scholarly journals Chlamydomonas IFT70/CrDYF-1 Is a Core Component of IFT Particle Complex B and Is Required for Flagellar Assembly

2010 ◽  
Vol 21 (15) ◽  
pp. 2696-2706 ◽  
Author(s):  
Zhen-Chuan Fan ◽  
Robert H. Behal ◽  
Stefan Geimer ◽  
Zhaohui Wang ◽  
Shana M. Williamson ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Chlamydomonas Reinhardtii ◽  
Posttranslational Modification ◽  
Model Organism ◽  
Intraflagellar Transport ◽  
Apparent Size ◽  
Model Organisms ◽  
Core Component ◽  
B Subunit ◽  
Flagellar Assembly

DYF-1 is a highly conserved protein essential for ciliogenesis in several model organisms. In Caenorhabditis elegans, DYF-1 serves as an essential activator for an anterograde motor OSM-3 of intraflagellar transport (IFT), the ciliogenesis-required motility that mediates the transport of flagellar precursors and removal of turnover products. In zebrafish and Tetrahymena DYF-1 influences the cilia tubulin posttranslational modification and may have more ubiquitous function in ciliogenesis than OSM-3. Here we address how DYF-1 biochemically interacts with the IFT machinery by using the model organism Chlamydomonas reinhardtii, in which the anterograde IFT does not depend on OSM-3. Our results show that this protein is a stoichiometric component of the IFT particle complex B and interacts directly with complex B subunit IFT46. In concurrence with the established IFT protein nomenclature, DYF-1 is also named IFT70 after the apparent size of the protein. IFT70/CrDYF-1 is essential for the function of IFT in building the flagellum because the flagella of IFT70/CrDYF-1–depleted cells were greatly shortened. Together, these results demonstrate that IFT70/CrDYF-1 is a canonical subunit of IFT particle complex B and strongly support the hypothesis that the IFT machinery has species- and tissue-specific variations with functional ramifications.

scholarly journals Chlamydomonas Basal Bodies as Flagella Organizing Centers

Cells ◽  
2018 ◽  
Vol 7 (7) ◽  
pp. 79 ◽  
Author(s):  
Jenna Wingfield ◽  
Karl-Ferdinand Lechtreck
Keyword(s):  
Structure Function ◽  
Chlamydomonas Reinhardtii ◽  
Developmental Disorders ◽  
Intraflagellar Transport ◽  
Specific Protein ◽  
Basal Bodies ◽  
Unicellular Green Alga ◽  
Flagellar Assembly ◽  
Flagellar Axoneme

During ciliogenesis, centrioles convert to membrane-docked basal bodies, which initiate the formation of cilia/flagella and template the nine doublet microtubules of the flagellar axoneme. The discovery that many human diseases and developmental disorders result from defects in flagella has fueled a strong interest in the analysis of flagellar assembly. Here, we will review the structure, function, and development of basal bodies in the unicellular green alga Chlamydomonas reinhardtii, a widely used model for the analysis of basal bodies and flagella. Intraflagellar transport (IFT), a flagella-specific protein shuttle critical for ciliogenesis, was first described in C. reinhardtii. A focus of this review will be on the role of the basal bodies in organizing the IFT machinery.

scholarly journals Review of Biological Effects of Acute and Chronic Radiation Exposure on Caenorhabditis elegans

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1966
Author(s):  
Rabin Dhakal ◽  
Mohammad Yosofvand ◽  
Mahsa Yavari ◽  
Ramzi Abdulrahman ◽  
Ryan Schurr ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Ionizing Radiation ◽  
Biological Effects ◽  
Model Organism ◽  
Model Organisms ◽  
C Elegans ◽  
Chronic Radiation ◽  
Biological Responses ◽  
Chronic Radiation Exposure ◽  
Acute And Chronic Exposure

Knowledge regarding complex radiation responses in biological systems can be enhanced using genetically amenable model organisms. In this manuscript, we reviewed the use of the nematode, Caenorhabditis elegans (C. elegans), as a model organism to investigate radiation’s biological effects. Diverse types of experiments were conducted on C. elegans, using acute and chronic exposure to different ionizing radiation types, and to assess various biological responses. These responses differed based on the type and dose of radiation and the chemical substances in which the worms were grown or maintained. A few studies compared responses to various radiation types and doses as well as other environmental exposures. Therefore, this paper focused on the effect of irradiation on C. elegans, based on the intensity of the radiation dose and the length of exposure and ways to decrease the effects of ionizing radiation. Moreover, we discussed several studies showing that dietary components such as vitamin A, polyunsaturated fatty acids, and polyphenol-rich food source may promote the resistance of C. elegans to ionizing radiation and increase their life span after irradiation.

scholarly journals Length regulation of multiple flagella that self-assemble from a shared pool of components

