Biomedical subjects

T D Raabe

Publications and source records attributed to T D Raabe.

9 recordsLinked to original sources

PubMed2005

Stimulation of acetylcholine receptor transcription by neuregulin-2 requires an N-box response element and is regulated by alternative splicing.

The neuregulin (Nrg) family of growth/differentiation factors is encoded by at least four genes in the mammalian genome: nrg-1, nrg-2, nrg-3 and nrg-4. Nrg-1 and Nrg-2 share the highest homology within the family, and the primary RNA transcripts from their encoding genes are subjected to extensive alternative splicing. Although little is known about the biological function of Nrg-2-4, their structural similarity with Nrg-1 suggests that they could account for some of the activities presently attributed to Nrg-1. Thus, at the neuromuscular junction Nrg-1 has been a favored candidate for the signal that activates selective acetylcholine receptor (AChR) transcription in synaptic myonuclei. However, we have recently shown that like Nrg-1, Nrg-2 can also activate AChR transcription in cultured myotubes and accumulates at the synaptic site. Synapse-specific and Nrg-1-induced AChR transcription require an enhancer sequence, the N-box, which is also mutated in some patients with congenital myasthenia gravis. Here, we show that Nrg-2-induced AChR transcription requires an N-box motif and is regulated by alternative splicing. We also show that unique Nrg-2 isoforms are differentially distributed between spinal cord and skeletal muscle, the tissues that harbor the cellular components of the neuromuscular synapse.

Alternative Splicing↗

PubMed1999

Neuregulins and ErbB receptors in cultured neonatal astrocytes.

Neuregulins (NRGs) are a family of growth factors involved in signaling between neurons and glial cells of the peripheral and central nervous system. NRGs are synthesized and secreted by a number of cell types including Schwann cells, neurons, and oligodendrocytes. NRG transduction signals are mediated by the erbB family of receptor tyrosine kinases. These NRGs may be important for paracrine or autocrine signaling during development, injury, and the normal functioning of the central nervous system. In this study, we characterize the NRGs and erbB receptors expressed by cultured neonatal rat astrocytes. Using immunoblotting protocols with pan-specific antibodies, we identified eleven NRG molecular weight isoforms from approximately 16 kDa to 105 kDa in cultured neonatal rat astrocytes. Immunocyotchemistry with isoform-specific antibodies revealed the expression of both major isoform families (NRGalpha, NRGbeta). Additionally, astrocyte-conditioned media contained two molecular weight isoforms of NRGs. We detected mRNA expression of NRGalpha and NRGbeta in astrocytes by amplifying mRNA transcripts with reverse transcription polymerase chain reaction. Furthermore, we confirm that cultured astrocytes express all four erbB receptors as detected by immunocytochemical and immunoblotting techniques. These data indicate that astrocytes contain and secrete NRGs.

Animals↗

PubMed1998

Neuregulins in glial cells.

The role of growth factors in controlling the development of glial cells in both the peripheral and central nervous systems has been investigated for a number of years. The recent discovery of a new family of growth factors termed the neuregulins (NRGs) has led to an explosion of information concerning the putative role of these growth factors in the development of Schwann cells (SC), oligodendrocytes (OLG), and astrocytes. Many of these previous studies have focused on the effects of exogenous NRGs on glial cell development and differentiation. We now review the evidence that these glial cells themselves produce NRGs and discuss the major implications of these findings with respect to glial cell development and diseases which affect glial cell function. We also discuss the potential role of endogenous NRGs following neural injury.

Animals↗

PubMed1997

Effect of neu differentiation factor isoforms on neonatal oligodendrocyte function.

Previous studies have suggested that neu differentiation factor (NDF), a member of the neuregulin (NRG) family of growth factors, may regulate the development of PNS and CNS glial cells. There is limited information concerning the potential role of NDF on the development of neonatal (immature) oligodendrocytes (OLG) into adult OLG. We now report the effect of the two major isoform families of NDF (NDF alpha and NDF beta) on the development of cultured rat neonatal OLG. Immunocytochemical and western blot analyses of neonatal OLG using anti-erb-B antibodies revealed that these immature OLG express all four members of NRG (erb-B) receptors. Treatment of neonatal OLG with varying concentrations of either NDF alpha or NDF beta did not have a mitogenic effect on cultured neonatal OLG. Pretreatment of immature OLG with either of the NDF isoforms also did not influence the subsequent mitogenicity of other known OLG mitogens. However, treatment of neonatal OLG with either isoform of NDF influenced the survival of these cells by protecting the cells from apoptosis. Additionally, treatment of neonatal OLG with either NDF alpha or NDF beta resulted in more extensive process formation compared to control, non-treated OLG.

Animals↗

PubMed1997

Neonatal oligodendrocytes contain and secrete neuregulins in vitro.

The factors that influence the development of oligodendrocyte (OLG) progenitors into mature OLGs remain elusive. Recent evidence has suggested that neu differentiation factor (NDF), which is a member of the neuregulin family of growth factors, influences the development of glial cells, including Schwann cells, astrocytes, and OLGs. Neurons are postulated to be the source of neuregulins, because neurons closely interact with these glial cells during development. In this report, we have identified the mRNA for both isoform families of NDF in cultured neonatal (immature) OLGs. We have also demonstrated that cultured neonatal OLGs contain and secrete NDF protein. These data raise the possibility that NDF could be used in an autocrine/paracrine loop by neonatal OLGs during development for survival, proliferation, and/or differentiation.

