We have two main projects in the lab investigating different aspects of cell biology in protozoan parasites. Kinetoplastids are a group of parasites that diverged early from the main eukaryotic lineage. They are an interesting model for cell biology since many of their organelles are single copy, meaning that organelle biogenesis and division must be precisely coordinated with the cell cycle. By understanding the mechanisms underlying this coordination, we hope to uncover conserved pathways by which eukaryotic cells regulate their subcellular architecture, and how those structures relate to organelle function. In addition, the parasitic life cycles of these organisms require adaptation to different energetic environments, and in some cases the ability to interact directly with the host. In both cases, the cell undergoes dramatic morphological transitions at the cellular and subcellular levels. We are probing these unique aspects of kinetoplastid biology in order to reveal novel proteins and systems that have evolved within this particular parasitic lineage.
Mitos in Motion: The dynamic mitochondrion of kinetoplastid parasites
Mitochondria make energy for the cell, and are a central part of many metabolic pathways. The shape and structure of mitochondria can vary according to cell type, and are thought to correlate with function. In addition, cells can change the shape of their mitochondria to adapt to different energy needs. Mitochondrial shape is controlled by the balance between membrane fusion and fission events, a process called mitochondrial dynamics. More fusion results in highly interconnected mitochondrial networks, while more fission results in many discrete, smaller mitochondria. These changes can be regulated according to the cell’s energetic state or its stage in the cell cycle. They also help ensure the health and fitness of mitochondria. Some aspects of these processes appear to be conserved across all mitochondria-containing cells.
Kinetoplastid parasites, such as Trypanosoma brucei and Crithidia fasciculata, are unusual in that each cell contains a single mitochondrion. These cells are therefore an extreme example of the coordination of mitochondrial biogenesis with the cell cycle. Furthermore, T. brucei regulates the shape and function of its mitochondrion according to its stage in the life cycle. When the parasite is in the mammalian host, mitochondrial metabolism is down-regulated, and the organelle is a single, straight tubule. In the midgut of the tse-tse fly vector, the parasites have an elaborately branched mitochondrion that is highly active. The processes involved in interconversion between these two states are not known but may be related to mitochondrial dynamics in other eukaryotes. We are currently using homology-based and screening approaches in order to identify proteins that are involved in the establishment and maintenance of mitochondrial shape in kinetoplastids, as well as those that mediate changes in mitochondrial shape as the parasites transition between insect and mammalian hosts.
Sticky parasites: How do kinetoplastids adhere to surfaces?
Pathogenic kinetoplastids such as T. brucei, T. cruzi, and Leishmania species are all transmitted by insects. Although the parasite life cycles in these insects vary widely between the different kinetoplastid species, they share a common feature: at some point the parasites need to convert from a free-swimming form to a form that can adhere to insect tissues. This attachment resembles a hemidesmosome, but its molecular composition is unknown. It’s also not known what signals, if any, direct the parasite to convert to the adherent form. Since adherence is necessary for effective colonization of the insect host, understanding how parasites stick may increase our understanding of transmission.
Crithidia fasciculata is a non-pathogenic kinetoplastid that only infects mosquitoes. It adheres to the epithelium of the mosquito hindgut and is spread between mosquitoes by defecation. The adherent form of the parasite is morphologically distinct from the free-swimming form and, like the pathogenic kinetoplastids, it adheres via a hemidesmosome-like structure. Interestingly, C. fasciculata will also adhere to tissue culture plastic, giving us a powerful in vitro model with which to study adherence. We have recently used RNAseq to identify transcripts associated with adherent parasites. We are currently investigating gene candidates identified by this approach through gene knockouts and adhesion assays. We are also, in collaboration with Michael Povelones at the University of Pennsylvania, studying the life cycle of Crithidia in the mosquito.