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The Drosophila homologue of Rootletin is required for mechanosensory function and ciliary rootlet formation in chordotonal sensory neurons.

Styczynska-Soczka K, Jarman AP - Cilia (2015)

Bottom Line: Knock-down of Rootletin results in loss of ciliary rootlet in these neurons and severe disruption of their sensory function.No evidence was found for a defect in cell division.Although our evidence is consistent with an anchoring role for the rootlet, severe loss of mechanosensory function of chordotonal (Ch) neurons upon Rootletin knock-down may suggest a direct role for the rootlet in the mechanotransduction mechanism itself.

View Article: PubMed Central - PubMed

Affiliation: Centre for Integrative Physiology, University of Edinburgh, Edinburgh, EH8 9XD UK.

ABSTRACT

Background: In vertebrates, rootletin is the major structural component of the ciliary rootlet and is also part of the tether linking the centrioles of the centrosome. Various functions have been ascribed to the rootlet, including maintenance of ciliary integrity through anchoring and facilitation of transport to the cilium or at the base of the cilium. In Drosophila, Rootletin function has not been explored.

Results: In the Drosophila embryo, Rootletin is expressed exclusively in cell lineages of type I sensory neurons, the only somatic cells bearing a cilium. Expression is strongest in mechanosensory chordotonal neurons. Knock-down of Rootletin results in loss of ciliary rootlet in these neurons and severe disruption of their sensory function. However, the sensory cilium appears largely normal in structure and in localisation of proteins suggesting no strong defect in ciliogenesis. No evidence was found for a defect in cell division.

Conclusions: The role of Rootletin as a component of the ciliary rootlet is conserved in Drosophila. In contrast, lack of a general role in cell division is consistent with lack of centriole tethering during the centrosome cycle in Drosophila. Although our evidence is consistent with an anchoring role for the rootlet, severe loss of mechanosensory function of chordotonal (Ch) neurons upon Rootletin knock-down may suggest a direct role for the rootlet in the mechanotransduction mechanism itself.

No MeSH data available.


Rootletin knock-down does not affect Ch lineage cell divisions or cilium formation. a, b Stage 17 embryos stained with anti-CPO (green) and anti-Futsch (magenta). a Wild type, showing lch5 and v’ch1 Ch neurons. Three cell types found in the chordotonal organ are indicated (neuron, scolopale, cap). Also indicated are ES neurons (lesA–C, v’es2). bRootletin knock-down, all these cell types are visible. c, d Stage 16 embryos stained with anti-Futsch (magenta) and anti-HRP (green). c Wild type, showing the HRP-positive cilia at the tips of the Ch neurons (lch5 and vchAB). dRootletin knock-down, showing that the ch neuron cilia are present. e, f Cilia of adult femoral Ch neurons detected with mCD8-GFP. The terminal dendrite of single neurons is shown with the non-ciliary inner segment (is) and cilium (ci, yellow line) indicated. e Wild type. Cilium length = 9.5 μm (f) Rootletin knock-down., Cilium length = 9.8 μm
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Fig2: Rootletin knock-down does not affect Ch lineage cell divisions or cilium formation. a, b Stage 17 embryos stained with anti-CPO (green) and anti-Futsch (magenta). a Wild type, showing lch5 and v’ch1 Ch neurons. Three cell types found in the chordotonal organ are indicated (neuron, scolopale, cap). Also indicated are ES neurons (lesA–C, v’es2). bRootletin knock-down, all these cell types are visible. c, d Stage 16 embryos stained with anti-Futsch (magenta) and anti-HRP (green). c Wild type, showing the HRP-positive cilia at the tips of the Ch neurons (lch5 and vchAB). dRootletin knock-down, showing that the ch neuron cilia are present. e, f Cilia of adult femoral Ch neurons detected with mCD8-GFP. The terminal dendrite of single neurons is shown with the non-ciliary inner segment (is) and cilium (ci, yellow line) indicated. e Wild type. Cilium length = 9.5 μm (f) Rootletin knock-down., Cilium length = 9.8 μm

Mentions: If Rootletin knock-down causes cell division disruption, this would be expected to affect the numbers and/or types of cells being formed in the sense organ lineages. However, there was no indication of a cell division defect in the sensory neuron lineages: Rootletin knock-down flies were viable with normal numbers and arrangements of bristles (produced by support cells of ES lineages) (data not shown). In order to visualise all cell types of the sense organ lineages, late embryos were stained with antibodies against Couch Potato, a protein expressed in all sense organ precursors and their progeny [22], and Futsch, a protein expressed in sensory neurons [23]. No differences were observed between knock-down embryos and controls (Fig. 2a, b). For instance, each chordotonal organ of the lateral lch5 cluster consisted of a single neuron and several supporting cells in their correct locations. Moreover, staining dividing sense organ precursor cells with γ-tubulin showed normal centrosomes and mitotic spindle (data not shown). Therefore, we found no evidence of any phenotypes that might indicate cell division defects resulting from centrosome cycle disruption.Fig. 2


The Drosophila homologue of Rootletin is required for mechanosensory function and ciliary rootlet formation in chordotonal sensory neurons.

