Role of SOCS and VHL Proteins in Neuronal Differentiation and Development.

The basic helix-loop-helix factors play a central role in neuronal differentiation and nervous system development, which involve the Notch and signal transducer and activator of transcription (STAT)/small mother against decapentaplegic signaling pathways. Neural stem cells differentiate into three nervous system lineages, and the suppressor of cytokine signaling (SOCS) and von Hippel-Lindau (VHL) proteins are involved in this neuronal differentiation. The SOCS and VHL proteins both contain homologous structures comprising the BC-box motif. SOCSs recruit Elongin C, Elongin B, Cullin5(Cul5), and Rbx2, whereas VHL recruits Elongin C, Elongin B, Cul2, and Rbx1. SOCSs form SBC-Cul5/E3 complexes, and VHL forms a VBC-Cul2/E3 complex. These complexes degrade the target protein and suppress its downstream transduction pathway by acting as E3 ligases via the ubiquitin-proteasome system. The Janus kinase (JAK) is the main target protein of the E3 ligase SBC-Cul5, whereas hypoxia-inducible factor is the primary target protein of the E3 ligase VBC-Cul2; nonetheless, VBC-Cul2 also targets the JAK. SOCSs not only act on the ubiquitin-proteasome system but also act directly on JAKs to suppress the Janus kinase-signal transduction and activator of transcription (JAK-STAT) pathway. Both SOCS and VHL are expressed in the nervous system, predominantly in brain neurons in the embryonic stage. Both SOCS and VHL induce neuronal differentiation. SOCS is involved in differentiation into neurons, whereas VHL is involved in differentiation into neurons and oligodendrocytes; both proteins promote neurite outgrowth. It has also been suggested that the inactivation of these proteins may lead to the development of nervous system malignancies and that these proteins may function as tumor suppressors. The mechanism of action of SOCS and VHL involved in neuronal differentiation and nervous system development is thought to be mediated through the inhibition of downstream signaling pathways, JAK-STAT, and hypoxia-inducible factor-vascular endothelial growth factor pathways. In addition, because SOCS and VHL promote nerve regeneration, they are expected to be applied in neuronal regenerative medicine for traumatic brain injury and stroke.

SOCS7-Derived BC-Box Motif Peptide Mediated Cholinergic Differentiation of Human Adipose-Derived Mesenchymal Stem Cells.

Adipose-derived mesenchymal stem cells (ADMSCs) are a type of pluripotent somatic stem cells that differentiate into various cell types such as osteoblast, chondrocyte, and neuronal cells. ADMSCs as donor cells are used to produce regenerative medicines at hospitals and clinics. However, it has not been reported that ADMSCs were differentiated to a specific type of neuron with a peptide. Here, we report that ADMSCs differentiate to the cholinergic phenotype of neurons by the SOCS7-derived BC-box motif peptide. At operations for patients with neurological disorders, a small amount of subcutaneous fat was obtained. Two weeks later, adipose-derived mesenchymal stem cells (ADMSCs) were isolated and cultured for a further 1 to 2 weeks. Flow cytometry analysis for characterization of ADMSCs was performed with CD73, CD90, and CD105 as positive markers, and CD14, CD31, and CD56 as negative markers. The results showed that cultured cells were compatible with ADMSCs. Immunocytochemical studies showed naïve ADMSCs immunopositive for p75NTR, RET, nestin, keratin, neurofilament-M, and smooth muscle actin. ADMSCs were suggested to be pluripotent stem cells. A peptide corresponding to the amino-acid sequence of BC-box motif derived from SOCS7 protein was added to the medium at a concentration of 2 µM. Three days later, immunocytochemistry analysis, Western blot analysis, ubiquitination assay, and electrophysiological analysis with patch cramp were performed. Immunostaining revealed the expression of neurofilament H (NFH), choline acetyltransferase (ChAT), and tyrosine hydroxylase (TH). In addition, Western blot analysis showed an increase in the expression of NFH, ChAT, and TH, and the expression of ChAT was more distinct than TH. Immunoprecipitation with JAK2 showed an increase in the expression of ubiquitin. Electrophysiological analysis showed a large holding potential at the recorded cells through path electrodes. The BC-box motif peptide derived from SOCS7 promoted the cholinergic differentiation of ADMSCs. This novel method will contribute to research as well as regenerative medicine for cholinergic neuron diseases.