scholarly journals Flagellar Length Control System: Testing a Simple Model Based on Intraflagellar Transport and Turnover

2005 ◽  
Vol 16 (1) ◽  
pp. 270-278 ◽  
Author(s):  
Wallace F. Marshall ◽  
Hongmin Qin ◽  
Mónica Rodrigo Brenni ◽  
Joel L. Rosenbaum
Keyword(s):  
Simple Model ◽  
Control Systems ◽  
Size Control ◽  
Intraflagellar Transport ◽  
Balance Point ◽  
Probe Size ◽  
Simple Model System ◽  
Length Regulation ◽  
Flagellar Assembly ◽  
Level Analysis

Flagellar length regulation provides a simple model system for addressing the general problem of organelle size control. Based on a systems-level analysis of flagellar dynamics, we have proposed a mechanism for flagellar length control in which length is set by the balance of continuous flagellar assembly and disassembly. The model proposes that the assembly rate is length dependent due to the inherent length dependence of intraflagellar transport, whereas disassembly is length independent, such that the two rates can only reach a balance point at a single length. In this report, we test this theoretical model by using three different measurements: 1) the quantity of intraflagellar transport machinery as a function of length, 2) the variation of flagellar length as a function of flagellar number, and 3) the rate of flagellar growth as a function of length. We find that the quantity of intraflagellar transport machinery is independent of length, that flagellar length is a decreasing function of flagellar number, and that flagellar growth rate in regenerating flagella depends on length and not on the time since regeneration began. These results are consistent with the balance-point model for length control. The three strategies used here are not limited to flagella and can in principle be adapted to probe size control systems for any organelle.

scholarly journals Length-dependent disassembly maintains four different flagellar lengths in Giardia

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Shane G McInally ◽  
Jane Kondev ◽  
Scott C Dawson
Keyword(s):  
Mathematical Modeling ◽  
Basal Body ◽  
Diffusion Barrier ◽  
Quantitative Imaging ◽  
Intraflagellar Transport ◽  
Ideal Model ◽  
Parasitic Protist ◽  
Length Regulation ◽  
Flagellar Assembly ◽  
Membrane Bound

With eight flagella of four different lengths, the parasitic protist Giardia is an ideal model to evaluate flagellar assembly and length regulation. To determine how four different flagellar lengths are maintained, we used live-cell quantitative imaging and mathematical modeling of conserved components of intraflagellar transport (IFT)-mediated assembly and kinesin-13-mediated disassembly in different flagellar pairs. Each axoneme has a long cytoplasmic region extending from the basal body, and transitions to a canonical membrane-bound flagellum at the ‘flagellar pore’. We determined that each flagellar pore is the site of IFT accumulation and injection, defining a diffusion barrier functionally analogous to the transition zone. IFT-mediated assembly is length-independent, as train size, speed, and injection frequencies are similar for all flagella. We demonstrate that kinesin-13 localization to the flagellar tips is inversely correlated to flagellar length. Therefore, we propose a model where a length-dependent disassembly mechanism controls multiple flagellar lengths within the same cell.

In vivo imaging shows continued association of several IFT A, B and dynein complexes while IFT trains U-turn at the tip

2021 ◽  
Author(s):  
Jenna L. Wingfield ◽  
Betlehem Mekonnen ◽  
Ilaria Mengoni ◽  
Peiwei Liu ◽  
Mareike Jordan ◽  
...  
Keyword(s):  
Simple Model ◽  
In Vivo Imaging ◽  
Fluorescent Protein ◽  
Intraflagellar Transport ◽  
Entry And Exit ◽  
Protein Carriers ◽  
Data Support ◽  
Flagellar Assembly ◽  
Protein Entry

Flagellar assembly depends on intraflagellar transport (IFT), a bidirectional motility of protein carriers, the IFT trains. The trains are periodic assemblies of IFT-A and IFT-B subcomplexes and the motors kinesin-2 and IFT dynein. At the tip, anterograde trains are remodeled for retrograde IFT, a process that in Chlamydomonas involves kinesin-2 release and train fragmentation. However, the degree of train disassembly at the tip remains unknown. Two-color imaging of fluorescent protein-tagged IFT components indicates that IFT-A and IFT-B proteins from a given anterograde train usually return in the same set of retrograde trains. Similarly, concurrent turnaround was typical for IFT-B proteins and the IFT dynein subunit D1bLIC-GFP but severance was observed as well. Our data support a simple model of IFT turnaround, in which IFT-A, IFT-B, and IFT dynein typically remain associated at the tip and anterograde trains convert directly into retrograde trains without disassembly but for possible splitting into strings of IFT complexes. Continuous association of IFT-A, IFT-B and IFT dynein during tip remodeling could balance protein entry and exit preventing the build-up of IFT material in flagella.