Styczynska-Soczka K, Jarman AP - Cilia (2015)

Rootletin knock-down does not affect Ch lineage cell divisions or cilium formation. a, b Stage 17 embryos stained with anti-CPO (green) and anti-Futsch (magenta). a Wild type, showing lch5 and v’ch1 Ch neurons. Three cell types found in the chordotonal organ are indicated (neuron, scolopale, cap). Also indicated are ES neurons (lesA–C, v’es2). bRootletin knock-down, all these cell types are visible. c, d Stage 16 embryos stained with anti-Futsch (magenta) and anti-HRP (green). c Wild type, showing the HRP-positive cilia at the tips of the Ch neurons (lch5 and vchAB). dRootletin knock-down, showing that the ch neuron cilia are present. e, f Cilia of adult femoral Ch neurons detected with mCD8-GFP. The terminal dendrite of single neurons is shown with the non-ciliary inner segment (is) and cilium (ci, yellow line) indicated. e Wild type. Cilium length = 9.5 μm (f) Rootletin knock-down., Cilium length = 9.8 μm
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig2: Rootletin knock-down does not affect Ch lineage cell divisions or cilium formation. a, b Stage 17 embryos stained with anti-CPO (green) and anti-Futsch (magenta). a Wild type, showing lch5 and v’ch1 Ch neurons. Three cell types found in the chordotonal organ are indicated (neuron, scolopale, cap). Also indicated are ES neurons (lesA–C, v’es2). bRootletin knock-down, all these cell types are visible. c, d Stage 16 embryos stained with anti-Futsch (magenta) and anti-HRP (green). c Wild type, showing the HRP-positive cilia at the tips of the Ch neurons (lch5 and vchAB). dRootletin knock-down, showing that the ch neuron cilia are present. e, f Cilia of adult femoral Ch neurons detected with mCD8-GFP. The terminal dendrite of single neurons is shown with the non-ciliary inner segment (is) and cilium (ci, yellow line) indicated. e Wild type. Cilium length = 9.5 μm (f) Rootletin knock-down., Cilium length = 9.8 μm
Mentions: If Rootletin knock-down causes cell division disruption, this would be expected to affect the numbers and/or types of cells being formed in the sense organ lineages. However, there was no indication of a cell division defect in the sensory neuron lineages: Rootletin knock-down flies were viable with normal numbers and arrangements of bristles (produced by support cells of ES lineages) (data not shown). In order to visualise all cell types of the sense organ lineages, late embryos were stained with antibodies against Couch Potato, a protein expressed in all sense organ precursors and their progeny [22], and Futsch, a protein expressed in sensory neurons [23]. No differences were observed between knock-down embryos and controls (Fig. 2a, b). For instance, each chordotonal organ of the lateral lch5 cluster consisted of a single neuron and several supporting cells in their correct locations. Moreover, staining dividing sense organ precursor cells with γ-tubulin showed normal centrosomes and mitotic spindle (data not shown). Therefore, we found no evidence of any phenotypes that might indicate cell division defects resulting from centrosome cycle disruption.Fig. 2

Bottom Line: Knock-down of Rootletin results in loss of ciliary rootlet in these neurons and severe disruption of their sensory function.No evidence was found for a defect in cell division.Although our evidence is consistent with an anchoring role for the rootlet, severe loss of mechanosensory function of chordotonal (Ch) neurons upon Rootletin knock-down may suggest a direct role for the rootlet in the mechanotransduction mechanism itself.

View Article: PubMed Central - PubMed

Affiliation: Centre for Integrative Physiology, University of Edinburgh, Edinburgh, EH8 9XD UK.

ABSTRACT

Background: In vertebrates, rootletin is the major structural component of the ciliary rootlet and is also part of the tether linking the centrioles of the centrosome. Various functions have been ascribed to the rootlet, including maintenance of ciliary integrity through anchoring and facilitation of transport to the cilium or at the base of the cilium. In Drosophila, Rootletin function has not been explored.

Results: In the Drosophila embryo, Rootletin is expressed exclusively in cell lineages of type I sensory neurons, the only somatic cells bearing a cilium. Expression is strongest in mechanosensory chordotonal neurons. Knock-down of Rootletin results in loss of ciliary rootlet in these neurons and severe disruption of their sensory function. However, the sensory cilium appears largely normal in structure and in localisation of proteins suggesting no strong defect in ciliogenesis. No evidence was found for a defect in cell division.

Conclusions: The role of Rootletin as a component of the ciliary rootlet is conserved in Drosophila. In contrast, lack of a general role in cell division is consistent with lack of centriole tethering during the centrosome cycle in Drosophila. Although our evidence is consistent with an anchoring role for the rootlet, severe loss of mechanosensory function of chordotonal (Ch) neurons upon Rootletin knock-down may suggest a direct role for the rootlet in the mechanotransduction mechanism itself.

No MeSH data available.