Animals↗

PubMed1996

Cultured neonatal Schwann cells contain and secrete neuregulins.

Neuregulins have become the focus of intense research due to their putative roles in the etiology of certain cancers and to their evolving roles in PNS and CNS development. Evidence has been presented that neuregulins are present in neuronal cells where they act as mediators of neuron-glial signaling. Consistent with this view, we now report that there is a dramatic differential response of Schwann cells to the two major isoforms of neu differentiation factor (NDF alpha and NDF beta). NDF beta is a potent mitogen, whereas NDF alpha is nonmitogenic for Schwann cells. In addition, we report that Schwann cells contain endogenous NDF as well as the mRNA for both isoforms of NDF. Evidence is also presented that several isoforms of NDF are released from cultured Schwann cells. Our data suggest that in addition to functioning as a neuron-glial mediator, endogenous neuregulins may function in an autocrine/paracrine loop or in a juxtacrine- (cell-to-cell) mediated signal between individual Schwann cells or between Schwann cells and neurons.

Animals↗

PubMed1996

Mechanisms for the maintenance and eventual degradation of neurofilament proteins in the distal segments of severed goldfish mauthner axons.

Cellular mechanisms that might affect the degradation of neurofilament proteins (NFPs) were examined in the distal segments of severed goldfish Mauthner axons (M-axons), which do not degenerate for more than 2 months after severance. Calpain levels, as determined by reactivity to a polyclonal antibody, remained constant for 80 d postseverance in distal segments of M-axons and then declined from 80 to 85 d postseverance. Calpain activity in rat brain, as determined by a spectrophotometric assay, was much higher than calpain activity in control and severed goldfish brain, spinal cord, muscle, or M-axons. Calpain activity was extremely low in M-axons compared with that in all other tissues and remained low for up to 80 d postseverance in distal segments of M-axons. Phosphorylated NFPs, as determined by Stains-All treatment of SDS gels, were maintained for up to 72 d postseverance and then decreased noticeably at 75 d postseverance when NFP breakdown products appeared on silver-stained gels. By 85 d post-severance, phosphorylated NFPs no longer were detected, and NFP breakdown products were the most prominent bands on silver-stained gels. These results suggest that the distal segments of M-axons survive for months after severance, because NFPs are maintained in a phosphorylated state that stabilizes and protects NFPs from degradation by low levels of calpain activity in the M-axon; the distal segments of severed M-axons degenerate eventually when NFPs no longer are maintained in a phosphorylated state and become susceptible to degradation, possibly by low levels of calpain activity in the M-axon.

Animals↗

PubMed1996

Phosphorylation of neurofilament proteins in isolated goldfish Mauthner axoplasm.

The six neurofilament proteins (NFPs) in the goldfish Mauthner axon (M-axon) have molecular sizes of 235, 145, 123, 105, 80, and 60 kDa. To determine if NFPs in the M-axon are phosphorylated, isolated Mauthner axoplasm (M-axoplasm) and a neurofilament-enriched extract (NFE) prepared from M-axoplasm were incubated with 32P, which resulted in the radiolabeling of NFPs as determined by their detection on autoradiograms. Kinase inhibitors directed against cyclic AMP-dependent kinases (PKAs) or cofactor-independent kinases significantly reduced the in vitro phosphorylation of NFPs in NFE, whereas inhibitors directed against protein kinase C did not significantly reduce the in vitro phosphorylation of NFPs in NFE. Experiments using two kinase inhibitors directed against different kinases significantly reduced the in vitro phosphorylation of NFPs in NFE to a greater extent than the reduction produced using any single kinase inhibitor. These data suggest that NFPs in the M-axon are phosphorylated and that the in vitro (and perhaps the in vivo) phosphorylation of NFPs is mediated by PKA and/or cofactor-independent kinases that copurify with NFPs.

Animals↗

PubMed1995

Calcium-activated proteolysis of neurofilament proteins in goldfish Mauthner axons.

We have examined the proteolytic breakdown of neurofilament proteins (NFPs) in isolated Mauthner axoplasm (M-axoplasm). Documentation of proteolytic breakdown of NFPs in M-axoplasm is important because NFPs are not degraded in distal segments of severed Mauthner axons (M-axons) maintained in vivo for up to 62 days at 20 degrees C. By incubating M-axoplasm with 2 mM calcium in vitro, we have demonstrated that M-axoplasm contains an endogenous calcium-activated neutral protease that degrades NFPs. This calcium-activated proteolysis of M-axoplasm NFPs produced novel bands on silver-stained gels. These novel bands were presumed to be NFP breakdown products because they reacted with antibodies to the alpha-intermediate filament antigen (anti-IFA) on immunoblots from these gels. Incubations of M-axoplasm with 2 mM calcium plus exogenous calpain produced novel bands similar to those observed for M-axoplasm incubated with 2 mM calcium. Incubations of M-axoplasm with 2mM calcium plus calpain inhibitors did not produce these novel bands. These in vitro data indicate that M-axoplasm contains calpain that degrades NFPs and produces novel bands similar to those observed from distal segments of severed M-axons maintained in vivo longer than 62 days postseverance. Factors that affect the activity of calpain or affect the ability of calpain to degrade NFPs could account for the delayed degradation of NFPs in distal segments of severed M-axons maintained in vivo.

Animals↗