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Vol. 14, Issue 5, 2041-2056, May 2003

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A Novel Dynein Light Intermediate Chain Colocalizes with the Retrograde Motor for Intraflagellar Transport at Sites of Axoneme Assembly in Chlamydomonas and Mammalian Cells

Catherine A. Perrone ^*, Douglas Tritschler ^*, Patrick Taulman , Raqual Bower ^*, Bradley K. Yoder , and Mary E. Porter ^* 

^* Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455; Department of Cell Biology, University of Alabama at Birmingham Medical Center, Birmingham, Alabama 35294

Submitted October 23, 2002; Revised December 18, 2002; Accepted January 7, 2003
Monitoring Editor: Mary Beckerle

The assembly of cilia and flagella depends on bidirectional intraflagellar transport (IFT). Anterograde IFT is driven by kinesin II, whereas retrograde IFT requires cytoplasmic dynein 1b (cDHC1b). Little is known about how cDHC1b interacts with its cargoes or how it is regulated. Recent work identified a novel dynein light intermediate chain (D2LIC) that colocalized with the mammalian cDHC1b homolog DHC2 in the centrosomal region of cultured cells. To see whether the LIC might play a role in IFT, we characterized the gene encoding the Chlamydomonas homolog of D2LIC and found its expression is up-regulated in response to deflagellation. We show that the LIC subunit copurifies with cDHC1b during flagellar isolation, dynein extraction, sucrose density centrifugation, and immunoprecipitation. Immunocytochemistry reveals that the LIC colocalizes with cDHC1b in the basal body region and along the length of flagella in wild-type cells. Localization of the complex is altered in a collection of retrograde IFT and length control mutants, which suggests that the affected gene products directly or indirectly regulate cDHC1b activity. The mammalian DHC2 and D2LIC also colocalize in the apical cytoplasm and axonemes of ciliated epithelia in the lung, brain, and efferent duct. These studies, together with the identification of an LIC mutation, xbx-1(ok279), which disrupts retrograde IFT in Caenorhabditis elegans, indicate that the novel LIC is a component of the cDHC1b/DHC2 retrograde IFT motor in a variety of organisms.

Abbreviations used: BAC, bacterial artificial chromosome; DHC, dynein heavy chain; D2LIC, light intermediate chain associated with mammalian DHC2; IC, intermediate chain; IFT, intraflagellar transport; LC, light chain; LIC, light intermediate chain; PBS, phosphate-buffered saline; RFLP, restriction fragment length polymorphism; RT-PCR, reverse transcription-polymerase chain reaction.

Note added in proof. Since this manuscript was accepted for publication, another report localizing DHC2 and D2LIC in cilia of brain, retina, and cultured cells has appeared (Mikami, et al., J. Cell Sci. [2002]. 115, 4801–4808).

Corresponding author. E-mail address: mary-p{at}biosci.cbs.umn.edu.

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