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Thomas G Fai ◽  
Lishibanya Mohapatra ◽  
Prathitha Kar ◽  
Jane Kondev ◽  
Ariel Amir
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Green Algae ◽  
Model Organism ◽  
Size Control ◽  
Intraflagellar Transport ◽  
Canonical Model ◽  
Active Disassembly ◽  
Pool Model ◽  
Length Regulation

The single-celled green algae Chlamydomonas reinhardtii with its two flagella—microtubule-based structures of equal and constant lengths—is the canonical model organism for studying size control of organelles. Experiments have identified motor-driven transport of tubulin to the flagella tips as a key component of their length control. Here we consider a class of models whose key assumption is that proteins responsible for the intraflagellar transport (IFT) of tubulin are present in limiting amounts. We show that the limiting-pool assumption is insufficient to describe the results of severing experiments, in which a flagellum is regenerated after it has been severed. Next, we consider an extension of the limiting-pool model that incorporates proteins that depolymerize microtubules. We show that this ‘active disassembly’ model of flagellar length control explains in quantitative detail the results of severing experiments and use it to make predictions that can be tested in experiments.

scholarly journals The short flagella 1 (SHF1) gene in Chlamydomonas encodes a Crescerin TOG-domain protein required for late stages of flagellar growth.

2021 ◽  
Author(s):  
Karina Perlaza ◽  
Mariya Mirvis ◽  
Hiroaki Ishikawa ◽  
Wallace F Marshall
Keyword(s):  
Model Organism ◽  
Size Control ◽  
Intraflagellar Transport ◽  
Loss Of Function ◽  
Wild Type ◽  
Cytoplasmic Microtubule ◽  
Flagellar Assembly ◽  
Flagellar Regeneration ◽  
Domain Protein ◽  
Late Stages

Length control of flagella represents a simple and tractable system to investigate the dynamics of organelle size. Models for flagellar length control in the model organism, Chlamydomonas reinhardtii have focused on the length-dependence of the intraflagellar transport (IFT) system which manages the delivery and removal of axonemal subunits at the tip of the flagella. One of these cargoes, tubulin, is the major axonemal subunit, and its frequency of arrival at the tip plays a central role in size control models. However, the mechanisms determining tubulin dynamics at the tip are still poorly understood. We discovered a loss-of-function mutation that leads to shortened flagella, and found that this was an allele of a previously described gene, SHF1, whose molecular identity had not previously been determined. We found that SHF1 encodes a Chlamydomonas ortholog of Crescerin, previously identified as a cilia specific TOG-domain array protein that can bind tubulin via its TOG domains and increase tubulin polymerization rates. In this mutant, flagellar regeneration occurs with the same initial kinetics as wild-type cells, but plateaus at a shorter length. Using a computational model in which the flagellar microtubules are represented by a differential equation for flagellar length combined with a stochastic model for cytoplasmic microtubule dynamics, we found that our experimental results are best described by a model in which Crescerin/SHF1 binds tubulin dimers in the cytoplasm and transports them into the flagellum. We suggest that this TOG-domain protein is necessary to efficiently and preemptively increase intra-flagella tubulin levels to offset decreasing IFT cargo at the tip as flagellar assembly progresses.

scholarly journals Networks of Causal Linkage Between Eigenmodes Characterize Behavioral Dynamics of Caenorhabditis elegans

2021 ◽  
Vol 17 (9) ◽  
pp. e1009329
Author(s):  
Erik Saberski ◽  
Antonia K. Bock ◽  
Rachel Goodridge ◽  
Vitul Agarwal ◽  
Tom Lorimer ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Animal Behavior ◽  
Escape Response ◽  
Model Organism ◽  
Model Organisms ◽  
High Dimensional ◽  
Behavioral Differences ◽  
Behavioral Dynamics ◽  
Mutant Strains ◽  
Behavioral States

Behavioral phenotyping of model organisms has played an important role in unravelling the complexities of animal behavior. Techniques for classifying behavior often rely on easily identified changes in posture and motion. However, such approaches are likely to miss complex behaviors that cannot be readily distinguished by eye (e.g., behaviors produced by high dimensional dynamics). To explore this issue, we focus on the model organism Caenorhabditis elegans, where behaviors have been extensively recorded and classified. Using a dynamical systems lens, we identify high dimensional, nonlinear causal relationships between four basic shapes that describe worm motion (eigenmodes, also called “eigenworms”). We find relationships between all pairs of eigenmodes, but the timescales of the interactions vary between pairs and across individuals. Using these varying timescales, we create “interaction profiles” to represent an individual’s behavioral dynamics. As desired, these profiles are able to distinguish well-known behavioral states: i.e., the profiles for foraging individuals are distinct from those of individuals exhibiting an escape response. More importantly, we find that interaction profiles can distinguish high dimensional behaviors among divergent mutant strains that were previously classified as phenotypically similar. Specifically, we find it is able to detect phenotypic behavioral differences not previously identified in strains related to dysfunction of hermaphrodite-specific neurons.

