NC1 domains and then characterized by two-dimensional gel electrophoresis and Western blotting. Both BMs were found to contain a novel Alpha1.Alpha2.Alpha5.Alpha6 network besides the classical Alpha1.Alpha2 network. The Alpha1.Alpha2.Alpha5.Alpha6 network represents a new arrangement in which a protomer (triple-helical isoform) containing the Alpha5 and Alpha6 chains is linked through NC1-NC1 interactions to an adjoining protomer composed of the Alpha1 and Alpha2 chains. Re-association studies revealed that the NC1 domains contain recognition sequences sufficient to encode the assembly of both networks. These findings, together with previous ones, indicate that the six chains of type IV collagen are distributed in three major networks (Alpha1.Alpha2, Alpha3.Alpha4.Alpha5, and Alpha1.Alpha2.Alpha5.Alpha6) whose chain composition is encoded by the NC1 domains. The existence of the Alpha1.Alpha2.Alpha5.Alpha6 network provides a molecular explanation for the concomitant loss of Alpha5 and Alpha6 chains from the BMs of patients with X-linked Alport's syndrome.
-
regulation of glomerular basement membrane collagen expression by lmx1b contributes to renal disease in nail patella syndrome
Nature Genetics, 2001Co-Authors: Roy Morello, Yoshifumi Ninomiya, Guang Zhou, Sandra D Dreyer, Scott J Harvey, Paul S Thorner, Jeffrey H Miner, William W Cole, Andreas WinterpachtAbstract:Basement membrane (BM) morphogenesis is critical for normal kidney function. Heterotrimeric type IV collagen, composed of different combinations of six Alpha-chains (1-6), is a major matrix component of all BMs (ref. 2). Unlike in other BMs, glomerular BM (GBM) contains primarily the Alpha 3(IV) and Alpha 4(IV) chains, together with the Alpha 5(IV) chain. A poorly understood, coordinated temporal and spatial switch in gene expression from ubiquitously expressed Alpha 1(IV) and Alpha 2(IV) collagen to the Alpha 3(IV), Alpha 4(IV) and Alpha 5(IV) chains occurs during normal embryogenesis of GBM (ref. 4). Structural abnormalities of type IV collagen have been associated with diverse biological processes including defects in molecular filtration in Alport syndrome, cell differentiation in hereditary leiomyomatosis, and autoimmunity in Goodpasture syndrome; however, the transcriptional and developmental regulation of type IV collagen expression is unknown. Nail patella syndrome (NPS) is caused by mutations in LMX1B, encoding a LIM homeodomain transcription factor. Some patients have nephrosis-associated renal disease characterized by typical ultrastructural abnormalities of GBM (refs. 8,9). In Lmx1b(-/-) mice, expression of both Alpha(3)IV and Alpha(4)IV collagen is strongly diminished in GBM, whereas that of Alpha1, Alpha2 and Alpha5(IV) collagen is unchanged. Moreover, LMX1B binds specifically to a putative enhancer sequence in intron 1 of both mouse and human COL4A4 and upregulates reporter constructs containing this enhancer-like sequence. These data indicate that LMX1B directly regulates the coordinated expression of Alpha 3(IV) and Alpha 4(IV) collagen required for normal GBM morphogenesis and that its dysregulation in GBM contributes to the renal pathology and nephrosis in NPS.
-
type iv collagen of the glomerular basement membrane evidence that the chain specificity of network assembly is encoded by the noncollagenous nc1 domains
Journal of Biological Chemistry, 2000Co-Authors: Ariel Boutaud, Yoshifumi Ninomiya, Dorinbogdan Borza, Olga Bondar, Kaiolaf Netzer, Narinder Singh, Yoshikazu SadoAbstract:The ultrafiltration function of the glomerular basement membrane (GBM) of the kidney is impaired in genetic and acquired diseases that affect type IV collagen. The GBM is composed of five (Alpha1 to Alpha5) of the six chains of type IV collagen, organized into an Alpha1.Alpha2(IV) and an Alpha3.Alpha4.Alpha5(IV) network. In Alport syndrome, mutations in any of the genes encoding the Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains cause the absence of the Alpha3. Alpha4.Alpha5 network, which leads to progressive renal failure. In the present study, the molecular mechanism underlying the network defect was explored by further characterization of the chain organization and elucidation of the discriminatory interactions that govern network assembly. The existence of the two networks was further established by analysis of the hexameric complex of the noncollagenous (NC1) domains, and the Alpha5 chain was shown to be linked to the Alpha3 and Alpha4 chains by interaction through their respective NC1 domains. The potential recognition function of the NC1 domains in network assembly was investigated by comparing the composition of native NC1 hexamers with hexamers that were dissociated and reconstituted in vitro and with hexamers assembled in vitro from purified Alpha1-Alpha5(IV) NC1 monomers. The results showed that NC1 monomers associate to form native-like hexamers characterized by two distinct populations, an Alpha1.Alpha2 and Alpha3.Alpha4.Alpha5 heterohexamer. These findings indicate that the NC1 monomers contain recognition sequences for selection of chains and protomers that are sufficient to encode the assembly of the Alpha1.Alpha2 and Alpha3.Alpha4.Alpha5 networks of GBM. Moreover, hexamer formation from the Alpha3, Alpha4, and Alpha5 NC1 monomers required co-assembly of all three monomers, suggesting that mutations in the NC1 domain in Alport syndrome may disrupt the assembly of the Alpha3.Alpha4.Alpha5 network by interfering with the assembly of the Alpha3.Alpha4.Alpha5 NC1 hexamer.
