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Regulation of phototransduction responsiveness and retinal degeneration by a phospholipase D-generated signaling lipid.

LaLonde MM, Janssens H, Rosenbaum E, Choi SY, Gergen JP, Colley NJ, Stark WS, Frohman MA - J. Cell Biol. (2005)

Bottom Line: Drosophila melanogaster phototransduction proceeds via a phospholipase C (PLC)-triggered cascade of phosphatidylinositol (PI) lipid modifications, many steps of which remain undefined.We describe the involvement of the lipid phosphatidic acid and the enzyme that generates it, phospholipase D (Pld), in this process.Pld() flies exhibit decreased light sensitivity as well as a heightened susceptibility to retinal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY 11794, USA.

ABSTRACT
Drosophila melanogaster phototransduction proceeds via a phospholipase C (PLC)-triggered cascade of phosphatidylinositol (PI) lipid modifications, many steps of which remain undefined. We describe the involvement of the lipid phosphatidic acid and the enzyme that generates it, phospholipase D (Pld), in this process. Pld() flies exhibit decreased light sensitivity as well as a heightened susceptibility to retinal degeneration. Pld overexpression rescues flies lacking PLC from light-induced, metarhodopsin-mediated degeneration and restores visual signaling in flies lacking the PI transfer protein, which is a key player in the replenishment of the PI 4,5-bisphosphate (PIP2) substrate used by PLC to transduce light stimuli into neurological signals. Altogether, these findings suggest that Pld facilitates phototransduction by maintaining adequate levels of PIP2 and by protecting the visual system from metarhodopsin-induced, low light degeneration.

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Pld overexpression causes activity and light-dependent retinal degeneration. Retinal tissue sections were prepared from flies raised with the following conditions: under a 12-h light/12-h dark cycle for 21 d, (A) +/ Rh1; (B) P{UAS-Pld}/ Rh1; and (D) P{UAS-Pld-H1095N}/ Rh1; in the dark for 21 d, (C) P{UAS-Pld}/ Rh1; or under continuous light for 1 d, (E) P{UAS-Pld}/ Rh1; (F) trp1; (G) P{UAS-Pld}/ Rh1, trp1. The bottom panel consists of electron micrographs corresponding to E–G, with R7 labeled. (B) The overexpression of Pld resulted in changes in photoreceptor cell integrity with disarrayed architecture (white circle, outlining a single ommatidium) and widespread intracellular vacuolation (arrow). Only four of the nine complete ommatidia in this section had seven intact photoreceptor cells. The Rh1 promoter drives Pld expression only in R1–6 cells. R7/8 photoreceptors were largely spared, as can be observed in the circled ommatidia or pointed at by the arrow. (C) Maintaining the same flies in the dark substantially decreased the phenotype, as all 10 complete ommatidia in this section contained 7 intact photoreceptor cells, and only limited vacuolation was observed (arrow). (D) No degenerative changes were observed when a catalytically inactive point mutant allele of Pld (H1095N) was overexpressed. (E and F) Retinal disorganization and degeneration was observed in Pld-overexpressing flies after 1 d of continuous light stimulation (only three out of six complete ommatidia retained seven photoreceptor cells; asterisk in EM image in bottom panel shows an example of a degenerating cell; arrow shows that vacuolization and disorganization is also apparent) but not in trp1 mutant flies. (G) Pld-induced degeneration was suppressed when Pld was overexpressed in the trp- background, as all seven complete ommatidia maintained seven photoreceptor cells, and vacuolation was not observed (EM image, bottom). Sections are representative of three experiments performed.
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fig5: Pld overexpression causes activity and light-dependent retinal degeneration. Retinal tissue sections were prepared from flies raised with the following conditions: under a 12-h light/12-h dark cycle for 21 d, (A) +/ Rh1; (B) P{UAS-Pld}/ Rh1; and (D) P{UAS-Pld-H1095N}/ Rh1; in the dark for 21 d, (C) P{UAS-Pld}/ Rh1; or under continuous light for 1 d, (E) P{UAS-Pld}/ Rh1; (F) trp1; (G) P{UAS-Pld}/ Rh1, trp1. The bottom panel consists of electron micrographs corresponding to E–G, with R7 labeled. (B) The overexpression of Pld resulted in changes in photoreceptor cell integrity with disarrayed architecture (white circle, outlining a single ommatidium) and widespread intracellular vacuolation (arrow). Only four of the nine complete ommatidia in this section had seven intact photoreceptor cells. The Rh1 promoter drives Pld expression only in R1–6 cells. R7/8 photoreceptors were largely spared, as can be observed in the circled ommatidia or pointed at by the arrow. (C) Maintaining the same flies in the dark substantially decreased the phenotype, as all 10 complete ommatidia in this section contained 7 intact photoreceptor cells, and only limited vacuolation was observed (arrow). (D) No degenerative changes were observed when a catalytically inactive point mutant allele of Pld (H1095N) was overexpressed. (E and F) Retinal disorganization and degeneration was observed in Pld-overexpressing flies after 1 d of continuous light stimulation (only three out of six complete ommatidia retained seven photoreceptor cells; asterisk in EM image in bottom panel shows an example of a degenerating cell; arrow shows that vacuolization and disorganization is also apparent) but not in trp1 mutant flies. (G) Pld-induced degeneration was suppressed when Pld was overexpressed in the trp- background, as all seven complete ommatidia maintained seven photoreceptor cells, and vacuolation was not observed (EM image, bottom). Sections are representative of three experiments performed.

