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A M Goffinet

Publications and source records attributed to A M Goffinet.

At least 37 records · Page 2Linked to original sources
PubMed1999

Reelin, the extracellular matrix protein deficient in reeler mutant mice, is processed by a metalloproteinase.

Reelin is the extracellular protein defective in reeler mice. It is believed that reelin acts via the extracellular matrix to influence the development of nearby neurons, but the mechanism remains thus far unknown. In the present work, we present in vivo and in vitro evidence that reelin is cleaved. This processing did not occur in Relnrl-Orl mutant mice in which reelin is not secreted and was prevented in explant cultures by brefeldin treatment, suggesting that it takes place extracellularly or in a postendoplasmic reticulum compartment. Reelin cleavage was inhibited by zinc chelators known to inhibit metalloproteinases but was unaffected by inhibitors of serine, cysteine, or aspartate proteinases. Furthermore, reelin cleavage was insensitive to inhibitors of matrixins, neprilysin, meprin, and peptidyl dipeptidase A, suggesting that the processing enzyme belongs to a different enzyme family. This enzyme and the physiological meaning of reelin processing remain to be characterized further.

Animals↗
PubMed1999

Evolutionarily conserved, alternative splicing of reelin during brain development.

Reelin is the protein defective in reeler mutant mice and plays a pivotal role in brain development. However, some uncertainties remain about the relationship between reelin and the reeler phenotype. It is generally believed that reelin, secreted by specific neuronal types such as Cajal-Retzius cells, acts at short distance via the extracellular matrix on target neurons, the response of which requires the Dab1 gene product. However, the pattern of reelin expression in some structures such as olfactory bulb, retina, and spinal cord suggests that the protein might be endowed with different functions. In the present study, we identify two uncommon, evolutionarily conserved splicing events in the 3' part of the transcript that result in different forms of the protein. First, a 6-nucleotide, brain-specific microexon is skipped in about 10% of reelin RNA. In addition, an alternative polyadenylation event involving 10-25% of reelin mRNA results in secretion of a truncated protein lacking the terminal, highly basic stretch. This alternative reelin is generally expressed in the same cells as the major form, but is almost undetectable in retina and spinal cord. Both alternative splicing events are present in mouse, rat, and man, suggesting that the corresponding reelin forms are functionally important.

Animals↗
PubMed1998

A new view of early cortical development.

Recently, several genes that regulate the development of the cerebral cortex and are potential pharmacological targets have been cloned. Reelin, an extracellular matrix glycoprotein secreted by Cajal-Retzius cells in the marginal zone, instructs the radial organization of the cortical plate. The response of cortical plate cells to reelin requires the tyrosine kinase adaptor disabled-1 (Dab1). Cyclin-dependent kinase 5 and its activator p35 are necessary for the development of the cortical plate, probably at a later stage than reelin/Dab1. The transcription factor Tbr-1 is essential for differentiation of preplate and Cajal-Retzius cells and for formation of thalamocortical connections, while D1x-1/2 are required for tangential migration. Some neurotrophin systems such as neurotrophin 4, brain-derived neurotrophic factor, and neuregulin and its receptor ErbB are also thought to assist in the regulation of cortical development. In addition, a few genes implicated in human cortical dysplasias have been characterized. LIS1 encodes a protein related to platelet-activating factor acetyl hydrolase that is defective in lissencephaly-1 of the Miller-Dieker type, while the double cortex malformation is related to mutations of a new gene dubbed doublecortn.

Animals↗
PubMed1998

Cyclin-dependent kinase 5-deficient mice demonstrate novel developmental arrest in cerebral cortex.

The cerebral cortex of mice with a targeted disruption in the gene for cyclin-dependent kinase 5 (cdk5) is abnormal in its structure. Bromodeoxyuridine labeling reveals that the normal inside-out neurogenic gradient is inverted in the mutants; earlier born neurons are most often found superficial to those born later. Despite this, the early preplate layer separates correctly and neurons with a normal, pyramidal morphology can be found between true marginal zone and subplate. Consistent with their identity as layer VI corticothalamic neurons, they can be labeled by DiI injections into thalamus. The DiI injections also reveal that the trajectories of the cdk5(-/-) thalamocortical axons are oblique and cut across the entire cortical plate, instead of being oriented tangentially in the subcortical white matter. We propose a model in which the cdk5(-/-) defect blocks cortical development at a heretofore undescribed intermediate stage, after the splitting of the preplate, but before the migration of the full complement of cortical neurons.

