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1.
Neurobiol Dis ; 40(3): 503-17, 2010 Dec.
Article in English | MEDLINE | ID: mdl-20659558

ABSTRACT

Mutations in the Leucine Rich Repeat Kinase 2 (LRRK2) gene, first described in 2004 have now emerged as the most important genetic finding in both autosomal dominant and sporadic Parkinson's disease (PD). While a formidable research effort has ensued since the initial gene discovery, little is known of either the normal or the pathological role of LRRK2. We have created lines of mice that express human wild-type (hWT) or G2019S Lrrk2 via bacterial artificial chromosome (BAC) transgenesis. In vivo analysis of the dopaminergic system revealed abnormal dopamine neurotransmission in both hWT and G2019S transgenic mice evidenced by a decrease in extra-cellular dopamine levels, which was detected without pharmacological manipulation. Immunopathological analysis revealed changes in localization and increased phosphorylation of microtubule binding protein tau in G2019S mice. Quantitative biochemical analysis confirmed the presence of differential phospho-tau species in G2019S mice but surprisingly, upon dephosphorylation the tau isoform banding pattern in G2019S mice remained altered. This suggests that other post-translational modifications of tau occur in G2019S mice. We hypothesize that Lrrk2 may impact on tau processing which subsequently leads to increased phosphorylation. Our models will be useful for further understanding of the mechanistic actions of LRRK2 and future therapeutic screening.


Subject(s)
Brain/metabolism , Protein Serine-Threonine Kinases/genetics , Synaptic Transmission/physiology , tau Proteins/metabolism , Animals , Autoradiography , Chromatography, High Pressure Liquid , Chromosomes, Artificial, Bacterial , Dopamine/metabolism , Humans , Immunoblotting , In Situ Hybridization , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 , Male , Mice , Mice, Transgenic , Microdialysis , Phosphorylation , Protein Processing, Post-Translational , Receptors, Dopamine D1/metabolism , Receptors, Dopamine D2/metabolism , Reverse Transcriptase Polymerase Chain Reaction
3.
J Biol Chem ; 275(15): 11465-9, 2000 Apr 14.
Article in English | MEDLINE | ID: mdl-10753964

ABSTRACT

Mutations in kerato-epithelin are responsible for a group of hereditary cornea-specific deposition diseases, 5q31-linked corneal dystrophies. These conditions are characterized by progressive accumulation of protein deposits of different ultrastructure. Herein, we studied the corneas with mutations at kerato-epithelin residue Arg-124 resulting in amyloid (R124C), non-amyloid (R124L), and a mixed pattern of deposition (R124H). We found that aggregated kerato-epithelin comprised all types of pathological deposits. Each mutation was associated with characteristic changes of protein turnover in corneal tissue. Amyloidogenesis in R124C corneas was accompanied by the accumulation of N-terminal kerato-epithelin fragments, whereby species of 44 kDa were the major constituents of amyloid fibrils. R124H corneas with prevailing non-amyloid inclusions showed accumulation of a new 66-kDa species altogether with the full-size 68-kDa form. Finally, in R124L cornea with non amyloid deposits, we found only the accumulation of the 68-kDa form. Two-dimensional gels revealed mutation-specific changes in the processing of the full-size protein in all affected corneas. It appears that substitutions at the same residue (Arg-124) result in cornea-specific deposition of kerato-epithelin via distinct aggregation pathways each involving altered turnover of the protein in corneal tissue.


Subject(s)
Amyloid/metabolism , Chromosomes, Human, Pair 5 , Corneal Dystrophies, Hereditary/genetics , Extracellular Matrix Proteins , Mutation , Neoplasm Proteins/genetics , Transforming Growth Factor beta , Animals , Cornea/chemistry , Humans , Molecular Weight , Neoplasm Proteins/analysis , Neoplasm Proteins/metabolism , Rabbits
4.
Invest Ophthalmol Vis Sci ; 40(10): 2213-9, 1999 Sep.
Article in English | MEDLINE | ID: mdl-10476785