scholarly journals Defective flagellar assembly and length regulation in LF3 null mutants in Chlamydomonas

2003 ◽  
Vol 163 (3) ◽  
pp. 597-607 ◽  
Author(s):  
Lai-Wa Tam ◽  
William L. Dentler ◽  
Paul A. Lefebvre
Keyword(s):  
Immunofluorescence Microscopy ◽  
Intraflagellar Transport ◽  
Wild Type ◽  
Western Blots ◽  
Functional Complex ◽  
Length Regulation ◽  
Flagellar Assembly ◽  
Unequal Length ◽  
Null Mutants ◽  
Novel Protein

Four long-flagella (LF) genes are important for flagellar length control in Chlamydomonas reinhardtii. Here, we characterize two new null lf3 mutants whose phenotypes are different from previously identified lf3 mutants. These null mutants have unequal-length flagella that assemble more slowly than wild-type flagella, though their flagella can also reach abnormally long lengths. Prominent bulges are found at the distal ends of short, long, and regenerating flagella of these mutants. Analysis of the flagella by electron and immunofluorescence microscopy and by Western blots revealed that the bulges contain intraflagellar transport complexes, a defect reported previously (for review see Cole, D.G., 2003. Traffic. 4:435–442) in a subset of mutants defective in intraflagellar transport. We have cloned the wild-type LF3 gene and characterized a hypomorphic mutant allele of LF3. LF3p is a novel protein located predominantly in the cell body. It cosediments with the product of the LF1 gene in sucrose density gradients, indicating that these proteins may form a functional complex to regulate flagellar length and assembly.

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 Analysis of Biological Noise in an Organelle Size Control System

2020 ◽  
Author(s):  
David Bauer ◽  
Hiroaki Ishikawa ◽  
Kimberly A. Wemmer ◽  
Jane Kondev ◽  
Wallace F. Marshall
Keyword(s):  
Control System ◽  
Cell Function ◽  
Size Control ◽  
Intraflagellar Transport ◽  
Intrinsic Noise ◽  
Gliding Motility ◽  
Length Variation ◽  
Biological Noise ◽  
Length Regulation ◽  
Cell Variation

AbstractAnalysis of fluctuation in organelle size provides a new way to probe the mechanisms of organelle size control systems. By analyzing cell-to-cell variation and within-cell fluctuations of flagellar length in Chlamydomonas, we show that the flagellar length control system exhibits both types of variation. Cell to cell variation is dominated by cell size, while within-cell variation results from dynamic fluctuations that are subject to a constraint, providing evidence for a homeostatic size control system. We analyzed a series of candidate genes affecting flagella and found that flagellar length variation is increased in mutations which increase the average flagellar length, an effect that we show is consistent with a theoretical model for flagellar length regulation based on length-dependent intraflagellar transport balanced by length-independent disassembly. Comparing the magnitude and time-scale of length fluctuations with simple models suggests that tubulin assembly is not directly coupled with IFT-mediated arrival and that observed fluctuations involve tubulin assembly and disassembly events involving large numbers of tubulin dimers. Cells with greater differences in their flagellar lengths show impaired swimming but improved gliding motility, raising the possibility that cells have evolved mechanisms to tune intrinsic noise in length. Taken together our results show that biological noise exists at the level of subcellular structures, with a corresponding effect on cell function, and can provide new insights into the mechanisms of organelle size control.

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 Intraflagellar transport particle size scales inversely with flagellar length: revisiting the balance-point length control model

2009 ◽  
Vol 187 (1) ◽  
pp. 81-89 ◽  
Author(s):  
Benjamin D. Engel ◽  
William B. Ludington ◽  
Wallace F. Marshall
Keyword(s):  
Intraflagellar Transport ◽  
Control Model ◽  
Differential Interference Contrast Microscopy ◽  
Protein Traffic ◽  
Balance Point ◽  
Interference Contrast Microscopy ◽  
Flagellar Assembly ◽  
Transport Particle ◽  
Flagellar Regeneration ◽  
Kinetics Of

The assembly and maintenance of eukaryotic flagella are regulated by intraflagellar transport (IFT), the bidirectional traffic of IFT particles (recently renamed IFT trains) within the flagellum. We previously proposed the balance-point length control model, which predicted that the frequency of train transport should decrease as a function of flagellar length, thus modulating the length-dependent flagellar assembly rate. However, this model was challenged by the differential interference contrast microscopy observation that IFT frequency is length independent. Using total internal reflection fluorescence microscopy to quantify protein traffic during the regeneration of Chlamydomonas reinhardtii flagella, we determined that anterograde IFT trains in short flagella are composed of more kinesin-associated protein and IFT27 proteins than trains in long flagella. This length-dependent remodeling of train size is consistent with the kinetics of flagellar regeneration and supports a revised balance-point model of flagellar length control in which the size of anterograde IFT trains tunes the rate of flagellar assembly.