Animals↗
PubMed1998

The role of the first postmitotic cortical cells in the development of thalamocortical innervation in the reeler mouse.

In the mutant mouse reeler, the tangential distribution of thalamocortical fibers is essentially normal, even though neurons of the cortical plate accumulate below the entire early-born preplate population (Caviness et al., 1998). This seems incompatible with the hypothesis that cells of the subplate (the lower component of the preplate in normal mammals) form an axonal scaffold that guides thalamic fibers and act as temporary targets for them (Blakemore and Molnár, 1990, Shatz et al., 1990). We used carbocyanine dyes to trace projections in wild-type and reeler mice between embryonic day 13 and postnatal day 3. Preplate formation and early extension of corticofugal fibers to form a topographic array are indistinguishable in the two phenotypes. So too are the emergence of thalamic axons in topographic order through the primitive internal capsule, their meeting with preplate axons, and their distribution over the preplate scaffold. Distinctive differences appear after the cortical plate begins to accumulate below the preplate of reeler, causing the preplate axons to form oblique fascicles, running through the cortical plate. Thalamic axons then pass through the plate within the same fascicles and accumulate in the "superplate" layer for approximately 2-3 d, before defasciculating and plunging down to terminate deep in the cortical plate, creating the curious "looping" pattern seen in the adult. Thus, thalamocortical innervation in reeler follows the same algorithm of development but in relation to the misplaced population of early-born neurons. Far from challenging the theory that preplate fibers guide thalamic axons, reeler provides strong evidence for it.

Animals↗
PubMed1998

Prenatal development of reelin-immunoreactive neurons in the human neocortex.

Reelin, the protein defective in reeler mutant mice, is a secreted glycoprotein involved in the architectonic development of the central nervous system, more particularly in the development of neocortical lamination. In mice, reelin mRNA and protein expression are most robust in horizontal neurons of the embryonic marginal zone (MZ). By using monoclonal anti-reelin antibodies (de Bergeyck et al. [1998] J. Neurosci. Methods), the morphology and evolution of reelin-expressing neurons were studied in the MZ of the prenatal human neocortex. At 11 gestational weeks (GW), the MZ contained a single layer of reelin-positive mono- or bipolar horizontal Cajal-Retzius (CR) cells. From 14 GW onward, the subpial granular layer (SGL) invaded the MZ, forming a transient layer of undifferentiated, initially reelin-negative granule cells. In parallel to the emergence of the SGL and the morphological differentiation of the CR cells, a second population of reelin-positive cells appeared within the SGL. These cells, termed CR-like cells, were intermediate in size and shape between the CR cells and SGL granule cells. Between 16 GW and 24 GW, the packing density of the reelin-producing cells remained remarkably stable, despite the continuous growth of the cortical surface. During this period, CR cells settled progressively deeper within the MZ, although they remained in contact with the pial surface through radially ascending processes. Most CR cells disappeared at around 27 GW, in parallel with the dissolution of the SGL. During the last weeks of gestation, reelin was expressed by a few medium-sized, often horizontal neurons. These observations show that different neuronal populations in the human MZ express reelin and suggest that a possible function of the SGL is to supply reelin-producing cells through a gradual transformation of reelin-negative precursor cells into reelin-immunoreactive CR-like cells, thus coping with the protracted neurogenesis and dramatic surface expansion of the human neocortex.

Calbindin 2↗
PubMed1998

A panel of monoclonal antibodies against reelin, the extracellular matrix protein defective in reeler mutant mice.

Reelin, the extracellular matrix protein defective in reeler mutant mice, plays a key role during brain development. We therefore raised antibodies directed against various reelin epitopes in order to facilitate biochemical and cell biological studies of this important molecule. Homozygous reeler mice with a large deletion of most of the reelin gene were immunized with fusion proteins and carrier-coupled peptides corresponding to parts of the reelin sequence. Monoclonal antibodies were produced using classical procedures, screened using ELISA and-or western blot prepared with the antigen, and tested by immunohistochemistry and immunoprecipitation assays to detect endogenous reelin. The labeling of Cajal-Retzius cells in the embryonic mouse telencephalon was selected as criterion for positivity in immunohistochemistry. A total of 11 monoclonal antibodies were obtained, providing useful additions to the widely used antibody CR-50. Five are directed against the N-terminal part of reelin, among which three recognize the region that has significant similarity with F-spondin, and two are specific for hinge region located downstream from the F-spondin similarity region and upstream from the reelin repeats. Six antibodies are directed against the C-terminal part of reelin, among which one anti-peptide antibody recognizes the highly basic C-terminal segment. Antibodies against the N-terminal region stain well in immunohistochemistry. By comparison, the labeling of embryonic Cajal-Retzius cells with antibodies directed against the C-terminal region is weaker, suggesting that this part of the molecule might be modified or not be as readily accessible in the tissue as the N-terminus.

