Kinesin-II Is Preferentially Targeted to Assembling Cilia and Is Required for Ciliogenesis and Normal Cytokinesis inTetrahymena

Abstract

We cloned two genes, KIN1 and KIN2, encoding kinesin-II homologues from the ciliate Tetrahymena thermophila and constructed strains lacking either KIN1 or KIN2 or both genes. Cells with a single disruption of either gene showed partly overlapping sets of defects in cell growth, motility, ciliary assembly, and thermoresistance. Deletion of both genes resulted in loss of cilia and arrests in cytokinesis. Mutant cells were unable to assemble new cilia or to maintain preexisting cilia. Double knockout cells were not viable on a standard medium but could be grown on a modified medium on which growth does not depend on phagocytosis. Double knockout cells could be rescued by transformation with a gene encoding an epitope-tagged Kin1p. In growing cells, epitope-tagged Kin1p preferentially accumulated in cilia undergoing active assembly. Kin1p was also detected in the cell body but did not show any association with the cleavage furrow. The cell division arrests observed in kinesin-II knockout cells appear to be induced by the loss of cilia and resulting cell paralysis.

Members of the NIMA-related Kinase Family Promote Disassembly of Cilia by Multiple Mechanisms

“…Strikingly, structures that closely resemble IFT particles described in Chlamydomonas (Kozminski et al, 1995) were frequently seen between the axonemes and the ciliary membranes of shortening cilia ( Figure 7B, small arrows). Such IFT particle-like structures are rarely seen in cilia of normal or in cilia of Nrk2p-GFP cells without cadmium treatment ( Figures 6A and 8A), which could be due to a slow rate of subunit exchange and low level of IFT components in already assembled cilia of Tetrahymena (Brown et al, 1999;Thazhath et al, 2004). This observation suggests that the resorption caused by Nrk2p-GFP is associated with an upregulation of IFT.…”

“…In cilia, the intraflagellar transport (IFT), a bidirectional motility system that operates inside cilia (Rosenbaum and Witman, 2002;Scholey, 2003), plays a key role in length regulation. The anterograde IFT supplies structural subunits during assembly (Piperno et al, 1996;Qin et al, 2005) and maintains the axoneme after assembly (Brown et al, 1999). The retrograde IFT recycles the IFT machinery (Signor et al, 1999) and removes ciliary structural components that turn over (Qin et al, 2004).…”

Allele-specific regulation of primary cilia function by the von Hippel–Lindau tumor suppressor

“…We observed an 80-min doubling time (Figure 1d), consistent with published growth rates. 15 We next asked what the effect might be of doxycycline withdrawal on ciliated KC12TR/VHL-TO. Cellular levels of VHL dropped significantly by 72 h after the removal of doxycycline, which was followed by a drop in ciliated cell frequency (Figure 1e).…”

Élongation de l’axonème et dynamique du transport intraflagellaire

“…Des études in vitro ont permis de révéler l'organisation moléculaire de ces deux complexes [18]. Les protéines des complexes IFT sont localisées le long de l'axonème, mais une proportion significative d'entre elles sont concentrées à la base du flagelle [16,[19][20][21][22]. L'expression de ces protéines fusionnées à la GFP (green fluorescent protein) permet de révéler leur mouvement au sein des cils et flagelles [19,21,23].…”

“…Une fois revenus à la base du flagelle, les trains sont réutilisés pour la construction de nouveaux trains antérogrades. L'inhibition de ce transport intraflagellaire empêche l'assemblage des cils et flagelles chez la plupart des organismes étudiés jusqu'à présent [13,15,20,25,26]. L'intégrité des moteurs comme des trains IFT est essentielle.…”