ABSTRACT

PURPOSE: Recently, the authors identified a gene, BIGH3, in which different mutations cause a group of hereditary corneal dystrophies: lattice type I and IIIA (CDLI and CDLIIIA), granular Groenouw type I (CDGGI), Avellino (CDA), and Reis-Bücklers' (CDRB). All these disorders are characterized by the progressive accumulation of corneal deposits with different structural organization. Experiments were conducted to determine the role of kerato-epithelin (KE), the product of BIGH3, in the pathogenesis of the diseases. METHODS: KE-15 and KE-2, two rabbit antisera raised against peptides from the 69-364 and 426 - 682 amino acid regions of KE respectively, were used for immunohistology of the corneas obtained after keratoplasty in six CDLI patients, three CDGGI patients, and one CDA patient. RESULTS: The nonamyloid deposits observed in CDGGI stained intensively with KE-15 and KE-2, whereas the amyloid deposits in all analyzed CDLI corneas reacted to KE-2 but not to KE-15. In the CDA cornea, where amyloid and nonamyloid inclusions were present, positive staining with both antisera was observed. CONCLUSIONS: Pathologic amyloid and nonamyloid deposits observed in CDLI, CDGGI-, and CDA-affected corneas are caused by KE accumulation. Different staining patterns of amyloid and nonamyloid deposits observed with antibodies against the amino and carboxyl termini of KE suggest that two mechanisms of KE misfolding are implicated in the pathogenesis of 5q31-linked corneal dystrophies.


Subject(s)
Chromosomes, Human, Pair 5 , Corneal Dystrophies, Hereditary/etiology , Extracellular Matrix Proteins , Genetic Linkage , Neoplasm Proteins/physiology , Transforming Growth Factor beta/physiology , Amyloid/metabolism , Animals , Blotting, Western , Cells, Cultured , Cornea/metabolism , Corneal Dystrophies, Hereditary/metabolism , Corneal Dystrophies, Hereditary/pathology , Corneal Dystrophies, Hereditary/surgery , DNA Primers/chemistry , Fibroblasts/metabolism , Humans , Immunoenzyme Techniques , Keratoplasty, Penetrating , Mutation, Missense , Peptide Fragments , Rabbits
5.
Am J Hum Genet ; 62(2): 320-4, 1998 Feb.
Article in English | MEDLINE | ID: mdl-9463327

ABSTRACT

Mutations in the BIGH3 gene on chromosome 5q31 cause four distinct autosomal dominant diseases of the human cornea: granular (Groenouw type I), Reis-Bücklers, lattice type I, and Avellino corneal dystrophies. All four diseases are characterized by both progressive accumulation of corneal deposits and eventual loss of vision. We have identified a specific recurrent missense mutation for each type of dystrophy, in 10 independently ascertained families. Genotype analysis with microsatellite markers surrounding the BIGH3 locus was performed in these 10 families and in 5 families reported previously. The affected haplotype could be determined in 10 of the 15 families and was different in each family. These data indicate that R555W, R124C, and R124H mutations occurred independently in several ethnic groups and that these mutations do not reflect a putative founder effect. Furthermore, this study confirms the specific importance of the R124 and R555 amino acids in the pathogenesis of autosomal dominant corneal dystrophies linked to 5q.


Subject(s)
Chromosomes, Human, Pair 5 , Corneal Dystrophies, Hereditary/genetics , Point Mutation , Chromosome Mapping , Corneal Dystrophies, Hereditary/classification , Exons , Genes, Dominant , Genetic Linkage , Genetic Markers , Haplotypes , Humans , Introns , Polymerase Chain Reaction , Polymorphism, Single-Stranded Conformational
6.
Nat Genet ; 15(3): 247-51, 1997 Mar.
Article in English | MEDLINE | ID: mdl-9054935

ABSTRACT

Granular dystrophy Groenouw type I (CDGG1), Reis-Bücklers (CDRB), lattice type I (CDL1) and Avellino (ACD) are four 5q31-linked human autosomal dominant corneal dystrophies. Clinically, they show progressive opacification of the cornea leading to severe visual handicap. The nature of the deposits remains unknown in spite of amyloid aetiology ascribed to the last two. We generated a YAC contig of the linked region and, following cDNA selection, recovered the beta ig-h3 gene. In six affected families we identified missense mutations. All detected mutations occurred at the CpG dinucleotide of two arginine codons: R555W in one CDGG1, R555Q in one CDRB, R124C in two CDL1 and R124H in two ACD families. This suggests, as the last two diseases are characterized by amyloid deposits, that R124 mutated kerato-epithelin (the product of beta ig-h3) forms amyloidogenic intermediates that precipitate in the cornea. Our data establish a common molecular origin for the 5q31-linked corneal dystrophies.