Amino Acid Sequence↗
PubMed1997

Genomic organization of the mouse reelin gene.

Reelin is the protein defective in reeler mice, an extensively studied model of brain development. The reelin gene (symbol Reln) codes for a protein of the extracellular matrix that contains eight successive repeats of 350 to 390 amino acids. In this work, we describe the genomic structure of the mouse reelin gene and the 5'-flanking genomic DNA sequences. The reelin gene is composed of 65 exons spread over approximately 450 kb of genomic DNA. We identified different reelin transcripts, formed by alternative splicing of a microexon as well as by use of two different polyadenylation sites. All splice sites conform to the GT-AG rule, except for the splice donor site of intron 30, which is GC instead of GT. A processed pseudogene is present in intron 42. Its nucleotide sequence is 86% identical to the sequence of the rat RDJ1 cDNA, which codes for a DnaJ-like protein of the Hsp40 family. Comparison of 8 intron positions in mouse and human reelin genes reveals a highly conserved genomic structure, suggesting a similar structure of the whole gene in both species. We identified two transcription start sites embedded within a CpG. The promoter region contains putative recognition sites for the transcription factors Sp1 and AP2 but lacks TATA and CAAT boxes. The presence of tandemly repeated regions in the Reelin protein suggests that gene duplication events occurred during evolution. By comparison of the amino acid sequences of the eight repeats and the positions of introns, we suggest a model for the evolution of the repeat coding portion of the reelin gene from a putative ancestral minigene.

Alternative Splicing↗
PubMed1997

A truncated Reelin protein is produced but not secreted in the 'Orleans' reeler mutation (Reln[rl-Orl]).

Reelin is the protein defective in reeler mutant mice [I. Bar, C. Lambert de Rouvroit, I. Royaux, D.B. Krizman, C. Dernoncourt, D. Ruelle, M.C. Beckers, A.M. Goffinet, A YAC contig containing the reeler locus with preliminary characterization of candidate gene fragments, Genomics 26 (1995) 543-549; G. D'Arcangelo, G.G. Miao, S.C. Chen, H.D. Soares, J.I. Morgan, T. Curran, A protein related to extracellular matrix proteins deleted in the mouse mutant reeler, Nature 374 (1995) 719-723; S. Hirotsune, T. Takahara, N. Sasaki, K. Hirose, A. Yoshiki, T. Ohashi, M. Kusakabe, Y. Murakami, M. Muramatsu, S. Watanabe, K. Nakao, M. Katsuki, Y. Hayashizaki, The reeler gene encodes a protein with an EGF-like motif expressed by pioneer neurons, Nature Genet. 10 (1995) 77-83]. In the Orleans allele of reeler (symbol: Reln[rl-Orl]), a 220 nucleotide deletion is present in the 3' region of the Reelin message, resulting in a frame shift with production of a predicted protein amputated from its C-terminal amino acids. In this study, we first show that the predicted truncated protein indeed exists in Orleans reeler mice, using several anti-Reelin antibodies. Three antibodies are directed against epitopes located in the N-terminal region of the protein, namely: monoclonal antibody CR-50 [M. Ogawa, T. Miyata, K. Nakajima, K. Yagyu, M. Seike, K. Ikenaka, H. Yamamoto, K. Mikoshiba, The reeler gene-associated antigen on Cajal-Retzius neurons is a crucial molecule for laminar organization of cortical neurons, Neuron 14 (1995) 899-912] (epitope region between Reelin residues 251-407), monoclonal antibody G10 (epitope located between amino acids 199 and 244) and the polyclonal antipeptide rp4 (positions 381-399). A fourth antibody, antipeptide rp5, reacts with the C-terminal (3443-3461) Reelin sequence. In normal embryos, all four antibodies stained cells in the marginal zone with features of Cajal-Retzius cells. While N-terminal specific antibodies detected Reelin immunoreactivity in mouse embryos homozygous for the reeler-Orleans mutation, no staining was obtained with the rp5 antibody, showing the presence of a truncated protein. Moreover, although Reelin could be detected at the surface of living Cajal-Retzius cells of normal mice, it was not revealed after vital staining of embryonic cortex from Orleans reeler mice. These results indicate that the C-terminal region of Reelin is essential for its secretion and suggest that the Orleans reeler phenotype is due to defective Reelin secretion rather than to secretion of an inactive protein.

