Abstract
Intraflagellar transport (IFT), the kinesin-2 and IFT-dynein-dependent bidirectional movement of multisubunit protein complexes called IFT-particles and associated cargo molecules along ciliary axonemes, is thought to be essential for the assembly and maintenance of virtually all eukaryotic cilia and flagella. Transport assays that allow measurements of the rates of movement of specific, fluorescently tagged, functional components of the IFT machinery, including motors, IFT particle subunits, and putative cargo, were first developed in Caenorhabditis elegans sensory cilia, and they have proved to be an important and valuable tool for dissecting the molecular mechanisms of IFT. We describe how these transport assays are performed in our laboratory and summarize the information that has been obtained by using them concerning the mechanisms of action and regulation of the motors that drive IFT, the composition and organization of the IFT-particles, and the identification of IFT-dynein subunits and ciliary tubulin isotypes as likely cargo proteins of kinesin-2-driven anterograde IFT.
| Original language | English (US) |
|---|---|
| Title of host publication | Methods in Enzymology |
| Pages | 285-304 |
| Number of pages | 20 |
| Volume | 524 |
| DOIs | |
| State | Published - 2013 |
Publication series
| Name | Methods in Enzymology |
|---|---|
| Volume | 524 |
| ISSN (Print) | 00766879 |
| ISSN (Electronic) | 15577988 |
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Keywords
- Cilium assembly
- Fluorescence microscopy
- Fluorescence recovery after photobleaching
- In vivo motility assays
- Intraflagellar transport assays
- Kinesin-2
- Mathematical modeling
- Tubulin
ASJC Scopus subject areas
- Biochemistry
- Molecular Biology
Cite this
Measuring rates of intraflagellar transport along Caenorhabditis elegans sensory cilia using fluorescence microscopy. / Brust-Mascher, Ingrid; Ou, Guangshuo; Scholey, Jonathan M.
Methods in Enzymology. Vol. 524 2013. p. 285-304 (Methods in Enzymology; Vol. 524).Research output: Chapter in Book/Report/Conference proceeding › Chapter
}
TY - CHAP
T1 - Measuring rates of intraflagellar transport along Caenorhabditis elegans sensory cilia using fluorescence microscopy
AU - Brust-Mascher, Ingrid
AU - Ou, Guangshuo
AU - Scholey, Jonathan M.
PY - 2013
Y1 - 2013
N2 - Intraflagellar transport (IFT), the kinesin-2 and IFT-dynein-dependent bidirectional movement of multisubunit protein complexes called IFT-particles and associated cargo molecules along ciliary axonemes, is thought to be essential for the assembly and maintenance of virtually all eukaryotic cilia and flagella. Transport assays that allow measurements of the rates of movement of specific, fluorescently tagged, functional components of the IFT machinery, including motors, IFT particle subunits, and putative cargo, were first developed in Caenorhabditis elegans sensory cilia, and they have proved to be an important and valuable tool for dissecting the molecular mechanisms of IFT. We describe how these transport assays are performed in our laboratory and summarize the information that has been obtained by using them concerning the mechanisms of action and regulation of the motors that drive IFT, the composition and organization of the IFT-particles, and the identification of IFT-dynein subunits and ciliary tubulin isotypes as likely cargo proteins of kinesin-2-driven anterograde IFT.
AB - Intraflagellar transport (IFT), the kinesin-2 and IFT-dynein-dependent bidirectional movement of multisubunit protein complexes called IFT-particles and associated cargo molecules along ciliary axonemes, is thought to be essential for the assembly and maintenance of virtually all eukaryotic cilia and flagella. Transport assays that allow measurements of the rates of movement of specific, fluorescently tagged, functional components of the IFT machinery, including motors, IFT particle subunits, and putative cargo, were first developed in Caenorhabditis elegans sensory cilia, and they have proved to be an important and valuable tool for dissecting the molecular mechanisms of IFT. We describe how these transport assays are performed in our laboratory and summarize the information that has been obtained by using them concerning the mechanisms of action and regulation of the motors that drive IFT, the composition and organization of the IFT-particles, and the identification of IFT-dynein subunits and ciliary tubulin isotypes as likely cargo proteins of kinesin-2-driven anterograde IFT.
KW - Cilium assembly
KW - Fluorescence microscopy
KW - Fluorescence recovery after photobleaching
KW - In vivo motility assays
KW - Intraflagellar transport assays
KW - Kinesin-2
KW - Mathematical modeling
KW - Tubulin
UR - http://www.scopus.com/inward/record.url?scp=84875245326&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84875245326&partnerID=8YFLogxK
U2 - 10.1016/B978-0-12-397945-2.00016-0
DO - 10.1016/B978-0-12-397945-2.00016-0
M3 - Chapter
C2 - 23498746
AN - SCOPUS:84875245326
SN - 9780123979452
VL - 524
T3 - Methods in Enzymology
SP - 285
EP - 304
BT - Methods in Enzymology
ER -