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Axonal neuregulin signals cells of the oligodendrocyte lineage through activation of HER4 and Schwann cells through HER2 and HER3.

Vartanian T, Goodearl A, Viehöver A, Fischbach G - J. Cell Biol. (1997)

Bottom Line: We have found that oligodendrocite-type II astrocyte (O2A) progenitor cells and mature oligodendrocytes express HER2 and HER4 but no HER3.Coimmunoprecipitation experiments indicate that receptor activation in Schwann cells results in the formation of HER2:HER3 heterodimers.The use of unique neuregulin receptor combinations in oligodendrocytes and Schwann cells likely results in recruitment of different signaling pathways and thus provides a basis for different biological responses.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
We are interested in the signaling between axons and glia that leads to myelination and maintenance of the myelin internode, and we have focused on the role of neuregulins and their receptors. Neuregulins are a family of ligands that includes heregulin, neu differentiation factor, glial growth factor, and the acetylcholine receptor-inducing activity. Three signal transducing transmembrane receptors for neuregulins, which bear significant homology to the EGF receptor, are currently known: HER2 (erbB2), HER3 (erbB3), and HER4 (erbB4). We have found that oligodendrocite-type II astrocyte (O2A) progenitor cells and mature oligodendrocytes express HER2 and HER4 but no HER3. Schwann cells express HER2 and HER3 but little HER4. In O2A progenitor cells and oligodendrocytes, recombinant neuregulin induces the rapid tyrosine phosphorylation of only HER4. HER2 is not phosphorylated in cells of the oligodendrocyte lineage, but a physical interaction between HER2 and HER4 was detected in coimmunoprecipitation experiments. In Schwann cells, neuregulin induces the phosphorylation of both HER2 and HER3. Coimmunoprecipitation experiments indicate that receptor activation in Schwann cells results in the formation of HER2:HER3 heterodimers. Neuregulin localized immunocytochemically was present on neurites of cultured dorsal root ganglion neurons, and it was released into the medium in a form that promoted receptor tyrosine phosphorylation. Neuregulins therefore meet important criteria expected of molecules involved in axonal-glial signaling. The use of unique neuregulin receptor combinations in oligodendrocytes and Schwann cells likely results in recruitment of different signaling pathways and thus provides a basis for different biological responses.

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DRG neurons express multiple neuregulin isoforms. (A) Neurons isolated from E15 DRGs were cultured for 3 wk as described, fixed, and stained with an antibody that recognizes a peptide within the COOH terminus of the neuregulin “a” form. Immunoreactive cells were recognized with Cy3-conjugated secondary antibodies and epifluorescence. Both processes and soma stain intensely for neuregulin. (B) Western blot of cytosolic and membrane fractions from DRG neurons. Neurons isolated from DRGs were  cultured for 3 wk and lysed in hypotonic buffer, and membrane and cytosolic fractions were prepared as detailed in Materials and Methods. Proteins were solubilized in SDS-DTT–containing buffer and resolved on 4–15% gradient gels, and the resulting blots were probed  with the antineuregulin antibody described above. Bar, 50 μm.
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Figure 7: DRG neurons express multiple neuregulin isoforms. (A) Neurons isolated from E15 DRGs were cultured for 3 wk as described, fixed, and stained with an antibody that recognizes a peptide within the COOH terminus of the neuregulin “a” form. Immunoreactive cells were recognized with Cy3-conjugated secondary antibodies and epifluorescence. Both processes and soma stain intensely for neuregulin. (B) Western blot of cytosolic and membrane fractions from DRG neurons. Neurons isolated from DRGs were cultured for 3 wk and lysed in hypotonic buffer, and membrane and cytosolic fractions were prepared as detailed in Materials and Methods. Proteins were solubilized in SDS-DTT–containing buffer and resolved on 4–15% gradient gels, and the resulting blots were probed with the antineuregulin antibody described above. Bar, 50 μm.

Mentions: Having established that CNS and PNS glia have distinct neuregulin receptors, we wanted to determine if neurons secreted neuregulins and if they were present on the cell surface. We chose DRG neurons to study because they possess both CNS and PNS myelinated axons. Sensory neurons were obtained from the dorsal root ganglia of embryonic day (E)15 rats and were freed of fibroblasts and Schwann cells by treatment with 5-fluorodeoxyuridine as described in the Materials and Methods section. Neuregulin staining, detected with an antibody recognizing a COOH terminus cytoplasmic domain, is present in both the soma and neurites of cultured DRG neurons (Fig. 7 A). Cultured sensory neurons express three forms of membraneassociated neuregulin by Western blot analysis with apparent molecular masses of 144, 84, and 55 kD (Fig. 7 B). The 84-kD band is consistent with a full-length Ig-domain–containing protein. A neuregulin isoform of ∼144 kD was previously noted in Western blots of rat spinal cord (63). An ∼144-kD isoform can be predicted from known alternative splice sites, but such a cDNA has not been cloned. A 55-kD band was present in both membrane and cytosolic fractions. The 55-kD band in membrane fractions likely represents the remaining transmembrane domain plus the cytosolic domain after extracellular proteolytic processing of full-length neuregulin. In the cytosol, this 55-kD band is the free soluble cytosolic domain, which has not been previously demonstrated as a stable product of neuregulin processing (Fig. 7 B). These results suggests that, in addition to cell surface neuregulins, a proportion of the neuregulins expressed by sensory neurons are processed to release the extracellular domain.


