@article {381,
	title = {Silencing of a putative inner arm dynein heavy chain results in flagellar immotility in Trypanosoma brucei.},
	journal = {Mol Biochem Parasitol},
	volume = {175},
	year = {2011},
	month = {2011 Jan},
	pages = {68-75},
	abstract = {The Trypanosoma brucei flagellum controls motility and is crucial for cell polarity and division. Unique features of trypanosome motility suggest that flagellar beat regulation in this organism is unusual and worthy of study. The flagellar axoneme, required for motility, has a structure that is highly conserved among eukaryotes. Of the several dyneins in the axonemal inner arm complex, dynein f is thought to control flagellar waveform shape. A T. brucei gene predicted to encode the dynein f alpha heavy chain, TbDNAH10, was silenced using RNA interference in procyclic T. brucei cells. This resulted in immotile flagella, showing no movement except for occasional slight twitches at the tips. Cell growth slowed dramatically and cells were found in large clusters. Microscopic analysis of silenced cultures showed many cells with detached flagella, sometimes entangled between multiple cells. DAPI staining showed an increased frequency of mis-positioned kinetoplasts and multinucleate cells, suggesting that these cells experience disruption at an early cell cycle stage, probably secondary to the motility defect. TEM images showed apparently normal axonemes and no discernable defects in inner arm structure. This study demonstrates the use of RNAi as an effective method to study very large genes such as dynein heavy chains (HCs), and the immotility phenotype of these dynein knockdowns suggests that an intact inner arm is necessary for flagellar beating in T. brucei. Since analogous mutants in Chlamydomonas reinhardtii retain motility, this phenotype likely reflects differences in requirements for motility and/or dynein assembly between the two organisms and these comparative studies will help elucidate the mechanisms of flagellar beat regulation.},
	keywords = {Cell Nucleus, Dyneins, Flagella, Locomotion, Microscopy, Electron, Transmission, Organelles, Protozoan Proteins, RNA Interference, Trypanosoma brucei brucei},
	issn = {1872-9428},
	doi = {10.1016/j.molbiopara.2010.09.005},
	author = {Springer, Amy L and Bruhn, David F and Kinzel, Kathryn W and Rosenthal, No{\"e}l F and Zukas, Randi and Klingbeil, Michele M}
}
@article {384,
	title = {Stem-loop silencing reveals that a third mitochondrial DNA polymerase, POLID, is required for kinetoplast DNA replication in trypanosomes.},
	journal = {Eukaryot Cell},
	volume = {7},
	year = {2008},
	month = {2008 Dec},
	pages = {2141-6},
	abstract = {Kinetoplast DNA (kDNA), the mitochondrial genome of trypanosomes, is a catenated network containing thousands of minicircles and tens of maxicircles. The topological complexity dictates some unusual features including a topoisomerase-mediated release-and-reattachment mechanism for minicircle replication and at least six mitochondrial DNA polymerases (Pols) for kDNA transactions. Previously, we identified four family A DNA Pols from Trypanosoma brucei with similarity to bacterial DNA Pol I and demonstrated that two (POLIB and POLIC) were essential for maintaining the kDNA network, while POLIA was not. Here, we used RNA interference to investigate the function of POLID in procyclic T. brucei. Stem-loop silencing of POLID resulted in growth arrest and the progressive loss of the kDNA network. Additional defects in kDNA replication included a rapid decline in minicircle and maxicircle abundance and a transient accumulation of minicircle replication intermediates before loss of the kDNA network. These results demonstrate that POLID is a third essential DNA Pol required for kDNA replication. While other eukaryotes utilize a single DNA Pol (Pol gamma) for replication of mitochondrial DNA, T. brucei requires at least three to maintain the complex kDNA network.},
	keywords = {Animals, DNA Replication, DNA, Kinetoplast, DNA-Directed DNA Polymerase, Mitochondria, Mitochondrial Proteins, Protozoan Proteins, RNA Interference, RNA, Double-Stranded, Trypanosoma brucei brucei},
	issn = {1535-9786},
	doi = {10.1128/EC.00199-08},
	author = {Chandler, Julian and Vandoros, Anthula V and Mozeleski, Brian and Klingbeil, Michele M}
}
