Kyoji Ohyama

Immunohistochemical localization of neurocan and L1 in the formation of thalamocortical pathway of developing rats

We used immunohistochemistry to examine possible molecular interactions between the subplate and growing thalamocortical axons in rat fetuses. In the cortical anlage of embryonic day 16 (E16), the subplate first appeared below the cortical plate. Among chondroitin sulfate proteoglycans, phosphacan was uniformly distributed throughout the cortical wall, whereas neurocan was localized only in the subplate at E16. Neural cell adhesion molecules, NCAM‐H, TAG‐1, and L1, were detected in the cortical anlage. Both cortical neurons and growing axons were diffusely immunopositive for NCAM‐H, and TAG‐1 immunoreactivity was found on immature neurons and cortical efferent axons but not on thalamocortical axons. L1 immunoreactivity was specifically localized on the growing thalamocortical axons. When the locations of neurocan and L1 were compared in the developing cortex, L1‐bearing axons were found to extend to neurocan‐immunopositive regions; neurocan immunoreactivity was intense in the subplate at E16, when small numbers of L1‐immunoreactive thalamocortical axons began to invade the cortex. At E17, many L1‐positive axons were observed in the subplate that expressed neurocan specifically. Double immunostaining showed that L1‐positive axons and neurocan immunoreactivity overlapped in the subplate at E17. After E18, neurocan expression gradually extended to the lower part of the cortical plate; it extended to the entire cortex by E21, 1 day before birth. By E21, L1‐bearing axons had invaded the lower part of the cortical plate. The present study demonstrated that the neurocan expression precedes growth of L1‐bearing thalamocortical afferent fibers. Because neurocan can bind to L1 molecule in vitro, these results suggest that neurocan and L1 play some important roles in pathfinding of the thalamocortical afferent fibers during rat corticogenesis. J. Comp. Neurol. 382:141‐152, 1997.

COORDINATE EXPRESSION OF L1 AND 6B4 PROTEOGLYCAN/PHOSPHACAN IS CORRELATED WITH THE MIGRATION OF MESENCEPHALIC DOPAMINERGIC NEURONS IN MICE

Mesencephalic dopaminergic (DA) neurons of mice are generated from embryonic day 10 to 12 (E10-12) in the ventricular zone of the mesencephalon. They first migrate toward the ventral mesencephalon, and then turn laterally, or tangentially, in the basal part of the mesencephalon. With immunohistochemical analysis of E10-E15 ICR mice, we found that cell adhesion molecule L1 was transiently expressed on the median part of tangential fibers coincident with the lateral migration of DA neurons from E11 to E13, when neurons move along the tangential fibers toward their final destinations: the reticular formation, the substantia nigra pars compact, and the ventral tegmental area. While L1 expression was not observed in DA neurons, they expressed a chondroitin sulfate proteoglycan, 6B4 proteoglycan/phosphacan, which has been shown to bind to L1/Ng-CAM in vitro. These results suggest that the heterophilic interaction between 6B4 proteoglycan on the neurons and L1 on the fibers is involved in the lateral migration of mesencephalic DA neurons in mice.

Cyclin E enhances P53-mediated transactivation

Plasmids expressing G1 and G2 cyclins were introduced into the Saos‐2 cell system monitoring p53‐mediated transactivation [(1993) Oncogene 8, 543]. Cyclin E, but not other cyclins, enhanced the p53‐mediated transactivation about 2‐fold. Co‐transfection of a CDK2 expression plasmid caused a 30% increase in the extent of the p53‐mediated transactivation. Moreover, the transfected p53 protein became phosphorylated coordinately with the enhanced transactivation. The close correlation between transactivation and p53 phosphorylation suggests that phosphorylation is involved in positive regulation for the transactivation by p53.

Mutual dependence of murine fetal cutaneous regeneration and peripheral nerve regeneration.

