• Molecules and Cells
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• v.44 no.8
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• pp.591-601
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• 2021

Cilia are highly specialized organelles that extend from the cell membrane and function as cellular signaling hubs. Thus, cilia formation and the trafficking of signaling molecules into cilia are essential cellular processes. TULP3 and Tubby (TUB) are members of the tubby-like protein (TULP) family that regulate the ciliary trafficking of G-protein coupled receptors, but the functions of the remaining TULPs (i.e., TULP1 and TULP2) remain unclear. Herein, we explore whether these four structurally similar TULPs share a molecular function in ciliary protein trafficking. We found that TULP3 and TUB, but not TULP1 or TULP2, can rescue the defective cilia formation observed in TULP3-knockout (KO) hTERT RPE-1 cells. TULP3 and TUB also fully rescue the defective ciliary localization of ARL13B, INPP5E, and GPR161 in TULP3 KO RPE-1 cells, while TULP1 and TULP2 only mediate partial rescues. Furthermore, loss of TULP3 results in abnormal IFT140 localization, which can be fully rescued by TUB and partially rescued by TULP1 and TULP2. TUB's capacity for binding IFT-A is essential for its role in cilia formation and ciliary protein trafficking in RPE-1 cells, whereas its capacity for PIP₂ binding is required for proper cilia length and IFT140 localization. Finally, chimeric TULP1 containing the IFT-A binding domain of TULP3 fully rescues ciliary protein trafficking, but not cilia formation. Together, these two TULP domains play distinct roles in ciliary protein trafficking but are insufficient for cilia formation in RPE-1 cells. In addition, TULP1 and TULP2 play other unknown molecular roles that should be addressed in the future.

• Molecules and Cells
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• v.43 no.12
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• pp.1002-1010
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• 2020

Cilia are important eukaryotic cellular compartments required for diverse biological functions. Recent studies have revealed that protein targeting into the proper ciliary subcompartments is essential for ciliary function. In Drosophila chordotonal cilium, where mechano-electric transduction occurs, two transient receptor potential (TRP) superfamily ion channels, TRPV and TRPN, are restricted to the proximal and distal subcompartments, respectively. To understand the mechanisms underlying the sub-ciliary segregation of the two TRPs, we analyzed their localization under various conditions. In developing chordotonal cilia, TRPN was directly targeted to the ciliary tip from the beginning of its appearance and was retained in the distal subcompartment throughout development, whereas the ciliary localization of TRPV was considerably delayed. Lack of intraflagella transport-related proteins affected TRPV from the initial stage of its pre-ciliary trafficking, whereas it affected TRPN from the ciliary entry stage. The ectopic expression of the two TRP channels in both ciliated and non-ciliated cells revealed their intrinsic properties related to their localization. Taken together, our results suggest that sub-ciliary segregation of the two TRP channels relies on their distinct intrinsic properties, and begins at the initial stage of their pre-ciliary trafficking.

• Molecules and Cells
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• v.45 no.4
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• pp.169-176
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• 2022

A primary cilium, a hair-like protrusion of the plasma membrane, is a pivotal organelle for sensing external environmental signals and transducing intracellular signaling. An interesting linkage between cilia and obesity has been revealed by studies of the human genetic ciliopathies Bardet-Biedl syndrome and Alström syndrome, in which obesity is a principal manifestation. Mouse models of cell type-specific cilia dysgenesis have subsequently demonstrated that ciliary defects restricted to specific hypothalamic neurons are sufficient to induce obesity and hyperphagia. A potential mechanism underlying hypothalamic neuron cilia-related obesity is impaired ciliary localization of G protein-coupled receptors involved in the regulation of appetite and energy metabolism. A well-studied example of this is melanocortin 4 receptor (MC4R), mutations in which are the most common cause of human monogenic obesity. In the paraventricular hypothalamus neurons, a blockade of ciliary trafficking of MC4R as well as its downstream ciliary signaling leads to hyperphagia and weight gain. Another potential mechanism is reduced leptin signaling in hypothalamic neurons with defective cilia. Leptin receptors traffic to the periciliary area upon leptin stimulation. Moreover, defects in cilia formation hamper leptin signaling and actions in both developing and differentiated hypothalamic neurons. The list of obesity-linked ciliary proteins is expending and this supports a tight association between cilia and obesity. This article provides a brief review on the mechanism of how ciliary defects in hypothalamic neurons facilitate obesity.

• BMB Reports
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• v.48 no.7
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• pp.380-387
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• 2015

Cilia are conserved subcellular organelles with diverse sensory and developmental roles. Recently, they have emerged as crucial organelles whose dysfunction causes a wide spectrum of disorders called ciliopathies. Recent studies on the pathological mechanisms underlying ciliopathies showed that the ciliary compartment is further divided into subdomains with specific roles in the biogenesis, maintenance and function of cilia. Several conserved sets of molecules that play specific roles in each subcompartment have been discovered. Here we review recent progress on our understanding of ciliary subcompartments, especially focusing on the molecules required for their structure and/or function. [BMB Reports 2015; 48(7): 380-387]

• Molecules and Cells
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• v.41 no.7
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• pp.676-683
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• 2018

Cilia are highly specialized antennae-like organelles that extend from the cell surface and act as cell signaling hubs. Intraflagellar transport (IFT) is a specialized form of intracellular protein trafficking that is required for the assembly and maintenance of cilia. Because cilia are so important, mutations in several IFT components lead to human disease. Thus, clarifying the molecular functions of the IFT proteins is a high priority in cilia biology. Live imaging in various species and cellular preparations has proven to be an important technique in both the discovery of IFT and the mechanisms by which it functions. Live imaging of Drosophila cilia, however, has not yet been reported. Here, we have visualized the movement of IFT in Drosophila cilia using time-lapse live imaging for the first time. We found that NOMPB-GFP (IFT88) moves according to distinct parameters depending on the ciliary segment. NOMPB-GFP moves at a similar speed in proximal and distal cilia toward the tip (${\sim}0.45{\mu}m/s$). As it returns to the ciliary base, however, NOMPB-GFP moves at ${\sim}0.12{\mu}m/s$ in distal cilia, accelerating to ${\sim}0.70{\mu}m/s$ in proximal cilia. Furthermore, while live imaging NOMPB-GFP, we observed one of the IFT proteins required for retrograde movement, Oseg4 (WDR35), is also required for anterograde movement in distal cilia. We anticipate our time-lapse live imaging analysis technique in Drosophila cilia will be a good starting point for a more sophisticated analysis of IFT and its molecular mechanisms.