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DCX in PC12 cells: CREB-mediated transcription and neurite outgrowth
O Shmueli1, A Gdalyahu, K Sorokina
1Department of Molecular Genetics, The Weizmann Institute of Science, 76100 Rehovot, Israel.
Abstract:
Mutations in doublecortin (DCX) result in X-linked lissencephaly in males. To explore the role of DCX in differentiation and signal transduction we overexpressed DCX in PC12 cells. Our results indicate that DCX stabilizes microtubules and inhibits neurite outgrowth in nerve growth factor-induced differentiation. However, neurite length is increased when differentiation is induced by epidermal growth factor and forskolin or by dibutyryl-cAMP. Furthermore, CREB-mediated transcription is downregulated, supporting the notion that cytoskeletal regulatory proteins can affect the transcriptional state of a cell. Using different constructs and mutations we reach the conclusion that microtubule stabilization is a key factor, but not the only one, in controlling neurite extension. Overexpression of a mutation found in a lissencephaly patient (S47R), completely blocks neurite outgrowth. We propose that these functions are important during normal and abnormal brain development.
Insights
Doublecortin (DCX) mutations cause X-linked lissencephaly. DCX stabilizes microtubules, affecting neurite outgrowth and gene transcription, with a specific mutation blocking outgrowth, impacting brain development.
Area of Science:
- Neuroscience
- Cell Biology
- Developmental Biology
Background:
- Mutations in the doublecortin (DCX) gene are linked to X-linked lissencephaly, a severe brain malformation in males.
- Understanding DCX's function is crucial for comprehending normal and abnormal brain development.
- The role of DCX in cellular differentiation and signal transduction pathways remains to be fully elucidated.
Purpose of the Study:
- To investigate the function of doublecortin (DCX) in neuronal differentiation and signal transduction.
- To explore how DCX overexpression impacts neurite outgrowth in PC12 cells under various differentiation conditions.
- To determine the specific role of microtubule stabilization and other factors in DCX-mediated neurite extension.
Main Methods:
- Overexpression of wild-type and mutant DCX constructs in PC12 cells.
- Induction of neuronal differentiation using nerve growth factor (NGF), epidermal growth factor (EGF), forskolin, and dibutyryl-cAMP (db-cAMP).
- Analysis of neurite outgrowth, length, and CREB-mediated transcription.
Main Results:
- DCX overexpression stabilized microtubules and inhibited neurite outgrowth induced by NGF.
- Neurite length increased with EGF, forskolin, or db-cAMP induction, suggesting context-dependent effects.
- CREB-mediated transcription was downregulated, indicating crosstalk between cytoskeletal regulation and gene expression. A specific lissencephaly mutation (S47R) completely blocked neurite outgrowth.
Conclusions:
- Microtubule stabilization by DCX is a key, but not sole, determinant of neurite extension.
- DCX influences neuronal differentiation through both cytoskeletal effects and modulation of transcriptional activity.
- These findings highlight the importance of DCX function in both normal brain development and lissencephaly pathogenesis.