Inner space exploration: the chemical biologist's guide to the cell

As scientists, we live in the throes of technology- pushed forward by it, prodding it on, prompted by it to explore new landscapes. New technologies reveal to us the intricacies of the natural world. Our environment becomes more vivid and real than it was before. For example, Archaea, the oddball phylogenetic domain that straddles ground between bacteria and eukaryotes, has long been considered a denizen only of extreme environ- ments and specialized ecological niches. Yet reports in the past several years that use polymerase chain reaction (PCR) amplification of ribosomal RNA have identified robust archaeal lineages in environs as normal as Wiscon- sin dirt [l]. That this intriguing phylogenetic cousin of ours, an analysis of which might yield hints to our own evolutionary history, lives not just at a sulfurous vent at the bottom of the ocean but also, simply, underfoot is rather wonderful. One is reminded that the mundane world visible to us now is just one face of a more complex, more elegant universe that we have no means to sense. The eukaryotic cell itself might turn out to be just this kind of complex universe. A recent meeting on new tech- nologies in cell biology and genomics reminded us of a classic movie from the 1960s called ‘The Fantastic Voyage’ (a comic remake in 1987 was entitled ‘Inner Space’). In this movie, a team of scientists and their space- craft were miniaturized for a journey through the human body, an ‘inner space’ no less cosmic or mystical than the galaxies and nebulae of outer space. These lucky scien- tists experienced a real-time image of the cells, tissues and systems that sustain us; they navigated, first hand, vital processes such as breathing and digestion that we take for granted ‘every day. What would those scientists have seen if they could have been shrunk further to the size of a single molecule, traveling along the surface or through the innards of a living cell? For years scientists have prodded cells with electrodes, freeze-fractured them for microscopy, immunoprecipi- tated their contents in twos and threes, and reconstituted their elements in vitro. Yet the obvious questions - what does it actually look like in there, are cytoplasmic enzymes organized or free floating, how do the mem- branes of subcellular organelles orchestrate themselves - have remained recalcitrant to study. The astounding advances in optical microscopy and miniaturization tech- niques presented at this recent meeting have made it pos- sible to analyze the physiology of single cells in the environment of complex tissues, and single molecules within isolated living cells. In concert with these tech- niques, chemical biology might now offer a tenable route to the intracellular frontier. Consider the problem of detecting transient interactions within a cell, which has long frustrated biochemists. Fleeting contacts between enzymes as one passes a sub- strate to the next, low affinity stickiness that might cluster proteins about a scaffold, such subtle contacts - if they occur - might be invisible or indistinguishable from artifact using many in vitro tests of affinity. Intracel- lular assays such as the yeast two-hybrid system and elegant derivatives thereof allow cellular constituents to remain in their native milieu but tether them to sensors designed to monitor intracellular associations. Clearly, the more subtle interaction, sensitive it will be to such baggage. Other approaches that outfit a protein with a unique chemical handle might serve as the basis of new detection techniques. Work in our lab [Z] has demonstrated that installation of a ketone - a func- tional group unique to the extracellular landscape -on cell surface carbohydrates can serve as a novel means to fluorescently tag or otherwise label them. Others have accomplished the analogous feat on a cytoplasmic protein, by installing in it a hexapeptide containing four cysteine residues that is recognized in situ by