Biomedical subjects

J H Miner

Publications and source records attributed to J H Miner.

At least 19 recordsLinked to original sources

PubMed2001

Mystery solved: discovery of a novel integrin ligand in the developing kidney.

Mutant mice lacking the integrin alpha8 subunit exhibit variable defects in kidney development with most mutants missing both kidneys. Several lines of evidence indicate that the known extracellular matrix ligands for integrin alpha8beta1 are either dispensable for or not involved in alpha8beta1 signaling during kidney development. This suggests the presence of an unknown ligand. A novel alpha8beta1 ligand, nephronectin, has now been identified. Nephronectin is a new extracellular matrix protein associated with the Wolffian duct and the ureteric bud, epithelial structures with well-defined roles in kidney development.

Animals↗

PubMed2001

Overexpression of alpha4 chain-containing laminins in human glial tumors identified by gene microarray analysis.

Differential gene expression in tumors often involves growth factors and extracellular matrix/basement membrane components. Here, 11,000- gene microarray was used to identify gene expression profiles in brain tumors including high-grade gliomas [glioblastoma multiforme (GBM) and anaplastic astrocytoma], low-grade astrocytomas, or benign extra-axial brain tumors (meningioma) in comparison with normal brain tissue. Histologically normal tissues adjacent to GBMs were also studied. All GBMs studied overexpressed 14 known genes compared with normal human brain tissue. Overexpressed genes belonged to two broad groups: (a) growth factor-related genes; and (b) structural/extracellular matrix-related genes. For most of these 14 genes, expression levels were lower in low-grade astrocytoma than in GBM and were barely detectable in normal brain. Despite normal-appearing histology, gene expression patterns of tissues immediately adjacent to GBM were similar to those of their respective primary GBMs. Two genes were consistently up-regulated in both high-grade and low-grade gliomas, as well as in histologically normal tissues adjacent to GBMs. These genes coded for the epidermal growth factor receptor (previously reported to be overexpressed in gliomas) and for the alpha4 chain of laminin, a major blood vessel basement membrane component. Changes in expression of this laminin chain have not been previously associated with malignant tumors. Overexpression of laminin alpha4 chain in GBM and astrocytoma grade II by gene microarray analysis was confirmed by semiquantitive reverse transcription-PCR and immunohistochemistry. Importantly, an alpha4 chain-containing laminin isoform, laminin-8 (alpha4beta1gamma1), was expressed mainly in blood vessel walls of GBMs and histologically normal tissues adjacent to GBMs, whereas another alpha4 chain-containing laminin isoform, laminin-9 (alpha4beta2gamma1), was expressed mainly in blood vessel walls of low-grade tumors and normal brain. GBMs that overexpressed laminin-8 had a shorter mean time to tumor recurrence (4.3 months) than GBMs with overexpression of laminin-9 (9.7 months, P = 0.0007). Up-regulation of alpha4 chain-containing laminins could be important for the development of glioma-induced neovascularization and glial tumor progression. Overexpression of laminin-8 may be predictive of glioma recurrence.

Adult↗

PubMed2001

Localization of Lutheran, a novel laminin receptor, in normal, knockout, and transgenic mice suggests an interaction with laminin alpha5 in vivo.

Laminins are major components of all basement membranes. One laminin that has garnered particular interest, due to its widespread expression pattern and importance during development, is the laminin alpha5 chain. In vitro studies have suggested that the Lutheran blood group glycoprotein/basal cell adhesion molecule (Lu), an Ig superfamily transmembrane protein, is a receptor for laminins containing the alpha5 chain. However, there are no in vivo studies showing that these proteins are capable of interacting in tissues. We have isolated the mouse ortholog of Lu and characterized its expression and localization in mouse tissues. Lu was primarily found on the basal surface of epithelial cells and on muscle cells adjacent to basement membranes containing laminin alpha5. In addition, there was both a dramatic reduction in the basal concentration of Lu in mice lacking laminin alpha5, and a significant increase in Lu protein in transgenic mice overexpressing laminin alpha5. Together, these data provide the first in vivo evidence for an interaction between Lu and laminin alpha5 and support the hypothesis that Lu is a laminin alpha5 receptor. We propose that laminin alpha5 is involved in concentrating Lu on the basal surface of epithelial cells. This may be one mechanism by which basement membrane signals are transmitted to the cell.

Animals↗

PubMed2001

CD2AP localizes to the slit diaphragm and binds to nephrin via a novel C-terminal domain.

