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Multiple ecto-nucleotidases in PC12 cells: identification and cellular distribution after heterologous expression
1Biozentrum der J.W. Goethe-Universität, AK Neurochemie, Zoologisches Institut, Frankfurt am Main, Germany.
Journal of Neurochemistry
|September 13, 2001
Summary
This study identifies key ecto-nucleotidases in PC12 cells, crucial for regulating extracellular nucleotides and supporting neural cell survival and differentiation. These enzymes are vital for neural signaling and development.
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Extracellular ATP and nucleotides modulate nervous system functions via surface enzymes called ecto-nucleotidases.
- Several ecto-nucleotidase families exist with overlapping substrate specificities, necessitating identification in neural cells.
Purpose of the Study:
- To identify ecto-nucleotidases present on PC12 cells, a model for neural cells.
- To characterize the activity and distribution of these enzymes on the cell surface and intracellularly.
Main Methods:
- Analysis of PC12 cell surface enzyme activity (ATPase, ADPase).
- Polymerase Chain Reaction (PCR) to detect ecto-nucleotidase mRNA.
- Single and double transfections of PC12 cells followed by fluorescence analysis for enzyme localization.
Main Results:
- PC12 cells exhibit significant ATPase and ADPase activity, primarily from ecto-nucleoside triphosphate diphosphohydrolase (NTPDase) family members.
- mRNA for NTPDase1-3, NTPDase5/6, ecto-nucleotide pyrophosphatase/phosphodiesterase3 (NPP3), tissue-non-specific alkaline phosphatase, and ecto-5'-nucleotidase were detected.
- Ecto-nucleotidases were found on the entire cell surface, accumulating in varicosities and neurite tips, with low ecto-5'-nucleotidase and alkaline phosphatase activity, and absent NPP3 activity.
Conclusions:
- PC12 cells express multiple ecto-nucleotidases, with NTPDases being the main drivers of ATP and ADP hydrolysis.
- These enzymes are localized to the cell surface and neurite tips, suggesting roles in neural signaling.
- PC12 cell ecto-nucleotidases are important for terminating nucleotide signaling and producing nucleosides that support cell survival and neurite differentiation.