Amino Acid Sequence↗
PubMed1997

Reln(rl-Alb2), an allele of reeler isolated from a chlorambucil screen, is due to an IAP insertion with exon skipping.

The reeler Albany2 mutation (Reln(rl-Alb2) in the mouse is an allele of reeler isolated during a chlorambucil mutagenesis screen. Homozygous animals had drastically reduced concentrations of reelin mRNA, in which an 85-nt exon was absent. At the genomic level, the mutation was shown to be due to an intracisternal A-particle insertion leading to exon skipping. This appears to be the first observation of retrotransposon insertion during chlorambucil mutagenesis.

Alleles↗
PubMed1997

Abnormalities in the cerebellum and brainstem in homozygous lurcher mice.

The lurcher mutation induces Purkinje cell degeneration in heterozygous mice, and neonatal death in homozygous animals. Using the D6Mit16 Simple Sequence Length Polymorphic marker in F2 hybrids between AKR +/+ mice and B6+/Lc mice, homozygous lurcher fetuses and newborns as well as heterozygous and normal littermates were identified, and their brain morphology was analysed. In homozygous lurcher embryos at embryonic day 18 and neonates the cerebellum was hypotrophic, particularly in the posterior half. Purkinje cells were smaller in the whole cerebellum and showed a maturational delay. Calretinin-positive cells were less frequently observed in the depth of the vermis than in normal mice. Both Purkinje cells and the vermal calretinin-positive cells were more abnormal in fetuses at day 19 and newborn mutants than one day earlier. An abnormal number of pycnotic cells were observed in the cerebellum, especially in newborn mutants. Brainstem abnormalities were characterized by abnormal curvature, caudal displacement of the pontine gray nuclei which were located caudally along the ventral border of the superior olivary complex, a drastic decrease in Purkinje cell axons in all the vestibular nuclei and the presence of dystrophic processes in at least two calbindin-positive cell groups of the dorsal pontine region. These results show that the mutation, which is semidominant in Purkinje cells, is recessive in other cell groups of the cerebellum and brainstem. They reveal that the sequence leading to Purkinje cell death appears to be similar in homozygous and heterozygous mice, although occurring earlier and worsening more quickly in the former. Lastly, they confirm the absence of effect of the mutation on the neurons of the inferior olivary complex.

Animals↗
PubMed1997

Aberrant splicing of a mouse disabled homolog, mdab1, in the scrambler mouse.

Although accurate long-distance neuronal migration is a cardinal feature of cerebral cortical development, little is known about control of this migration. The scrambler (scm) mouse shows abnormal cortical lamination that is indistinguishable from reeler. Genetic and physical mapping of scm identified yeast artificial chromosomes containing an exon of mdab1, a homolog of Drosophila disabled, which encodes a phosphoprotein that binds nonreceptor tyrosine kinases. mdab1 transcripts showed abnormal splicing in scm homozygotes, with 1.5 kb of intracisternal A particle retrotransposon sequence inserted into the mdab1 coding region in antisense orientation, producing a mutated and truncated predicted protein. Therefore, mdab1 is most likely the scm gene, thus implicating nonreceptor tyrosine kinases in neuronal migration and lamination in developing cerebral cortex.

Animals↗
PubMed1997

Reelin mRNA expression during mouse brain development.

Using in situ hybridization, expression of the mRNA for reelin, the gene most probably responsible for the reeler trait, was studied during mouse brain development, from embryonic day 13 to maturity. The highest level of expression was found in Cajal-Retzius neurons, while a high signal was also seen in the olfactory bulb, the external granular layer of the cerebellum and, particularly at early developmental stages, in hypothalamic differentiation fields, tectum and spinal cord. A moderate to low level of expression was found in the septal area, striatal fields, habenular nuclei, some thalamic nuclei, particularly the lateral geniculate, the retina and some nuclei of the reticular formation in the central field of the medulla. Paradoxically, no reelin expression was detected in radial glial cells, the cortical plate, Purkinje cells, inferior olivary neurons and many other areas that are characteristically abnormal in reeler mutant mice. Together with other preliminary studies, the present observations suggest that the action of reelin is indirect, possibly mediated by the extracellular matrix. Most of the data can be explained by supposing that reelin is a cell-repulsive molecule which prevents migrating neurons from invading reelin-rich areas, and thus facilitates the deployment of radial glial cell processes and the formation of early architectonic patterns.

Animals↗