CD2AP, an adapter protein containing multiple SH3 domains, plays a critical role in kidney function. Mice lacking CD2AP die soon after birth because of kidney failure. In the kidney, CD2AP is expressed in glomerular podocytes, which suggests that it may play a role in a specialized adhesion complex known as the slit diaphragm. One of the major components of the slit diaphragm is nephrin, a podocyte-specific protein. Here we demonstrate that CD2AP localizes to the slit diaphragm in podocytes using immunoelectron microscopy and that nephrin and CD2AP co-immunoprecipitate from a podocyte cell line. The specificity of this interaction was verified by mapping studies, which demonstrated that a novel domain at the C terminus of CD2AP interacts with the C-terminal portion of the nephrin cytoplasmic domain. These studies lend further support to the idea that CD2AP plays a role in the structural integrity of the slit diaphragm.

Adaptor Proteins, Signal Transducing↗

PubMed2001

The laminins in the murine inner ear: developmental transitions and expression in cochlear basement membranes.

The laminins are a family of heterotrimeric extracellular matrix molecules that form suprastructural networks in basement membranes and elsewhere. They interact with integrin receptors, playing key roles in modulating programs of cytodifferentiation and maintaining tissue homeostasis in animals. Earlier studies have demonstrated an extensive laminin network in both the developing and adult cochlea, primarily associated with the basement membranes. These studies, however, did not address the laminin chain composition of these networks. In this study, we used antibodies specific for the known laminin chains to examine the composition of laminins in both the developing and adult murine cochlea. The results illustrate a complex and dynamic postnatal developmental regulation pattern for most of these chains, and suggest that an unusually large number of laminin heterotrimers are present in both the developing and adult cochlea. The laminin composition at postnatal day 2 is relatively simple. By postnatal day 7, however, activation of several laminin chains results in a very complex laminin composition. In the basement membrane underlying the region of the basilar membrane under the developing organ of Corti, eight of the 11 known basement membrane laminins are possible by co-localization inference. Dynamic changes in expression continue through day 14, but simplify by adulthood. Thus, the most dynamic period for laminin expression in the mouse cochlea coincides with terminal cytodifferentiation of the cochlear epithelial structures. Considering the well established role of laminins in regulating both embryonic and organ development in other systems, these data suggest a closer look at the role of the laminins in cochlear development and function may be warranted.

Aging↗

PubMed2001

Regulation of glomerular basement membrane collagen expression by LMX1B contributes to renal disease in nail patella syndrome.

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.

Animals↗

PubMed2001

CD2-associated protein and the kidney.

During the past 3 years there have been significant advances in our understanding of the biology of the glomerular podocyte. In particular, two proteins, CD2-associated protein and nephrin, have been identified as critical podocyte proteins that are both required for normal glomerular filtration. In addition to supporting the idea that the slit diaphragm plays a crucial role in glomerular function, these results suggest novel insights into the pathogenesis of glomerular diseases. The present review addresses these recent advances and discusses the implications of the findings.

Adaptor Proteins, Signal Transducing↗

PubMed2001

Glomerular laminin isoform transitions: errors in metanephric culture are corrected by grafting.

Glomerular basement membrane (GBM) assembly and maturation are marked by the replacement of laminin-1 (containing alpha 1-, beta 1-, and gamma 1-chains) with laminin-11 (consisting of alpha 5-, beta 2-, and gamma 1-chains). Similarly, the alpha 1- and alpha 2-chains of type IV collagen are replaced by collagen alpha 3-, alpha 4-, and alpha 5(IV)-chains. The cellular origins of these molecules and mechanisms for isoform removal and substitution are unknown. To explore glomerular laminin isoform transitions in vitro, we assessed metanephric organ cultures. Standard culture conditions do not support endothelial cell differentiation, and glomerular structures that form in vitro are avascular. Nevertheless, extensive podocyte development occurs in these cultures, including the formation of foot processes and assembly of a GBM-like matrix. Here, we show that the podocyte-specific markers, glomerular epithelial protein 1 and nephrin, which are normally expressed in capillary loop stage glomeruli in vivo, are also expressed by glomerular figures that form in organ culture. However, the GBM-like segments that form in vitro do not undergo normal laminin isoform switching. Instead, both laminin alpha 1- and alpha 5-chains are present, as is the beta 1-chain, but not beta 2. When avascular organ-cultured kidneys are grafted into anterior eye chambers, however, kidney-derived angioblasts establish extensive vasculature by 6 days, and glomeruli are lined by endothelial cells. We evaluated embryonic day 12 (E12) vascular endothelial growth factor receptor (Flk1)-lacZ kidneys that had first been grown in organ culture for 6--7 days and then grafted into wild-type mice. Correct laminin isoform substitution occurred and correlated with the appearance of endothelial cells expressing Flk1. Our findings indicate that endothelial cells, and/or factors present in the circulation, mediate normal GBM laminin isoform transitions in vivo.