Subject(s)
Chromosomes, Human, Pair 5 , Corneal Dystrophies, Hereditary/genetics , Extracellular Matrix Proteins , Neoplasm Proteins/genetics , Point Mutation , Transforming Growth Factor beta , Alternative Splicing , Base Sequence , Cells, Cultured , Chromosome Mapping , Chromosomes, Artificial, Yeast , Cornea/metabolism , DNA Primers , Dinucleoside Phosphates , Exons , Genes, Dominant , Genetic Linkage , Humans , Introns , Molecular Sequence Data , Polymerase Chain Reaction , Skin/metabolism
7.
Eur J Hum Genet ; 4(4): 214-8, 1996.
Article in English | MEDLINE | ID: mdl-8875187

ABSTRACT

Granular Groenouw type I (CDGG1) and lattice type 1 (CDL1) corneal dystrophies are two distinct potentially blinding conditions. These two entities were recently mapped to a region on chromosome 5q. We have investigated 2 families of Swiss origin with CDGG1 and CDL1 by linkage analysis. Our data show a maximum lod score of 5.38 at theta = 0.00 for marker D5S393 in CDL1 and 4.17 at theta = 0.00 for D5S658 in CDGG1. When combined, these families show a maximum low score of 9.22 for D5S393 at theta = 0.00. This confirms previous reports. Furthermore, we describe a recombination centromeric to D5S399 in a member of the CDL1 family. Haplotype analysis in the 4 branches of the CDGG1 family demonstrated a common chromosomal region including D5S393 and D5S399 in all the affected members. By combining our data with previously reported mapping information and assuming that CDGG1 and CDL1 are allelic manifestations of the same gene, we can refine the location of the CDGG1/CDL1 gene to a 1-cM region on chromosome 5q. Using candidate genes in the 5q22-q32 interval, we investigated the possibility that mutations in the SPARC or LOX genes cause these corneal diseases. Several recombinations occurred between these two genes and CDGG1/CDL1 in our 2 families, thus excluding this hypothesis.


Subject(s)
Chromosomes, Human, Pair 5/genetics , Corneal Dystrophies, Hereditary/genetics , Alleles , Corneal Dystrophies, Hereditary/classification , Female , Genetic Linkage , Genetic Markers , Haplotypes , Humans , Male , Models, Genetic , Osteonectin/genetics , Pedigree , Protein-Lysine 6-Oxidase/genetics , Recombination, Genetic , Switzerland
8.
Genomics ; 30(2): 141-8, 1995 Nov 20.
Article in English | MEDLINE | ID: mdl-8586412

ABSTRACT

The human TAX-1 gene (HGMW-approved symbol TAX1) is located on chromosome 1 (1q32.1) and encodes the neuronal cell adhesion molecule TAG-1/axonin-1. The gene product, termed TAG-1 in the rat and axonin-1 in the chicken, is composed of six immunoglobulin (Ig)-like and four fibronectin type III (FNIII)-like domains. It is found predominantly on the axons of particular nerve fiber tracts during neural development, and it has been demonstrated to function as a potent substratum for neurite outgrowth in vitro. Here we report the cloning and structural characterization of the TAX-1 gene. The transcribed region of the TAX-1 gene extends over about 40 kb. Like its chicken homologue, the human TAX-1 gene consists of 23 exons. Two GT/CA microsatellites were localized in the first intron; a polymorphism was found for one of them. Reporter gene analysis with serially truncated fragments of the 5'-flanking region indicated that a 164-bp fragment located immediately upstream of the putative transcription initiation site was sufficient to function as a basal promoter.


Subject(s)
Cell Adhesion Molecules, Neuronal/genetics , Promoter Regions, Genetic , Animals , Base Sequence , Chickens , Chromosome Mapping , Cloning, Molecular , Contactin 2 , Exons , Genes, Reporter , Humans , Introns , Molecular Sequence Data , Repetitive Sequences, Nucleic Acid , Sequence Homology, Nucleic Acid
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