scholarly journals Identifying RNA splicing factors using IFT genes in Chlamydomonas reinhardtii

Open Biology ◽  
2018 ◽  
Vol 8 (3) ◽  
pp. 170211 ◽  
Author(s):  
Huawen Lin ◽  
Zhengyan Zhang ◽  
Carlo Iomini ◽  
Susan K. Dutcher
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Splice Site ◽  
Rna Splicing ◽  
Splice Site Mutation ◽  
Intraflagellar Transport ◽  
Splicing Factors ◽  
Site Mutation ◽  
Nonsense Mediated Decay ◽  
Partial Restoration ◽  
Flagellar Assembly

Intraflagellar transport moves proteins in and out of flagella/cilia and it is essential for the assembly of these organelles. Using whole-genome sequencing, we identified splice site mutations in two IFT genes, IFT81 ( fla9 ) and IFT121 ( ift121-2 ), which lead to flagellar assembly defects in the unicellular green alga Chlamydomonas reinhardtii . The splicing defects in these ift mutants are partially corrected by mutations in two conserved spliceosome proteins, DGR14 and FRA10. We identified a dgr14 deletion mutant, which suppresses the 3′ splice site mutation in IFT81 , and a frameshift mutant of FRA10 , which suppresses the 5′ splice site mutation in IFT121 . Surprisingly, we found dgr14-1 and fra10 mutations suppress both splice site mutations. We suggest these two proteins are involved in facilitating splice site recognition/interaction; in their absence some splice site mutations are tolerated. Nonsense mutations in SMG1 , which is involved in nonsense-mediated decay, lead to accumulation of aberrant transcripts and partial restoration of flagellar assembly in the ift mutants. The high density of introns and the conservation of noncore splicing factors, together with the ease of scoring the ift mutant phenotype, make Chlamydomonas an attractive organism to identify new proteins involved in splicing through suppressor screening.

scholarly journals Chlamydomonas alpha-tubulin is posttranslationally modified in the flagella during flagellar assembly.

1983 ◽  
Vol 97 (1) ◽  
pp. 258-263 ◽  
Author(s):  
S W L'Hernault ◽  
J L Rosenbaum
Keyword(s):  
Protein Synthesis ◽  
Chlamydomonas Reinhardtii ◽  
Cell Body ◽  
Posttranslational Modification ◽  
De Novo ◽  
Alpha Tubulin ◽  
Flagellar Assembly ◽  
Matrix Fraction ◽  
De Novo Protein Synthesis

The principal alpha-tubulin within Chlamydomonas reinhardtii flagellar axonemes differs from the major alpha-tubulin in the cell body. We show that these two isoelectric variants of alpha-tubulin are related to one another since posttranslational modification of the cell body precursor form converts it to the axonemal form. During flagellar assembly, precursor alpha-tubulin enters the flagella and is posttranslationally modified within the flagellar matrix fraction prior to or at the time of its addition to the growing axonemal microtubules. Experiments designed to identify the nature of this posttranslational modification have also been conducted. When flagella are induced to assemble in the absence of de novo protein synthesis, tritiated acetate can be used to posttranslationally label alpha-tubulin in vivo and, under these conditions, no other flagellar polypeptides exhibit detectable labeling.

scholarly journals Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans

2020 ◽  
Author(s):  
Troy A. McDiarmid ◽  
Vinci Au ◽  
Donald G. Moerman ◽  
Catharine H. Rankin
Keyword(s):  
Caenorhabditis Elegans ◽  
Machine Vision ◽  
Negative Selection ◽  
Large Scale ◽  
Genome Engineering ◽  
Model Organism ◽  
Model Organisms ◽  
Behavioral Phenotypes ◽  
Functional Studies ◽  
Marker Selection