Amino Acid Sequence↗

PubMed2000

The presynaptic calcium channel is part of a transmembrane complex linking a synaptic laminin (alpha4beta2gamma1) with non-erythroid spectrin.

Nerve regeneration studies at the neuromuscular junction (NMJ) suggest that synaptic basal lamina components tell the returning axon where to locate neurotransmitter release machinery, including synaptic vesicle clusters and active zones. Good candidates for these components are the synaptic laminins (LNs) containing alpha4, alpha5, or beta2 chains. Results from a beta2 laminin knockout mouse have suggested a linkage of this extracellular laminin to cytosolic synaptic vesicle clusters. Here we report such a transmembrane link at the electric organ synapse, which is homologous to the NMJ. We immunopurified electric organ synaptosomes and found on their surface two laminins of 740 and 900 kDa. The 740 kDa laminin has a composition of alpha4beta2gamma1 (laminin-9). Immunostaining reveals that as in the NMJ, alpha4 and beta2 chains are concentrated at the electric organ synapse. Using detergent-solubilized synaptosomes, we immunoprecipitated a complex containing alpha4beta2gamma1 laminin, the voltage-gated calcium channel, and the cytoskeletal protein spectrin. Other presynaptic proteins such as 900 kDa laminin are not found in this complex. We hypothesize that alpha4beta2gamma1 laminin in the synaptic basal lamina attaches to calcium channel, which in turn is attached to cytosolic spectrin. Spectrin could then organize synaptic vesicle clusters by binding vesicle-associated proteins.

Animals↗

PubMed2000

Defective glomerulogenesis in the absence of laminin alpha5 demonstrates a developmental role for the kidney glomerular basement membrane.

Laminins are major components of all basement membranes. They are a diverse group of alpha/beta/gamma heterotrimers formed from five alpha, three beta, and three gamma chains. Laminin alpha5 is a widely expressed chain found in many embryonic and adult basement membranes. During embryogenesis, alpha5 has a role in disparate developmental processes, including neural tube closure, digit septation, and placentation. Here, we analyzed kidney development in Lama5 mutant embryos and found a striking defect in glomerulogenesis associated with an abnormal glomerular basement membrane (GBM). This correlates with failure of the developmental switch in laminin alpha chain deposition in which alpha5 replaces alpha1 in the GBM at the capillary loop stage of glomerulogenesis. In the absence of a normal GBM, glomerular epithelial cells were in disarray, and endothelial and mesangial cells were extruded from within the constricting glomerulus, leading to a complete absence of vascularized glomeruli. In addition, a minority of Lama5 mutant mice lacked one or both kidneys, indicating that laminin alpha5 is also important in earlier kidney development. Our results demonstrate a dual role for laminin alpha5 in kidney development, illustrate a novel defect in glomerulogenesis, and indicate a heretofore unappreciated developmental role for the GBM in influencing the behavior of epithelial and endothelial cells.

Animals↗

PubMed2000

The synaptic vesicle protein SV2 is complexed with an alpha5-containing laminin on the nerve terminal surface.

Interactions between growing axons and synaptic basal lamina components direct the formation of neuromuscular junctions during nerve regeneration. Isoforms of laminin containing alpha5 or beta2 chains are potential basal lamina ligands for these interactions. The nerve terminal receptors are unknown. Here we show that SV2, a synaptic vesicle transmembrane proteoglycan, is complexed with a 900-kDa laminin on synaptosomes from the electric organ synapse that is similar to the neuromuscular junctions. Although two laminins are present on synaptosomes, only the 900-kDa laminin is associated with SV2. Other nerve terminal components are absent from this complex. The 900-kDa laminin contains an alpha5, a beta1, and a novel gamma chain. To test whether SV2 directly binds the 900-kDa laminin, we looked for interaction between purified SV2 and laminin-1, a laminin isoform with a similar structure. We find SV2 binds with high affinity to purified laminin-1. Our results suggest that a synaptic vesicle component may act as a laminin receptor on the presynaptic plasma membrane; they also suggest a mechanism for activity-dependent adhesion at the synapse.