Mammalian fetal cutaneous wounds made at certain developmental stages show complete regeneration. It is reported that wound healing in both adult and fetal skin is disrupted by denervation. Furthermore, fetal cutaneous regeneration has unique aspects such as epidermal wrinkle texture regeneration and dermal regeneration that depend on developmental stage. Therefore, we have examined the relationship of fetal cutaneous regeneration with denervation. We made cutaneous wounds on fetal mice at various developmental time points including embryonic days (E)13, E15, and E17, and compared the regenerating patterns of peripheral nerves in the skin. We found that when the fetuses are wounded at an early stage of development, peripheral nerves regenerate quicker than at later stages of development when peripheral nerve regeneration is delayed. Next, we denervated the intercostal nerves and made wounds at the denervated sites on E13 and E15. We found that epidermal wrinkling and dermal regeneration were disrupted by denervation. These findings indicate that components of fetal cutaneous regeneration and peripheral nerve regeneration are mutually dependent.

Allele-specific activation of the c-myc gene in an atypical Burkitt's lymphoma carrying the t(2;8) chromosomal translocation 250 kb downstream from c-myc

The genetic structure and regulation of the c-myc gene was comprehensively studied for the first time in Burkitts lymphoma with t(2;8) translocation. In a Burkitts lymphoma cell line, KOBK101, the immunoglobulin kappa-encoding gene on chromosome 2, accompanied by its enhancer, was translocated to the pvt-1 locus located about 250 kb downstream from c-myc on chromosome 8. Only the c-myc allele on the translocated chromosome carried aberrant SalI and KpnI sites in the first intron, so the two c-myc alleles and their transcripts were analyzed separately. The c-myc allele on the untranslocated chromosome conserved the normal c-myc sequence and was transcriptionally silent. In contrast, the c-myc allele on the translocated chromosome was actively transcribed at three- to fivefold higher levels, as compared with non-malignant B-cell lines. Additionally, it carried predominant multiple mutations consisting of 64 nucleotide substitutions, three short deletions, and a one-base insertion, most of which clustered in the first exon and intron. The 24-base deletion in the first intron completely overlapped the binding site of a putative negative transcriptional factor of the 138-kDa phosphoprotein, MIF. Thus, the multiple mutations and the deregulated, allele-specific expression of c-myc were associated with the chromosomal translocation in cis. Together activation by the long-distance immunoglobulin kappa enhancer, and the alleviation of negative regulation by the mutations, seemed to cause the allele-specific activation of c-myc.

1142 Immunohistochemical localization of neural cell adhesion molecules in the developing rat cerebral cortex

1142 IMMUNOHISTOCHEMICALLOCALIZATIONOFNEURALCELLADHESION MOLECULESINTHEDEYELOPINGRATCEREBRALCORTEX. TETSUYAFUKUDA ‘.HITOSHI KAWANO ’ _ KYOJI OHYAMA ’ _ KOKI KAWAMURA ’ _ ATSUHIKO OOHIRA ’ _ AND KOSEI TAKEUCHI ” ’ Department of Anatomv. Keio Universitv. School of Medicine. Tokvo 160. ’ Denartment of Perinatologv and Neuroulycoscience. Institute for Developmental Research. Kasugai . Aichi 480-03. and 3 DeDartment of Exnerimental Biologv. Tokvo Metropolitan Institute of Gerontolouvv. Tokvo 173. Janan Mammahan cerebral cortex has a highly integrated cytoarchitecture which consists of a number of correctly arranged neurons and fine fiber networks This neural construction is formed through the development and various cell surface molecules play important roles for the formation of the cerebral cortex. In this study, localization of neural cell adhesion molecules belongmg to Ig superfamily, such as NCAM, L 1 and TAG1, was exammed immunohistochemically m the developmg rat cerebral cortex. Immunoreactions for embryonic type of NCAM (highly polysialylated NCAM, NCAM-H) and TAG-l were detected in the cortical anlage as early as embryonic day 14, whereas L I -immunoreactivity first occurred at embryomc day 16 Although NCAM-immunoreactivity is positive until the adulthood, immunoreactions for NCAM-H, Ll and TAG-l decreased remarkably m the mature bram The physiological roles of these molecules m the formation of cerebral cortex are discussed