Mentions: To explore further the possibility that Pld-generated PA can be converted to DAG and activate the TRP ion channels, we used several approaches. First, because Pld overexpression rescued norpA7 metarhodopsin-mediated retinal degeneration, the levels of DAG that were generated by the Pld overexpression in the norpA mutant background were clearly not sufficient to trigger TRP-dependent cell death. Accordingly, Pld overexpression was examined in wild-type flies in which the physiologically normal DAG contribution from the light stimulation of PLC and subsequent PIP2 hydrolysis would be present. Overexpression of Pld in wild-type flies resulted in the progressive retinal degeneration with clear loss of photoreceptor cells and increased intracellular vacuoles (Fig. 5 B) relative to controls (Fig. 5 A). This degeneration was light (PLC) dependent, as dark-reared, Pld-overexpressing flies displayed substantially less disruption of ommatidial morphology (Fig. 5 C).


Regulation of phototransduction responsiveness and retinal degeneration by a phospholipase D-generated signaling lipid.

LaLonde MM, Janssens H, Rosenbaum E, Choi SY, Gergen JP, Colley NJ, Stark WS, Frohman MA - J. Cell Biol. (2005)

Pld overexpression causes activity and light-dependent retinal degeneration. Retinal tissue sections were prepared from flies raised with the following conditions: under a 12-h light/12-h dark cycle for 21 d, (A) +/ Rh1; (B) P{UAS-Pld}/ Rh1; and (D) P{UAS-Pld-H1095N}/ Rh1; in the dark for 21 d, (C) P{UAS-Pld}/ Rh1; or under continuous light for 1 d, (E) P{UAS-Pld}/ Rh1; (F) trp1; (G) P{UAS-Pld}/ Rh1, trp1. The bottom panel consists of electron micrographs corresponding to E–G, with R7 labeled. (B) The overexpression of Pld resulted in changes in photoreceptor cell integrity with disarrayed architecture (white circle, outlining a single ommatidium) and widespread intracellular vacuolation (arrow). Only four of the nine complete ommatidia in this section had seven intact photoreceptor cells. The Rh1 promoter drives Pld expression only in R1–6 cells. R7/8 photoreceptors were largely spared, as can be observed in the circled ommatidia or pointed at by the arrow. (C) Maintaining the same flies in the dark substantially decreased the phenotype, as all 10 complete ommatidia in this section contained 7 intact photoreceptor cells, and only limited vacuolation was observed (arrow). (D) No degenerative changes were observed when a catalytically inactive point mutant allele of Pld (H1095N) was overexpressed. (E and F) Retinal disorganization and degeneration was observed in Pld-overexpressing flies after 1 d of continuous light stimulation (only three out of six complete ommatidia retained seven photoreceptor cells; asterisk in EM image in bottom panel shows an example of a degenerating cell; arrow shows that vacuolization and disorganization is also apparent) but not in trp1 mutant flies. (G) Pld-induced degeneration was suppressed when Pld was overexpressed in the trp- background, as all seven complete ommatidia maintained seven photoreceptor cells, and vacuolation was not observed (EM image, bottom). Sections are representative of three experiments performed.
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Related In: Results  -  Collection

