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. 2011:499:259-81.
doi: 10.1016/B978-0-12-386471-0.00013-4.

Using Caenorhabditis elegans to study serpinopathies

Olivia S Long  1 , Sager J Gosai, Joon Hyeok Kwak, Dale E King, David H Perlmutter, Gary A Silverman, Stephen C Pak
Affiliations

Using Caenorhabditis elegans to study serpinopathies

Olivia S Long et al. Methods Enzymol. 2011.

Abstract

Protein misfolding, polymerization, and/or aggregation are hallmarks of serpinopathies and many other human genetic disorders including Alzheimer's, Huntington's, and Parkinson's disease. While higher organism models have helped shape our understanding of these diseases, simpler model systems, like Caenorhabditis elegans, offer great versatility for elucidating complex genetic mechanisms underlying these diseases. Moreover, recent advances in automated high-throughput methodologies have promoted C. elegans as a useful tool for drug discovery. In this chapter, we describe how one could model serpinopathies in C. elegans and how one could exploit this model to identify small molecule compounds that can be developed into effective therapeutic drugs.

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Figures

Figure 13.1
Figure 13.1
A schematic representation of a C. elegans expression plasmid. (A) The canonical vector used to build all expression vectors (Okkema et al., 1993). (B) An expression vector for expression of an aggregation-prone serpin protein in C. elegans. I, synthetic intron; UTR, untranslated region.
Figure 13.2
Figure 13.2
Transgenesis flow chart. (A) Plasmid DNA for injection are prepared. (B) DNA is injected into young adult hermaphrodites. (C) Transgenic progeny are isolated and enriched. (D) Stably integrated transgenic lines are established by exposure to irradiation.
Figure 13.3
Figure 13.3
Schematic of microinjection into C. elegans gonad.
Figure 13.4
Figure 13.4
Automated worm transfer using COPAS BIOSORT. A screenshot of the COPAS BIOSORT interface is shown. Dots in the scatter plots represent an individual animal. Animals are gated based on their TOF and EXT values which are parameters for size and granularity, respectively (polygon, upper scatter plot). Animals are further gated based on their GFP expression (polygon, lower scatter plot). Only animals that satisfy both criteria are dispensed into wells for drug screening.
Figure 13.5
Figure 13.5
Image acquisition using the ArrayScanVTI. Brightfield (left), mCherry (center), and GFP (right) images of transgenic worms expressing an aggregation-prone serpin-GFP fusion. The SpotDetector BioApplication is used to automatically quantify the number, area, and intensity of GFP-positive, serpin aggregates (green spots) within each worm. Images obtained (in part) from Gosai et al., 2010.
See this image and copyright information in PMC

References

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