AbstractImproved genome engineering methods that enable automation of large and precise edits are essential for systematic investigations of genome function. We adapted peel-1 negative selection to an optimized Dual-Marker Selection (DMS) cassette protocol for CRISPR-Cas9 genome engineering in Caenorhabditis elegans and observed robust increases in multiple measures of efficiency that were consistent across injectors and four genomic loci. The use of Peel-1-DMS selection killed animals harboring transgenes as extrachromosomal arrays and spared genome edited integrants, often circumventing the need for visual screening to identify genome edited animals. To demonstrate the applicability of the approach, we created deletion alleles in the putative proteasomal subunit pbs-1 and the uncharacterized gene K04F10.3 and used machine vision to automatically characterize their phenotypic profiles, revealing homozygous essential and heterozygous behavioral phenotypes. These results provide a robust and scalable approach to rapidly generate and phenotype genome edited animals without the need for screening or scoring by eye.Author summaryThe ability to directly manipulate the genome and observe the resulting effects on the traits of an organism is a powerful approach to investigate gene function. CRISPR-based approaches to genome engineering have revolutionized such functional studies across model organisms but still face major challenges that limit the scope and complexity of projects that can be achieved in practice. Automating genome engineering and phenotyping would enable large-scale investigations of genome function in animals. Here, we describe the adaptation of peel-1 negative selection to an optimized dual-marker selection cassette CRISPR-Cas9 genome engineering method in C. elegans and combine it with automated machine vision phenotyping to achieve functional studies without the need for screening or scoring by eye. To demonstrate the applicability of the approach, we generated novel deletion alleles in two understudied genes, pbs-1 and K04F10.3, and used machine vision to characterize their phenotypic profiles, revealing homozygous lethal and heterozygous behavioral phenotypes. Our results open the door to systematic investigations of genome function in this model organism.

scholarly journals The short flagella 1 (SHF1) gene in Chlamydomonas encodes a Crescerin TOG-domain protein required for late stages of flagellar growth

2021 ◽  
Author(s):  
Karina Perlaza ◽  
Mary Mirvis ◽  
Hiroaki Ishikawa ◽  
Wallace Marshall
Keyword(s):  
Model Organism ◽  
Size Control ◽  
Intraflagellar Transport ◽  
Loss Of Function ◽  
Wild Type ◽  
Cytoplasmic Microtubule ◽  
Flagellar Assembly ◽  
Flagellar Regeneration ◽  
Domain Protein ◽  
Late Stages

Length control of flagella represents a simple and tractable system to investigate the dynamics of organelle size. Models for flagellar length control in the model organism, Chlamydomonas reinhardtii have focused on the length-dependence of the intraflagellar transport (IFT) system which manages the delivery and removal of axonemal subunits at the tip of the flagella. One of these cargoes, tubulin, is the major axonemal subunit, and its frequency of arrival at the tip plays a central role in size control models. However, the mechanisms determining tubulin dynamics at the tip are still poorly understood. We discovered a loss-of-function mutation that leads to shortened flagella, and found that this was an allele of a previously described gene, SHF1, whose molecular identity had not previously been determined.  We found that SHF1 encodes a Chlamydomonas ortholog of Crescerin, previously identified as a cilia-specific TOG-domain array protein that can bind tubulin via its TOG domains and increase tubulin polymerization rates. In this mutant, flagellar regeneration occurs with the same initial kinetics as wild-type cells, but plateaus at a shorter length. Using a computational model in which the flagellar microtubules are represented by a differential equation for flagellar length combined with a stochastic model for cytoplasmic microtubule dynamics, we found that our experimental results are best described by a model in which Crescerin/SHF1 binds tubulin dimers in the cytoplasm and transports them into the flagellum. We suggest that this TOG-domain protein is necessary to efficiently and preemptively increase intra-flagella tubulin levels to offset decreasing IFT cargo at the tip as flagellar assembly progresses.

scholarly journals Functional analysis of an individual IFT protein: IFT46 is required for transport of outer dynein arms into flagella

2007 ◽  
Vol 176 (5) ◽  
pp. 653-665 ◽  
Author(s):  
Yuqing Hou ◽  
Hongmin Qin ◽  
John A. Follit ◽  
Gregory J. Pazour ◽  
Joel L. Rosenbaum ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Chlamydomonas Reinhardtii ◽  
Intraflagellar Transport ◽  
The Other ◽  
Suppressor Mutation ◽  
Central Pair ◽  
Insertional Mutant ◽  
Flagellar Assembly ◽  
Dynein Arms ◽  
Bidirectional Movement

Intraflagellar transport (IFT), which is the bidirectional movement of particles within flagella, is required for flagellar assembly. IFT particles are composed of ∼16 proteins, which are organized into complexes A and B. We have cloned Chlamydomonas reinhardtii and mouse IFT46, and show that IFT46 is a highly conserved complex B protein in both organisms. A C. reinhardtii insertional mutant null for IFT46 has short, paralyzed flagella lacking dynein arms and with central pair defects. The mutant has greatly reduced levels of most complex B proteins, indicating that IFT46 is necessary for complex B stability. A partial suppressor mutation restores flagellar length to the ift46 mutant. IFT46 is still absent, but levels of the other IFT particle proteins are largely restored, indicating that complex B is stabilized in the suppressed strain. Axonemal ultrastructure is restored, except that the outer arms are still missing, although outer arm subunits are present in the cytoplasm. Thus, IFT46 is specifically required for transporting outer arms into the flagellum.

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