Axonal neuregulin signals cells of the oligodendrocyte lineage through activation of HER4 and Schwann cells through HER2 and HER3.

Vartanian T, Goodearl A, Viehöver A, Fischbach G - J. Cell Biol. (1997)

DRG neurons express multiple neuregulin isoforms. (A) Neurons isolated from E15 DRGs were cultured for 3 wk as described, fixed, and stained with an antibody that recognizes a peptide within the COOH terminus of the neuregulin “a” form. Immunoreactive cells were recognized with Cy3-conjugated secondary antibodies and epifluorescence. Both processes and soma stain intensely for neuregulin. (B) Western blot of cytosolic and membrane fractions from DRG neurons. Neurons isolated from DRGs were  cultured for 3 wk and lysed in hypotonic buffer, and membrane and cytosolic fractions were prepared as detailed in Materials and Methods. Proteins were solubilized in SDS-DTT–containing buffer and resolved on 4–15% gradient gels, and the resulting blots were probed  with the antineuregulin antibody described above. Bar, 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139859&req=5

Figure 7: DRG neurons express multiple neuregulin isoforms. (A) Neurons isolated from E15 DRGs were cultured for 3 wk as described, fixed, and stained with an antibody that recognizes a peptide within the COOH terminus of the neuregulin “a” form. Immunoreactive cells were recognized with Cy3-conjugated secondary antibodies and epifluorescence. Both processes and soma stain intensely for neuregulin. (B) Western blot of cytosolic and membrane fractions from DRG neurons. Neurons isolated from DRGs were cultured for 3 wk and lysed in hypotonic buffer, and membrane and cytosolic fractions were prepared as detailed in Materials and Methods. Proteins were solubilized in SDS-DTT–containing buffer and resolved on 4–15% gradient gels, and the resulting blots were probed with the antineuregulin antibody described above. Bar, 50 μm.
Mentions: Having established that CNS and PNS glia have distinct neuregulin receptors, we wanted to determine if neurons secreted neuregulins and if they were present on the cell surface. We chose DRG neurons to study because they possess both CNS and PNS myelinated axons. Sensory neurons were obtained from the dorsal root ganglia of embryonic day (E)15 rats and were freed of fibroblasts and Schwann cells by treatment with 5-fluorodeoxyuridine as described in the Materials and Methods section. Neuregulin staining, detected with an antibody recognizing a COOH terminus cytoplasmic domain, is present in both the soma and neurites of cultured DRG neurons (Fig. 7 A). Cultured sensory neurons express three forms of membraneassociated neuregulin by Western blot analysis with apparent molecular masses of 144, 84, and 55 kD (Fig. 7 B). The 84-kD band is consistent with a full-length Ig-domain–containing protein. A neuregulin isoform of ∼144 kD was previously noted in Western blots of rat spinal cord (63). An ∼144-kD isoform can be predicted from known alternative splice sites, but such a cDNA has not been cloned. A 55-kD band was present in both membrane and cytosolic fractions. The 55-kD band in membrane fractions likely represents the remaining transmembrane domain plus the cytosolic domain after extracellular proteolytic processing of full-length neuregulin. In the cytosol, this 55-kD band is the free soluble cytosolic domain, which has not been previously demonstrated as a stable product of neuregulin processing (Fig. 7 B). These results suggests that, in addition to cell surface neuregulins, a proportion of the neuregulins expressed by sensory neurons are processed to release the extracellular domain.

Bottom Line: We have found that oligodendrocite-type II astrocyte (O2A) progenitor cells and mature oligodendrocytes express HER2 and HER4 but no HER3.Coimmunoprecipitation experiments indicate that receptor activation in Schwann cells results in the formation of HER2:HER3 heterodimers.The use of unique neuregulin receptor combinations in oligodendrocytes and Schwann cells likely results in recruitment of different signaling pathways and thus provides a basis for different biological responses.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
We are interested in the signaling between axons and glia that leads to myelination and maintenance of the myelin internode, and we have focused on the role of neuregulins and their receptors. Neuregulins are a family of ligands that includes heregulin, neu differentiation factor, glial growth factor, and the acetylcholine receptor-inducing activity. Three signal transducing transmembrane receptors for neuregulins, which bear significant homology to the EGF receptor, are currently known: HER2 (erbB2), HER3 (erbB3), and HER4 (erbB4). We have found that oligodendrocite-type II astrocyte (O2A) progenitor cells and mature oligodendrocytes express HER2 and HER4 but no HER3. Schwann cells express HER2 and HER3 but little HER4. In O2A progenitor cells and oligodendrocytes, recombinant neuregulin induces the rapid tyrosine phosphorylation of only HER4. HER2 is not phosphorylated in cells of the oligodendrocyte lineage, but a physical interaction between HER2 and HER4 was detected in coimmunoprecipitation experiments. In Schwann cells, neuregulin induces the phosphorylation of both HER2 and HER3. Coimmunoprecipitation experiments indicate that receptor activation in Schwann cells results in the formation of HER2:HER3 heterodimers. Neuregulin localized immunocytochemically was present on neurites of cultured dorsal root ganglion neurons, and it was released into the medium in a form that promoted receptor tyrosine phosphorylation. Neuregulins therefore meet important criteria expected of molecules involved in axonal-glial signaling. The use of unique neuregulin receptor combinations in oligodendrocytes and Schwann cells likely results in recruitment of different signaling pathways and thus provides a basis for different biological responses.

Show MeSH
Related in: MedlinePlus