A 295-kDA intermediate filament-associated protein in radial glia and developing muscle cells in vivo and in vitro.

The RC2 antibody is frequently used to label mouse radial glial cells in all parts of the nervous system where neuronal migration occurs during embryonic and early postnatal life. The antigen recognized by this antibody still needs to be identified. We have characterized further its localization in vivo, its expression and subcellular localization in vitro, as well as its molecular nature. Histologic investigations of whole mouse embryos reveal an equally intense expression of RC2 immunostaining in radial glial cells in brain and spinal cord and in skeletal muscle. In glial cells cultures, the RC2 antibody recognizes an epitope located on the glial cytoskeleton and identified as an intermediate filament associated protein (IFAP) at the ultrastructural level. RC2 immunostaining in those cells is strongly dependent on the presence of a serum-derived activity. Serum-removal causes a decrease of the staining while adding serum back to the cells induces reexpression of RC2 immunoreactivity. By Western blotting, we find that in intermediate filament (IF) preparations obtained from cultured cerebellar glia, the RC2 antibody recognizes a 295-kDa protein whose expression is also dependent on the presence of serum in culture medium. In developing muscle cells, RC2 immunostaining is observed from the myoblast stage and disappears after complete myotube fusion. Both in vivo and in vitro, staining is first seen as a loose capping around myoblasts nuclei and progressively concentrates into Z-disks in association with the muscle IF protein desmin. The RC2 antibody also recognizes a 295-kDa protein band in muscle tissue protein extracts. Thus, the RC2 antibody recognizes a developmentally regulated cytoskeletal protein that is expressed, like other previously identified IFAPs, by cells of the glial and myogenic lineages and whose expression in vitro seems to be controlled by a signaling mechanism known to modulate astroglial morphology.

Radial glia phenotype: origin, regulation, and transdifferentiation.

Radial glial cells play a major guidance role for migrating neurons during central nervous system (CNS) histogenesis but also play many other crucial roles in early brain development. Being among the earliest cells to differentiate in the early CNS, they provide support for neuronal migration during embryonic brain development; provide instructive and neurotrophic signals required for the survival, proliferation, and differentiation of neurons; and may be multipotential progenitor cells that give rise to various cell types, including neurons. Radial glial cells constitute a major cell type of the developing brain in numerous nonmammalian and mammalian vertebrates, increasing in complexity in parallel with the organization of the nervous tissue they help to build. In mammalian species, these cells transdifferentiate into astrocytes when neuronal migration is completed, whereas, in nonmammalian species, they persist into adulthood as a radial component of astroglia. Thus, our perception of radial glia may have to change from that of path-defining cells to that of specialized precursor cells transiently fulfilling a guidance role during brain histogenesis. In that respect, their apparent change of phenotype from radial fiber to astrocyte probably constitutes one of the most common transdifferentiation events in mammalian development.

Identification of PSF, the polypyrimidine tract-binding protein-associated splicing factor, as a developmentally regulated neuronal protein.

The polypyrimidine tract-binding protein-associated splicing factor (PSF), which plays an essential role in mammalian spliceosomes, has been found to be expressed by differentiating neurons in developing mouse brain. The sequence of a fragment of mouse PSF was found to be remarkably similar to that of human PSF. Both the expression of PSF mRNA in cortex and cerebellum and PSF immunoreactivity in all brain areas were high during embryonic and early postnatal life and almost disappeared in adult tissue, except in the hippocampus and olfactory bulb where various neuronal populations remained PSF-immunopositive. Double-labeling experiments with anti-PSF antibody and anti-neurofilaments or anti-glial fibrillary acidic protein antibodies on sections of cortex, hippocampus, and cerebellum indicate that PSF is expressed by differentiating neurons but not by astrocytic cells. In vitro, mouse PSF was found to be expressed by differentiating cortical and cerebellar neurons. Radial glia or astrocyte nuclei were not immunopositive; however, oligodendrocytes differentiating in vitro were found to express PSF. The restricted expression of PSF suggests that this splicing factor could be involved in the control of neuronal-specific splicing events occurring at particular stages of neuronal differentiation and maturation.