scholarly journals Williams Syndrome As a Model for Elucidation of the Pathway Genes - the Brain - Cognitive Functions: Genetics and Epigenetics

Acta Naturae ◽  
2014 ◽  
Vol 6 (1) ◽  
pp. 9-22 ◽  
Author(s):  
Е. А. Nikitina ◽  
A. V. Medvedeva ◽  
G. А. Zakharov ◽  
Е. V. Savvateeva-Popova
Keyword(s):  
Simple Model ◽  
Williams Syndrome ◽  
Cognitive Functions ◽  
Single Gene ◽  
Model Systems ◽  
Epigenetic Events ◽  
Simple Model System ◽  
And Behavior ◽  
Brain Behavior ◽  
The Brain

Genomic diseases or syndromes with multiple manifestations arise spontaneously and unpredictably as a result of contiguous deletions and duplications generated by unequal recombination in chromosomal regions with a specific architecture. The Williams syndrome is believed to be one of the most attractive models for linking genes, the brain, behavior and cognitive functions. It is a neurogenetic disorder resulting from a 1.5 Mb deletion at 7q11.23 which covers more than 20 genes; the hemizigosity of these genes leads to multiple manifestations, with the behavioral ones comprising three distinct domains: 1) visuo-spatial orientation; 2) verbal and linguistic defect; and 3) hypersocialisation. The shortest observed deletion leads to hemizigosity in only two genes: eln and limk1. Therefore, the first gene is supposed to be responsible for cardiovascular pathology; and the second one, for cognitive pathology. Since cognitive pathology diminishes with a patients age, the original idea of the crucial role of genes straightforwardly determining the brains morphology and behavior was substituted by ideas of the brains plasticity and the necessity of finding epigenetic factors that affect brain development and the functions manifested as behavioral changes. Recently, non-coding microRNAs (miRs) began to be considered as the main players in these epigenetic events. This review tackles the following problems: is it possible to develop relatively simple model systems to analyze the contribution of both a single gene and the consequences of its epigenetic regulation in the formation of the Williams syndromes cognitive phenotype? Is it possible to use Drosophila as a simple model system?

scholarly journals The DHC1b (DHC2) Isoform of Cytoplasmic Dynein Is Required for Flagellar Assembly

1999 ◽  
Vol 144 (3) ◽  
pp. 473-481 ◽  
Author(s):  
Gregory J. Pazour ◽  
Bethany L. Dickert ◽  
George B. Witman
Keyword(s):  
Molecular Motors ◽  
Cell Movement ◽  
Intraflagellar Transport ◽  
Cytoplasmic Dynein ◽  
Cdna Clones ◽  
Western Blots ◽  
Heavy Chains ◽  
Different Types ◽  
Flagellar Assembly ◽  
Soluble Fractions

Dyneins are microtubule-based molecular motors involved in many different types of cell movement. Most dynein heavy chains (DHCs) clearly group into cytoplasmic or axonemal isoforms. However, DHC1b has been enigmatic. To learn more about this isoform, we isolated Chlamydomonas cDNA clones encoding a portion of DHC1b, and used these clones to identify a Chlamydomonas cell line with a deletion mutation in DHC1b. The mutant grows normally and appears to have a normal Golgi apparatus, but has very short flagella. The deletion also results in a massive redistribution of raft subunits from a peri-basal body pool (Cole, D.G., D.R. Diener, A.L. Himelblau, P.L. Beech, J.C. Fuster, and J.L. Rosenbaum. 1998. J. Cell Biol. 141:993–1008) to the flagella. Rafts are particles that normally move up and down the flagella in a process known as intraflagellar transport (IFT) (Kozminski, K.G., K.A. Johnson, P. Forscher, and J.L. Rosenbaum. 1993. Proc. Natl. Acad. Sci. USA. 90:5519–5523), which is essential for assembly and maintenance of flagella. The redistribution of raft subunits apparently occurs due to a defect in the retrograde component of IFT, suggesting that DHC1b is the motor for retrograde IFT. Consistent with this, Western blots indicate that DHC1b is present in the flagellum, predominantly in the detergent- and ATP-soluble fractions. These results indicate that DHC1b is a cytoplasmic dynein essential for flagellar assembly, probably because it is the motor for retrograde IFT.

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