A5089541729- Protist diversity and phylogeny
- Genetic and Kidney Cyst Diseases
- Microtubule and mitosis dynamics
- Algal biology and biofuel production
- Micro and Nano Robotics
- Photosynthetic Processes and Mechanisms
- Biocrusts and Microbial Ecology
- Genetic Syndromes and Imprinting
- Marine and coastal ecosystems
- Biomedical Research and Pathophysiology
- Renal and related cancers
- Hedgehog Signaling Pathway Studies
- Insect symbiosis and bacterial influences
- Fetal and Pediatric Neurological Disorders
- Microbial Community Ecology and Physiology
- Cerebrospinal fluid and hydrocephalus
- Methane Hydrates and Related Phenomena
- Parasitic Diseases Research and Treatment
- Fungal and yeast genetics research
- Nuclear Structure and Function
- Tissue Engineering and Regenerative Medicine
- Cellular Mechanics and Interactions
- Polyamine Metabolism and Applications
- Solar and Space Plasma Dynamics
- RNA Research and Splicing
University of Georgia
2016-2025
University of Massachusetts Chan Medical School
2004-2014
Czech Academy of Sciences, Institute of Physiology
2009
University of Cologne
1992-2005
Institut Curie
2000
Centre National de la Recherche Scientifique
2000
University of Minnesota
1996
Max Planck Institute for Biophysical Chemistry
1993
Mutations in human CEP290 cause cilia-related disorders that range severity from isolated blindness to perinatal lethality. Here, we describe a Chlamydomonas reinhardtii mutant which most of the gene is deleted. Immunoelectron microscopy indicated located flagellar transition zone close association with prominent microtubule–membrane links there. Ultrastructural analysis revealed defects these connectors, resulting loss attachment membrane microtubules. Biochemical flagella have abnormal...
In humans, seven evolutionarily conserved genes that cause the cilia-related disorder Bardet-Biedl syndrome (BBS) encode proteins form a complex termed BBSome. The function of BBSome in cilium is not well understood. We purified BBSome-like from Chlamydomonas reinhardtii flagella and found it contains at least BBS1, -4, -5, -7, -8 undergoes intraflagellar transport (IFT) association with subset IFT particles. C. insertional mutants defective -7 assemble motile, full-length but lack ability...
The assembly of the axoneme, structural scaffold cilia and flagella, requires translocation a vast quantity tubulin into growing cilium, but mechanisms that regulate targeting, quantity, timing transport are largely unknown. In Chlamydomonas, GFP-tagged α-tubulin enters as an intraflagellar (IFT) cargo by diffusion. IFT-based GFP-tubulin is elevated in IFT trains carry more tubulin. Cells possessing both nongrowing selectively target latter. preferential delivery boosts concentration soluble...
Chlamydomonas reinhardtii hydin is a central pair protein required for flagellar motility, and mice with Hydin defects develop lethal hydrocephalus. To determine if in cause hydrocephalus through mechanism involving cilia, we compared the morphology, ultrastructure, activity of cilia wild-type mutant strains. The length density brains animals normal. ciliary axoneme normal respect to 9 + 2 microtubules, dynein arms, radial spokes but one two microtubules lacks specific projection. are unable...
Mutations in Hydin cause hydrocephalus mice, and HYDIN is a strong candidate for causing humans. The gene conserved ciliated species, including Chlamydomonas reinhardtii. An antibody raised against C. reinhardtii hydin was specific an ∼540-kD flagellar protein that missing from axonemes of strains lack the central pair (CP). specifically decorated C2 microtubule CP apparatus. 80% knock down resulted short flagella lacking C2b projection microtubule; were arrested at switch points between...
The BBSome is a complex of seven proteins, including BBS4, that cycled through cilia by intraflagellar transport (IFT). Previous work has shown the membrane-associated signaling protein phospholipase D (PLD) accumulates abnormally in Chlamydomonas reinhardtii bbs mutants. Here we show PLD component wild-type but enriched ∼150-fold bbs4 cilia; this accumulation occurs progressively over time and results altered ciliary lipid composition. When BBSomes were introduced into cells, was rapidly...
Significance Bardet–Biedl syndrome (BBS) is a rare disease caused by dysfunctional cilia. In bbs mutants, the composition of ciliary membrane altered due to defects in BBSome, conserved complex BBS proteins. To determine molecular function we used single particle vivo imaging. Transport protein phospholipase D (PLD) BBSome-dependent, and PLD comigrates with BBSomes on intraflagellar transport (IFT) trains. accumulates inside cilia after removal its export sequence (CES) or absence BBSomes....
Cilia and flagella are microtubule-based organelles that protrude from the cell body. Ciliary assembly requires intraflagellar transport (IFT), a motile system delivers cargo body to flagellar tip for assembly. The process controlling injections of IFT proteins into compartment is, therefore, crucial ciliogenesis. Extensive biochemical genetic analyses have determined molecular machinery IFT, but these studies do not explain what regulates injection rate. Here, we provide evidence result...
