Overexpression of HuD accelerates neurite outgrowth and increases GAP-43 mRNA expression in cortical neurons and retinoic acid-induced embryonic stem cells in vitro

Exp Neurol. 2001 Apr;168(2):250-8. doi: 10.1006/exnr.2000.7599.


The neuron-specific RNA-binding protein HuD binds to a U-rich regulatory element of the 3' untranslated region (3' UTR) of the GAP-43 mRNA and stabilizes the mRNA. We have previously shown that overexpression of HuD in PC12 cells increases GAP-43 protein expression and induces the spontaneous formation of multiple neurites (K. D. Anderson et al. 2000. J. Neurochem. 75: 1103-1114). In this study, we examined the effects of HuD overexpression on the initial stages of neurite outgrowth and on GAP-43 gene expression using two in vitro systems: E19 rat cortical neurons and retinoic acid (RA)-induced embryonic stem (ES) cells. Normal neurite outgrowth of cortical neurons in vitro occurs over a 3-day period with a concomitant increase in GAP-43 and HuD expression. Cortical cells were infected with a replication-deficient HSV-1 vector containing the HuD cDNA in the sense orientation (HSV-HuD). Overexpression of HuD accelerated the formation of neurites. Immunocytochemical analysis showed that excess HuD resulted in a threefold increase in the number of GAP-43-positive cells undergoing morphological differentiation after 24 h of treatment. Using in situ hybridization, we found that the increased HuD expression resulted in a twofold increase in the levels of GAP-43 mRNA. Similarly, overexpression of HuD in RA-induced embryonic stem cells was found to increase the number of GAP-43-positive cells undergoing process outgrowth. In conclusion, our results demonstrate that HuD functions in the initiation of neurite outgrowth in a manner due, at least in part, to its regulation of GAP-43 expression.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cells, Cultured
  • Cerebral Cortex / metabolism
  • ELAV Proteins
  • Embryo, Mammalian
  • Female
  • GAP-43 Protein / genetics
  • GAP-43 Protein / metabolism*
  • Genetic Vectors
  • Keratolytic Agents / pharmacology
  • Nerve Tissue Proteins / drug effects
  • Nerve Tissue Proteins / metabolism*
  • Neurites / metabolism*
  • Neurons / metabolism
  • Pregnancy
  • RNA, Messenger / metabolism*
  • RNA-Binding Proteins / drug effects
  • RNA-Binding Proteins / metabolism*
  • Rats
  • Stem Cells / drug effects
  • Stem Cells / metabolism*
  • Tretinoin / pharmacology


  • ELAV Proteins
  • GAP-43 Protein
  • Keratolytic Agents
  • Nerve Tissue Proteins
  • RNA, Messenger
  • RNA-Binding Proteins
  • Tretinoin