Methods in cell biology

Although C. elegans is primarily touted for its facile genetics, there has been a burgeoning interest in studying cell biological processes in this organism. Strong genetics (Brenner, 1974; Jorgensen and Mango, 2002), the development of fluorescent protein tags (Chalfie et al., 1994; Yang et al., 1996; Zhang et al., 2004), the availability of RNA interference strategies to disrupt gene function (Fire et al., 1998; Timmons and Fire, 1998), and the ability to perform studies on primary cultures of embryonic cells (Christensen et al., 2002; Zhang et al., 2002), have all led to an increase in the number of cell biological problems addressed in the worm. Furthermore, the transparency of the organism affords a unique opportunity to study the roles of cell biological processes in a living multicellular animal. A serious obstacle to studying cell biological phenomena in C. elegans is the small size of its cells. However, advances in imaging techniques have allowed faint signals to be significantly amplified, and small structures to be visualized, allowing examination of transport, export, and import processes, as well as examination of cytoskeletal and chromatin structure. Here we have compiled a set of protocols that broadly fit under the category of Cell Biology. We begin with a brief discussion of various microscopical techniques employed by C. elegans researchers to study aspects of the cell. We then describe methods for studying protein-protein and protein-DNA interactions in C. elegans. We also describe methods used to study specific cell biological problems (e.g. endocytosis, chromatin, programmed cell death). Finally, we conclude this chapter with a discussion of primary embryonic cell culture and its uses. Contributors of sections or protocols are acknowledged in parentheses following the section or protocol titles. In some cases, protocols are based on previously published work, which is then cited after the protocol title. The on-line format of this chapter should easily allow for revisions and additions to the protocols presented here. Researchers willing to share protocols not presented here or with comments on existing protocols are encouraged to submit these to ude.rellefekcor@mahahs.