Intraflagellar transport (IFT) trains, multimegadalton assemblies of IFT proteins and motors, traffic in cilia. To study how trains assemble, we employed fluorescence protein-tagged Chlamydomonas reinhardtii. IFT-A motor are recruited from the cell body to basal pool, assembled into move through cilium, disperse back body. In contrast this ‘open’ system, IFT-B retrograde reenter pool a portion is reused directly anterograde indicating ‘semi-open’ system. Similar systems were also observed...
The assembly and maintenance of most cilia flagella rely on intraflagellar transport (IFT). Recent in vitro studies have suggested that, together, the calponin-homology domain within IFT81 N-terminus highly basic IFT74 form a module for IFT tubulin. By using Chlamydomonas mutants IFT74, we tested this hypothesis vivo. Modification predicted tubulin-binding residues did not significantly affect anterograde length steady-state but slowed down flagellar regeneration, phenotype similar to that...
The protein nephrocystin-4 (NPHP4) is widespread in ciliated organisms, and defects NPHP4 cause nephronophthisis blindness humans. To learn more about NPHP4's function, we have studied it Chlamydomonas reinhardtii. stably incorporated into the distal part of flagellar transition zone, close to membrane CEP290, another zone protein. Therefore, these two proteins, which are independently each other, define different domains zone. A nphp4 null mutant forms flagella with nearly normal length,...
Intraflagellar transport (IFT), which is essential for the formation and function of cilia in most organisms, trafficking IFT trains (i.e. assemblies particles) that carry cargo within cilium. Defects cause several human diseases. contain complexes IFT-A IFT-B. To dissect functions these complexes, we studied a
Cilia, essential motile and sensory organelles, have several compartments: the basal body, transition zone, middle distal axoneme segments. The segment accommodates key functions, including cilium assembly activities. While contains doublet microtubules (incomplete B-tubules fused to complete A-tubules), only A-tubule extensions, its existence requires coordination of microtubule length at nanometer scale. We show that three conserved proteins, two which are mutated in ciliopathy Joubert...
Mutations in the channel protein PKD2 cause autosomal dominant polycystic kidney disease, but function of cilia remains unclear. Here, we show that targets and anchors mastigonemes, filamentous polymers glycoprotein MST1, to extracellular surface Chlamydomonas cilia. PKD2–mastigoneme complexes physically connect axonemal doublets 4 8, positioning them perpendicular plane ciliary beating. pkd2 mutant lack cells swim with reduced velocity, indicating a motility-related complex. Association...
Hydroxyproline-rich glycoproteins (HRGPs) are a ubiquitous class of protein in the extracellular matrices and cell walls plants algae, yet little is known their native structures or interactions. Here, we used electron cryomicroscopy (cryo-EM) to determine structure hydroxyproline-rich mastigoneme, an filament isolated from cilia alga Chlamydomonas reinhardtii. The demonstrates that mastigonemes formed two HRGPs (a MST1 wrapped around single copy MST3) both have hyperglycosylated...
The small Ran GTPase, a key regulator of nucleocytoplasmic transport, is also involved in microtubule assembly and nuclear membrane formation. Herein, we show by immunofluorescence, immunoelectron microscopy, biochemical analysis that fraction tightly associated with the centrosome throughout cell cycle. interaction mediated centrosomal matrix A kinase anchoring protein (AKAP450). Accordingly, when AKAP450 delocalized from centrosome, delocalized, as consequence, regrowth or altered, despite...
Centrin, a 20 kDa calcium-binding protein, is constituent of contractile basal body-associated fibers in protists and various centrosomal structures. A construct inducing centrin RNAi was used to study the effect deficiency Chlamydomonas. Transformants contained variable amounts residual (down 5% wild-type) lacked fibers. They displayed flagellar number phenotype with mostly nonflagellate cells, suggesting that required for body assembly. Furthermore, bodies often failed dock plasma membrane...
Most motile cilia and flagella have nine outer doublet two central pair (CP) microtubules. Outer microtubules are continuous with the triplet of basal body, templated by body microtubules, grow addition new subunits to their distal ("plus") ends. In contrast, CP not raising question how these assembled polarity is established. assembly in Chlamydomonas reinhardtii was analyzed electron microscopy wide-field super-resolution immunofluorescence microscopy. To analyze independently from...
The microtubule (MT) plus-end tracking protein EB1 is present at the tips of cilia and flagella; end-binding 1 (EB1) remains tip during flagellar shortening in absence intraflagellar transport (IFT), predominant system flagella. To investigate how accumulates tip, we used vivo imaging fluorescent protein-tagged (EB1-FP) Chlamydomonas reinhardtii. After photobleaching, signal recovered within minutes, indicating an exchange with unbleached entering flagella from cell body. moved independent...
Outer dynein arms (ODAs) are multiprotein complexes that drive flagellar beating. Based on genetic and biochemical analyses, ODAs preassemble in the cell body then move into flagellum by intraflagellar transport (IFT). To study ODA vivo, we expressed essential intermediate chain 2 tagged with mNeonGreen (IC2-NG) to rescue corresponding Chlamydomonas reinhardtii mutant oda6. IC2-NG moved IFT; was of low processivity increased frequency during growth. As expected, IFT diminished oda16, lacking...