Animals↗

PubMed2000

Gelatinase B (MMP-9) is not essential in the normal kidney and does not influence progression of renal disease in a mouse model of Alport syndrome.

Matrix metalloproteinases are matrix degrading enzymes implicated in many biological processes, including development and inflammation. Gelatinase B (gelB; also known as MMP-9) is expressed in the kidney and is hypothesized to be involved in basement membrane remodeling and in preventing pathogenic accumulation of extracellular matrix in the kidney. Inhibition of gelB activity in metanephric organ culture disrupts branching morphogenesis of the ureteric bud, suggesting that gelB plays a role in kidney development in vivo. We studied kidneys of gelB-deficient mice to search for developmental, histological, molecular, ultrastructural, and functional defects. Surprisingly, no differences between gelB-/- and control kidneys were detected, and renal function was normal in gelB mutants. In addition, gelB-/- embryonic kidneys developed normally in organ culture. Gelatinase B-deficient mice were bred with Col4a3-/- mice, a model for Alport syndrome, to determine whether gelB influences the progression of glomerulonephritis. This is an important question, as it has been hypothesized that proteases are involved in damaging Alport glomerular basement membrane. However, the presence or absence of gelB did not affect the rate of progression of renal disease. Thus, gelB does not have a discernible role in the normal kidney and gelB is not involved in the progression of glomerulonephritis in a mouse model of Alport syndrome.

Animals↗

PubMed2000

Identification and expression of mouse netrin-4.

Netrins are secreted proteins that serve as potent axon guidance molecules in vertebrates and invertebrates. We report the identification of a novel mammalian member of this family. Netrin-4 is similar in predicted size and secondary structure to the other three netrins; all contain, in order, an amino-terminal signal sequence, a laminin-type globular domain of the 'VI' type, three laminin-type epidermal growth factor (EGF) repeats, and a carboxyl-terminal 'netrin module'. In terms of primary sequence, however, netrin-4 is a distant relative of netrins-1-3, and its globular domain is more closely related to those of laminins than to those of other netrins. Netrin-4 is broadly expressed in both neural and non-neural tissues of embryonic and adult mice. In embryonic spinal cord, it is selectively expressed by cells at the lateral margins of the floor plate. In postnatal brain, it is selectively expressed in subsets of neurons, including cerebellar granule and hippocampal pyramidal cells.

Age Factors↗

PubMed2000

CD2AP is expressed with nephrin in developing podocytes and is found widely in mature kidney and elsewhere.

CD2-associated protein (CD2AP) is an adapter molecule that can bind to the cytoplasmic domain of nephrin, a component of the glomerular slit diaphragm. Mice lacking CD2AP exhibit a congenital nephrotic syndrome characterized by extensive foot process effacement, suggesting that CD2AP-nephrin interactions are critical to maintaining slit diaphragm function. We have examined the patterns of expression of both CD2AP and nephrin in developing mouse and human kidney. Both proteins were first detected in developing podocytes at the capillary loop stage of glomerulogenesis and eventually became concentrated near the glomerular basement membrane. CD2AP was also observed diffusely in collecting duct and apically in many cells of proximal and distal tubule. Kidneys from Cd2ap -/- mice initially exhibited normal nephrin localization, but as the mice aged and foot processes became effaced, nephrin disappeared. In laminin-beta(2) mutant mice exhibiting nephrotic syndrome, CD2AP in glomeruli was aberrantly localized in a primarily punctate pattern. Extensive extrarenal expression of CD2AP was observed in endothelial and epithelial cells, in many cases with a specific subcellular localization. Together, these results suggest that CD2AP is not only involved in maintaining the slit diaphragm but may also have a general role in maintaining specialized subcellular architecture. The severity of kidney disease in Cd2ap mutant mice may have eclipsed manifestation of defects in other tissues.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Expression patterns of laminin alpha1 and alpha5 in human lung during development.