-
glomerular basement membrane identification of a novel disulfide cross linked network of α3 α4 and α5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
-
glomerular basement membrane identification of a novel disulfide cross linked network of Alpha3 Alpha4 and Alpha5 chains of type iv collagen and its implications for the pathogenesis of alport syndrome
Journal of Biological Chemistry, 1998Co-Authors: Sripad Gunwar, Yoshifumi Ninomiya, Yoshikazu Sado, Milton E Noelken, Fernando Ballester, Billy G HudsonAbstract:Glomerular basement membrane (GBM) plays a crucial function in the ultrafiltration of blood plasma by the kidney. This function is impaired in Alport syndrome, a hereditary disorder that is caused by mutations in the gene encoding type IV collagen, but it is not known how the mutations lead to a defective GBM. In the present study, the supramolecular organization of type IV collagen of GBM was investigated. This was accomplished by using pseudolysin (EC 3.4.24.26) digestion to excise truncated triple-helical protomers for structural studies. Two distinct sets of truncated protomers were solubilized, one at 4 degrees C and the other at 25 degrees C, and their chain composition was determined by use of monoclonal antibodies. The 4 degrees C protomers comprise the Alpha1(IV) and Alpha2(IV) chains, whereas the 25 degrees C protomers comprised mainly Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains along with some Alpha1(IV) and Alpha2(IV) chains. The structure of the 25 degrees C protomers was examined by electron microscopy and was found to be characterized by a network containing loops and supercoiled triple helices, which are stabilized by disulfide cross-links between Alpha3(IV), Alpha4(IV), and Alpha5(IV) chains. These results establish a conceptual framework to explain several features of the GBM abnormalities of Alport syndrome. In particular, the Alpha3(IV). Alpha4(IV).Alpha5(IV) network, involving a covalent linkage between these chains, suggests a molecular basis for the conundrum in which mutations in the gene encoding the Alpha5(IV) chain cause defective assembly of not only Alpha5(IV) chain but also the Alpha3(IV) and Alpha4(IV) chains in the GBM of patients with Alport syndrome.
Donald H Burns - One of the best experts on this subject based on the ideXlab platform.
-
benzodiazepine binding site occupancy by the novel gabaa receptor subtype selective drug 7 1 1 dimethylethyl 6 2 ethyl 2h 1 2 4 triazol 3 ylmethoxy 3 2 fluorophenyl 1 2 4 triazolo 4 3 b pyridazine tpa023 in rats primates and humans
Journal of Pharmacology and Experimental Therapeutics, 2010Co-Authors: John R Atack, Dean Foster Wong, Tim D Fryer, Christine Ryan, Sandra Sanabria, Yun Zhou, Robert F Dannals, Raymond E Gibson, Donald H BurnsAbstract:The GABA(A) receptor Alpha2/Alpha3 subtype-selective compound 7-(1,1-dimethylethyl)-6-(2-ethyl-2H-1,2,4-triazol-3-ylmethoxy)-3-(2-fluorophenyl)-1,2,4-triazolo[4,3-b]pyridazine (TPA023; also known as MK-0777) is a triazolopyridazine that has similar, subnanomolar affinity for the benzodiazepine binding site of Alpha1-, Alpha2-, Alpha3-, and Alpha5-containing GABA(A) receptors and has partial agonist efficacy at the Alpha2 and Alpha3 but not the Alpha1 or Alpha5 subtypes. The purpose of the present study was to define the relationship between plasma TPA023 concentrations and benzodiazepine binding site occupancy across species measured using various methods. Thus, occupancy was measured using either in vivo [(3)H]flumazenil binding or [(11)C]flumazenil small-animal positron emission tomography (microPET) in rats, [(123)I]iomazenil gamma-scintigraphy in rhesus monkeys, and [(11)C]flumazenil PET in baboons and humans. For each study, plasma-occupancy curves were derived, and the plasma concentration of TPA023 required to produce 50% occupancy (EC(50)) was calculated. The EC(50) values for rats, rhesus monkeys, and baboons were all similar and ranged from 19 to 30 ng/ml, although in humans, the EC(50) was slightly lower at 9 ng/ml. In humans, a single 2-mg dose of TPA023 produced in the region of 50 to 60% occupancy in the absence of overt sedative-like effects. Considering that nonselective full agonists are associated with sedation at occupancies of less than 30%, these data emphasize the relatively nonsedating nature of TPA023.
