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RET signaling-induced SPHK1 gene expression plays a role in both GDNF-induced differentiation and MEN2-type

Masashi Murakami1, Masatoshi Ichihara1, Sayaka Sobue1

  • 1Department of Medical Technology, Nagoya University Graduate School of Health Sciences, Nagoya, JapanDepartment of Pathology, Nagoya University School of Medicine, Nagoya, JapanDepartment of Molecular Carcinogenesis, Nagoya University Graduate School of Medicine, Nagoya, JapanDepartment of Cell Signaling, Gifu University School of Medicine, Gifu, JapanGifu International Institute of Biotechnology, Kakamigahara, Japan.

Journal of Neurochemistry
|June 9, 2007
PubMed

Insights

Glial cell line-derived neurotrophic factor (GDNF) signaling activates sphingosine kinase 1 (SPHK1), crucial for neurodevelopment. This pathway, when mutated in MEN2A, drives tumor growth, highlighting SPHK1 as an oncogenic target.

Area of Science:

  • Molecular Biology
  • Cell Signaling
  • Oncology

Background:

  • Glial cell line-derived neurotrophic factor (GDNF) family ligands signal through RET, a receptor tyrosine kinase.
  • RET signaling is vital for neuronal development and its dysregulation, particularly mutations in Multiple Endocrine Neoplasia type 2 (MEN2), drives oncogenesis.
  • Sphingosine kinase 1 (SPHK1) is an enzyme involved in cell growth and survival pathways.

Purpose of the Study:

  • To investigate the role of GDNF/RET signaling in regulating sphingosine kinase 1 (SPHK1) expression and activity.
  • To determine the involvement of SPHK1 in GDNF-mediated neuronal differentiation and cell growth.
  • To elucidate the oncogenic potential of SPHK1 in RET-driven cancers, specifically MEN2A.

Main Methods:

  • Utilized human neuroblastoma cells (TGW) and NIH3T3 fibroblasts.
  • Employed small interfering RNA (siRNA) to silence SPHK1 expression.
  • Investigated SPHK enzyme activity, SPHK1 protein and mRNA levels, and cell proliferation assays.
  • Analyzed promoter activity and utilized specific inhibitors for ERK1/2 and PI3 kinase pathways.

Main Results:

  • GDNF/RET signaling significantly up-regulated SPHK enzyme activity, SPHK1 protein, and SPHK1 mRNA in TGW cells.
  • SPHK1 silencing inhibited GDNF-induced neurite formation, GAP43 expression, and cell growth.
  • Constitutively active MEN2A-mutated RET, but not wild-type RET, increased SPHK activity and SPHK1 expression in NIH3T3 cells.
  • SPHK1 inhibition reduced cell growth and anchorage-independent colony formation in MEN2A-transfected cells.
  • Promoter analysis identified activator protein 2 and specificity protein 1 binding sites crucial for GDNF-induced SPHK1 transcription.
  • ERK1/2 and PI3 kinase pathways were implicated in GDNF-induced SPHK1 transcription.

Conclusions:

  • GDNF/RET signaling positively regulates SPHK1, which plays a critical role in neuronal differentiation and cell growth.
  • SPHK1 exhibits oncogenic properties in the context of MEN2A, contributing to tumor cell proliferation and survival.
  • The findings identify SPHK1 as a potential therapeutic target for RET-driven tumors.

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