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1.  Mitochondrial fission and fusion in Dictyostelium discoideum: a search for proteins involved in membrane dynamics 
BMC Research Notes  2012;5:505.
Mitochondrial morphology is maintained by two distinct membrane events -fission and fusion. Altering these conserved processes can disrupt mitochondrial morphology and distribution, thereby disrupting the organelle’s functionality and impeding cellular function. In higher eukaryotes, these processes are mediated by a family of dynamin-related proteins (DRP’s). In the lower eukaryotes, for instance Dictyostelium discoideum, mitochondrial fission and fusion have been implicated but not yet established. To understand the overall mechanism of these dynamics across organisms, we developed an assay to identify fission and fusion events in Dictyostelium and to assess the involvement of the mitochondrial proteins, MidA, CluA, and two DRP’s, DymA and DymB.
Using laser scanning confocal microscopy we show, for the first time, that lower eukaryotes mediate mitochondrial fission and fusion. In Dictyostelium, these processes are balanced, occurring approximately 1 event/minute. Quantification of the rates in midA-, cluA-, dymA-, or dymB- strains established that MidA appears to play an indirect role in the regulation of fission and fusion, while the DRP’s are not essential for these processes. Rates of fission and fusion were significantly reduced in cluA-cells, indicating that CluA is necessary for maintaining both fission and fusion.
We have successfully demonstrated that Dictyostelium mitochondria undergo the dynamic processes of fission and fusion. The classical mediators of membrane dynamics - the DRP’s – are not necessary for these dynamics, whereas CluA is necessary for both processes. This work contributes to our overall understanding of mitochondrial dynamics and ultimately will provide additional insight into mitochondrial disease.
PMCID: PMC3492061  PMID: 22980139
Mitochondria; Fission; Fusion; Mitochondrial morphology; Dictyostelium discoideum; MidA; CluA; Dynamin
2.  A Simple Microscopy Assay to Teach the Processes of Phagocytosis and Exocytosis 
CBE Life Sciences Education  2012;11(2):180-186.
Phagocytosis and exocytosis are two cellular processes involving membrane dynamics. While it is easy to understand the purpose of these processes, it can be extremely difficult for students to comprehend the actual mechanisms. As membrane dynamics play a significant role in many cellular processes ranging from cell signaling to cell division to organelle renewal and maintenance, we felt that we needed to do a better job of teaching these types of processes. Thus, we developed a classroom-based protocol to simultaneously study phagocytosis and exocytosis in Tetrahymena pyriformis. In this paper, we present our results demonstrating that our undergraduate classroom experiment delivers results comparable with those acquired in a professional research laboratory. In addition, students performing the experiment do learn the mechanisms of phagocytosis and exocytosis. Finally, we demonstrate a mathematical exercise to help the students apply their data to the cell. Ultimately, this assay sets the stage for future inquiry-based experiments, in which the students develop their own experimental questions and delve deeper into the mechanisms of phagocytosis and exocytosis.
PMCID: PMC3366903  PMID: 22665590
3.  The WD repeat protein, Mdv1p, functions as a molecular adaptor by interacting with Dnm1p and Fis1p during mitochondrial fission 
The Journal of Cell Biology  2002;158(3):445-452.
Yeast mitochondrial fission is a multistep process during which the dynamin-related GTPase, Dnm1p, assembles into punctate structures that associate with the outer mitochondrial membrane and mediate mitochondrial division. Steps in the Dnm1p-dependent process of fission are regulated by the actions of the WD repeat protein, Mdv1p, and the mitochondrial outer membrane protein, Fis1p. Our previous studies suggested a model where Mdv1p functions to regulate fission at a post-Dnm1p assembly step and Fis1p functions at two distinct steps, at an early point, to regulate Dnm1p assembly, and later, together with Mdv1p, to facilitate Dnm1p-dependent mitochondrial fission. To test this model, we have examined the physical and functional relationship between Mdv1p and Fis1p and present genetic, biochemical, and two-hybrid data indicating that a Fis1p–Mdv1p complex is required to regulate mitochondrial fission. To further define the role of Mdv1p in fission, we examined the structural features of Mdv1p required for its interactions with Dnm1p and Fis1p. Data from two-hybrid analyses and GFP-tagged domains of Mdv1p indicate that it contains two functionally distinct domains that enable it to function as a molecular adaptor to regulate sequential interactions between Dnm1p and Fis1p and catalyze a rate-limiting step in mitochondrial fission.
PMCID: PMC2173813  PMID: 12163467
mitochondria; membranes; fission; dynamin-related GTPase; WD repeat

Results 1-3 (3)