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Originally published as MBC in Press, 10.1091/mbc.E06-08-0666 on November 8, 2006

Vol. 18, Issue 1, 242-252, January 2007

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High-Content Microscopy Identifies New Neurite Outgrowth RegulatorsFormula Formula

Vibor Laketa*, Jeremy C. Simpson*, Stephanie Bechtel{dagger}, Stefan Wiemann{dagger}, and Rainer Pepperkok*

*Cell Biology and Biophysics Unit, European Molecular Biology Laboratory-Heidelberg, 69117 Heidelberg, Germany; and {dagger}Division of Molecular Genome Analysis, German Cancer Research Center, 69120 Heidelberg, Germany

Submitted August 2, 2006; Revised October 2, 2006; Accepted October 27, 2006
Monitoring Editor: Anne Ridley

Neurons, with their long axons and elaborate dendritic arbour, establish the complex circuitry that is essential for the proper functioning of the nervous system. Whereas a catalogue of structural, molecular, and functional differences between axons and dendrites is accumulating, the mechanisms involved in early events of neuronal differentiation, such as neurite initiation and elongation, are less well understood, mainly because the key molecules involved remain elusive. Here we describe the establishment and application of a microscopy-based approach designed to identify novel proteins involved in neurite initiation and/or elongation. We identified 21 proteins that affected neurite outgrowth when ectopically expressed in cells. Complementary time-lapse microscopy allowed us to discriminate between early and late effector proteins. Localization experiments with GFP-tagged proteins in fixed and living cells revealed a further 14 proteins that associated with neurite tips either early or late during neurite outgrowth. Coexpression experiments of the new effector proteins provide a first glimpse on a possible functional relationship of these proteins during neurite outgrowth. Altogether, we demonstrate the potential of the systematic microscope-based screening approaches described here to tackle the complex biological process of neurite outgrowth regulation.

Formula Formula The online version of this contains supplemental material at MBC Online (http://www.molbiolcell.org).

This article was published online ahead of print in MBC in Press (http://www.molbiolcell.org/cgi/doi/10.1091/mbc.E06-08-0666) on November 8, 2006.

Address correspondence to: Vibor Laketa (laketa{at}embl.de)

Abbreviations used: NGF, nerve growth factor; qRT-PCR, quantitative reverse transcription PCR; ORF, open reading frame; Gsk3beta, Glycogen synthase kinase 3beta; GFP, green fluorescent protein.




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