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fig5: Pld overexpression causes activity and light-dependent retinal degeneration. Retinal tissue sections were prepared from flies raised with the following conditions: under a 12-h light/12-h dark cycle for 21 d, (A) +/ Rh1; (B) P{UAS-Pld}/ Rh1; and (D) P{UAS-Pld-H1095N}/ Rh1; in the dark for 21 d, (C) P{UAS-Pld}/ Rh1; or under continuous light for 1 d, (E) P{UAS-Pld}/ Rh1; (F) trp1; (G) P{UAS-Pld}/ Rh1, trp1. The bottom panel consists of electron micrographs corresponding to E–G, with R7 labeled. (B) The overexpression of Pld resulted in changes in photoreceptor cell integrity with disarrayed architecture (white circle, outlining a single ommatidium) and widespread intracellular vacuolation (arrow). Only four of the nine complete ommatidia in this section had seven intact photoreceptor cells. The Rh1 promoter drives Pld expression only in R1–6 cells. R7/8 photoreceptors were largely spared, as can be observed in the circled ommatidia or pointed at by the arrow. (C) Maintaining the same flies in the dark substantially decreased the phenotype, as all 10 complete ommatidia in this section contained 7 intact photoreceptor cells, and only limited vacuolation was observed (arrow). (D) No degenerative changes were observed when a catalytically inactive point mutant allele of Pld (H1095N) was overexpressed. (E and F) Retinal disorganization and degeneration was observed in Pld-overexpressing flies after 1 d of continuous light stimulation (only three out of six complete ommatidia retained seven photoreceptor cells; asterisk in EM image in bottom panel shows an example of a degenerating cell; arrow shows that vacuolization and disorganization is also apparent) but not in trp1 mutant flies. (G) Pld-induced degeneration was suppressed when Pld was overexpressed in the trp- background, as all seven complete ommatidia maintained seven photoreceptor cells, and vacuolation was not observed (EM image, bottom). Sections are representative of three experiments performed.
Mentions: To explore further the possibility that Pld-generated PA can be converted to DAG and activate the TRP ion channels, we used several approaches. First, because Pld overexpression rescued norpA7 metarhodopsin-mediated retinal degeneration, the levels of DAG that were generated by the Pld overexpression in the norpA mutant background were clearly not sufficient to trigger TRP-dependent cell death. Accordingly, Pld overexpression was examined in wild-type flies in which the physiologically normal DAG contribution from the light stimulation of PLC and subsequent PIP2 hydrolysis would be present. Overexpression of Pld in wild-type flies resulted in the progressive retinal degeneration with clear loss of photoreceptor cells and increased intracellular vacuoles (Fig. 5 B) relative to controls (Fig. 5 A). This degeneration was light (PLC) dependent, as dark-reared, Pld-overexpressing flies displayed substantially less disruption of ommatidial morphology (Fig. 5 C).

Bottom Line: Drosophila melanogaster phototransduction proceeds via a phospholipase C (PLC)-triggered cascade of phosphatidylinositol (PI) lipid modifications, many steps of which remain undefined.We describe the involvement of the lipid phosphatidic acid and the enzyme that generates it, phospholipase D (Pld), in this process.Pld() flies exhibit decreased light sensitivity as well as a heightened susceptibility to retinal degeneration.

View Article: PubMed Central - PubMed

Affiliation: Program in Molecular and Cellular Biology, Center for Developmental Genetics, Stony Brook University, Stony Brook, NY 11794, USA.

ABSTRACT
Drosophila melanogaster phototransduction proceeds via a phospholipase C (PLC)-triggered cascade of phosphatidylinositol (PI) lipid modifications, many steps of which remain undefined. We describe the involvement of the lipid phosphatidic acid and the enzyme that generates it, phospholipase D (Pld), in this process. Pld() flies exhibit decreased light sensitivity as well as a heightened susceptibility to retinal degeneration. Pld overexpression rescues flies lacking PLC from light-induced, metarhodopsin-mediated degeneration and restores visual signaling in flies lacking the PI transfer protein, which is a key player in the replenishment of the PI 4,5-bisphosphate (PIP2) substrate used by PLC to transduce light stimuli into neurological signals. Altogether, these findings suggest that Pld facilitates phototransduction by maintaining adequate levels of PIP2 and by protecting the visual system from metarhodopsin-induced, low light degeneration.

Show MeSH
Related in: MedlinePlus