The Logistics of Cellular Traffic
David Gershlick, University of Leeds
In every cell proteins are continuously crafted and assimilated into the cells of intricate organisms. After synthesis the proteins get directed by a complex concert of cellular machinery in order to assume their appropriate role. The eukaryotic cell can be roughly divided into several different sub-sections. Surrounded by a liquid membrane, analogous to the rubber of a balloon, cells contain a watery molecular soup, which is further accompanied by membranous sub-compartments (referred to as organelles). These organelles form specialised environments with distinct roles and characteristics. Proteins fulfil functions within the membranes, at the membrane periphery or in the liquid throughout the cell. Organising these processes is a multifaceted task with numerous components needing to deliver particular proteins to specific destinations.
Scientists choosing the task of unravelling these phenomena had a complex assignment. Early techniques allowed cells to be sliced into thin sections and imaged or split open and separated. It was the 1974 Nobel laureate George Palade who coupled the observed structures with particular functions. Palade, faced with an unnerving array of cross sections of cells, wanted to understand how proteins leave the cell. The approach involved adding a radioactive element, which was incorporated into freshly made proteins, then checking where the radioactive proteins were at different time points. Essentially, the route a protein took through the cell was tracked. These observations set the stage for the introduction of the secretory pathway as a functional system of organelles.
In the following years many conceptual breakthroughs were made. One such event was the isolation and characterisation of the ‘coated vesicle’. Vesicles are small spherical compartments shuttling from one organelle to another. Filled with proteins and other components they provide a mechanism for the transport of proteins, without having to cross a membrane. Vesicle budding/fusion events were characterised by in vitro reconstitution from isolated organelles. The rate at which these vesicles bud, migrate and fuse is unexpectedly high. It was once calculated that in mammalian cells there are approximately 155 of one particular type of vesicle budding per second. Cells are alive with hundreds of independent vesicles, sometimes travelling the length of the cell to specifically deliver their valuable contents.
In the late seventies researchers realised that there was a plethora of functional proteins waiting to be discovered. Yeast was the perfect organism for this work. They are single cells, with genomes simpler than mammalian or plant, but a seemingly as complex cellular architecture. Yeast geneticists led the way over the following 20 years with several key studies all with a shared principle. The genomes of whole populations of yeast were randomly mutated and screened to look for protein sorting defects, and the responsible mutation isolated. Although there was a degree of overlap in the studies, often new essential proteins were discovered. These methods identified a large array of effectors, allowing the mechanisms of specific processes to be elucidated.
It was understood that if a protein needs to reach a particular cellular destination to fulfil its role then it cannot do so passively. They need to be directed somehow to fill the place reserved for them in the relevant vesicle or compartment. Thus, to differentiate proteins from one another, they have specific signals. This led to the distinction of ‘cargo’ that is transported, from ‘receptors’ that mediate transport steps. In a further layer of complexity, these receptors must continuously recycle to pick up another round of cargo, much like a postman returning to pick up more letters to deliver. Often receptors pass through multiple compartments to deliver cargo. These mechanisms and protein interactions occupy scientists (including myself) to this day.
With the discovery of fluorescent proteins the study of protein trafficking had a technical revolution. Fluorescent proteins light up in a distinctive manner, in a background of effectively invisible peers. Making fused chimeras consisting of a protein of interest attached to a fluorescent protein has become commonplace. Fluorescent microscopy allows observation of the location of a protein within the context of a three-dimensional cellular environment avoiding having to slice the cell into sections. Impressive recent advances allow single molecules to be observed, as well as the imaging of vesicles in living cells.
By complementing technical developments with scientific progress the conserved mechanisms that marshal a very complex system are being exposed. Associated with defects of the pathways there are various human disorders, where understanding membrane trafficking holds hope for effective therapies. A range of microorganisms are known to hijack these pathways obtaining access to the protected inner cell, a better understanding of these perturbations not only sheds light on the processes mediating homeostasis in healthy cells but would also drive medical innovations. In my field of plant biology we study these processes to not only to gain an understanding of cell biology but also to work towards global issues. Comprehending the secretory pathway allows us to progress to the goals of creating storage compartments in plant cells for industrial and pharmaceutical proteins, to generate extra-nutritious food and even to produce biofuel in a more sustainable and yet profitable manner.
The revolutionary breakthroughs described in this article seem to have occurred every 5-10 years, and perhaps it would be prudent to anticipate another such progression. However, I believe, such predictions are misplaced. Science funding seems to have changed impetus from the so called ‘blue-sky’ research to an applied focus. Each of the advances discussed above are a direct result of curiosity driven blue-sky work, as any major novel innovations would also likely be. That is not to say that applied research is not valuable, but with the majority of researchers having to focus on foreseeable impact in order to justify funding, it would seem obvious that the likelihood of major unexpected breakthroughs decreases. However, if the historical advances have taught us anything, it is that progress can happen in unexpected places at unexpected times and it is an exciting time to observe the subtle mysteries of the cell being gradually disentangled.