Laminins are trimeric glycoprotein components of basement membranes. Each laminin has three structurally similar chains, designated alpha, beta, and gamma. Five laminin alpha chains are now known. In previous studies using monoclonal antibody 4C7, laminin alpha1 was thought to be present in basement membranes of human lung throughout development and in the adult, but recent expression studies have demonstrated that 4C7 identifies laminin alpha5 rather than alpha1. To determine the temporal and spatial patterns of laminin alpha1 and laminin alpha5 in developing human lung, we prepared complementary DNA probes specific for laminin alpha1 and alpha5 messenger RNAs (mRNAs). By Northern analysis, laminin alpha1 mRNA was prominent in first-trimester fetal lung, but was not detectable at 23 wk or at later times. In contrast, laminin alpha5 mRNA was readily detected in early fetal lung and remained present thereafter. Immunohistochemical staining demonstrated laminin alpha1 only in early fetal lung, whereas laminin alpha5 was persistent from the early fetal period. In situ hybridization localized laminin alpha1 expression to distal epithelium in the first-trimester lung, and laminin alpha5 to all epithelium and developing pulmonary arteries from the first trimester through the perinatal period. These studies indicate that laminin alpha1 expression is restricted to early human lung morphogenesis, whereas the expression of laminin alpha5 in human lung is continuous from early lung development through adult life. It is evident that laminin alpha1 and laminin alpha5 have different roles in the development of the human lung.

Blotting, Northern↗

PubMed1999

Cell elongation induces laminin alpha2 chain expression in mouse embryonic mesenchymal cells: role in visceral myogenesis.

Bronchial smooth muscle (SM) mesenchymal cell precursors change their shape from round to spread/elongated while undergoing differentiation. Here we show that this change in cell shape induces the expression of laminin (LM) alpha2 chain not present in round mesenchymal cells. LM alpha2 expression is reversible and switched on and off by altering the cell's shape in culture. In comparison, the expression of LM beta1 and gamma1 remains unchanged. Functional studies showed that mesenchymal cell spreading and further differentiation into SM are inhibited by an antibody against LM alpha2. Dy/dy mice express very low levels of LM alpha2 and exhibit congenital muscular dystrophy. Lung SM cells isolated from adult dy/dy mice spread defectively and synthesized less SM alpha-actin, desmin, and SM-myosin than controls. These deficiencies were completely corrected by exogenous LM-2. On histological examination, dy/dy mouse airways and gastrointestinal tract had shorter SM cells, and lungs from dy/dy mice contained less SM-specific protein. The intestine, however, showed compensatory hyperplasia, perhaps related to its higher contractile activity. This study therefore demonstrated a novel role for the LM alpha2 chain in SM myogenesis and showed that its decrease in dy/dy mice results in abnormal SM.

Actins↗

PubMed1999

Insertional mutation of the collagen genes Col4a3 and Col4a4 in a mouse model of Alport syndrome.

Mice homozygous for the transgenic insertion in line OVE250 exhibit severe progressive glomerulonephritis. Ultrastructural changes in the glomerular basement membrane (GBM) at 2 weeks of age resemble those in Alport syndrome. The transgenic insertion site was mapped by FISH to mouse chromosome 1 close to Pax3. Genetic and molecular analyses identified a deletion of genomic DNA at the transgene insertion site. Exons 1 through 12 of the collagen IV gene Col4a4, exons 1 and 2 of the adjacent Col4a3 gene, and the intergenic promoter region are deleted. Transcripts of Col4a3 and Col4a4 are undetectable in mutant kidney, and both proteins are missing from the GBM. Persistent cellular proliferation in mutant kidneys suggests that interaction with the extracellular matrix may be important for cell maturation. Evolutionarily conserved sequence elements in the promoter regions of human and mouse Col4a3 and Col4a4 include a 19-bp element that was tandemly duplicated in the human lineage and a CTC box element common to several genes encoding extracellular matrix proteins. This new animal model of Alport syndrome, Col4Delta3-4, lacks both alpha3 and alpha4 chains of collagen IV and exhibits an earlier disease onset than mice lacking alpha3 only.

Animals↗

PubMed1999

Congenital nephrotic syndrome in mice lacking CD2-associated protein.

CD2-associated protein (CD2AP) is an 80-kilodalton protein that is critical for stabilizing contacts between T cells and antigen-presenting cells. In CD2AP-deficient mice, immune function was compromised, but the mice died at 6 to 7 weeks of age from renal failure. In the kidney, CD2AP was expressed primarily in glomerular epithelial cells. Knockout mice exhibited defects in epithelial cell foot processes, accompanied by mesangial cell hyperplasia and extracellular matrix deposition. Supporting a role for CD2AP in the specialized cell junction known as the slit diaphragm, CD2AP associated with nephrin, the primary component of the slit diaphragm.

Adaptor Proteins, Signal Transducing↗