Journal article
Subunit Interactions and Organization of the Chlamydomonas reinhardtii Intraflagellar Transport Complex A Proteins
The Journal of biological chemistry, Vol.287(15), pp.11689-11703
04/06/2012
Handle:
https://hdl.handle.net/2376/106812
PMCID: PMC3320918
PMID: 22170070
Abstract
Background:
The structure of intraflagellar transport complex A is poorly understood.
Results:
Interactions between IFT A proteins are identified.
Conclusion:
Three of the IFT A proteins can form a stable subcomplex.
Significance:
Determining the structure of IFT A will be crucial to understanding the molecular basis of its ciliary function.
Chlamydomonas reinhardtii
intraflagellar transport (IFT) particles can be biochemically resolved into two smaller assemblies, complexes A and B, that contain up to six and 15 protein subunits, respectively. We provide here the proteomic and immunological analyses that verify the identity of all six
Chlamydomonas
A proteins. Using sucrose density gradient centrifugation and antibody pulldowns, we show that all six A subunits are associated in a 16 S complex in both the cell bodies and flagella. A significant fraction of the cell body IFT43, however, exhibits a much slower sedimentation of ∼2 S and is not associated with the IFT A complex. To identify interactions between the six A proteins, we combined exhaustive yeast-based two-hybrid analysis, heterologous recombinant protein expression in
Escherichia coli
, and analysis of the newly identified complex A mutants,
ift121
and
ift122
. We show that IFT121 and IFT43 interact directly and provide evidence for additional interactions between IFT121 and IFT139, IFT121 and IFT122, IFT140 and IFT122, and IFT140 and IFT144. The mutant analysis further allows us to propose that a subset of complex A proteins, IFT144/140/122, can form a stable 12 S subcomplex that we refer to as the IFT A core. Based on these results, we propose a model for the spatial arrangement of the six IFT A components.
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Details
- Title
- Subunit Interactions and Organization of the Chlamydomonas reinhardtii Intraflagellar Transport Complex A Proteins
- Creators
- Robert H Behal - From theMark S Miller - From theHongmin Qin - theBen F Lucker - From theAlexis Jones - From theDouglas G Cole - From the
- Publication Details
- The Journal of biological chemistry, Vol.287(15), pp.11689-11703
- Academic Unit
- Center for Reproductive Biology
- Publisher
- American Society for Biochemistry and Molecular Biology; 9650 Rockville Pike, Bethesda, MD 20814, U.S.A
- Grant note
- R01-GM61920 / National Institutes of Health
- Identifiers
- 99900546732901842
- Language
- English
- Resource Type
- Journal article