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Two frizzled planar cell polarity signals in the Drosophila wing are differentially organized by the Fat/Dachsous pathway.

Hogan J, Valentine M, Cox C, Doyle K, Collier S - PLoS Genet. (2011)

Bottom Line: There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein.The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model.Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Marshall University, Huntington, West Virginia, United States of America.

ABSTRACT
The regular array of distally pointing hairs on the mature Drosophila wing is evidence for the fine control of Planar Cell Polarity (PCP) during wing development. Normal wing PCP requires both the Frizzled (Fz) PCP pathway and the Fat/Dachsous (Ft/Ds) pathway, although the functional relationship between these pathways remains under debate. There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein. The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model. Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.

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Reduced ds expression can modify pkpk hair polarity without affecting wing morphogenesis.All micrographs show dorsal wing surface of MS1096-Gal4/+; pk30, UAS-ds(IR)/pk30, tubP-GAL80ts female flies. Black arrows indicate local hair polarity. (A) Wing from fly cultured at 18°C. (B) Enlargement of anterior region outlined by red box in (A). (C) Wing from fly cultured at 30°C. (D) Enlargement of anterior region outlined by red box in (C). (E) Wing from fly cultured at 18°C then shifted to 30°C at 30 hours a.p.f. (F) Enlargement of anterior region outlined by red box in (E). (G) Wing from (A) rescaled, colored green and overlaid on wing (E) colored red. (H) Wing from (C) rescaled, colored green and overlaid on wing (E) colored red.
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pgen-1001305-g008: Reduced ds expression can modify pkpk hair polarity without affecting wing morphogenesis.All micrographs show dorsal wing surface of MS1096-Gal4/+; pk30, UAS-ds(IR)/pk30, tubP-GAL80ts female flies. Black arrows indicate local hair polarity. (A) Wing from fly cultured at 18°C. (B) Enlargement of anterior region outlined by red box in (A). (C) Wing from fly cultured at 30°C. (D) Enlargement of anterior region outlined by red box in (C). (E) Wing from fly cultured at 18°C then shifted to 30°C at 30 hours a.p.f. (F) Enlargement of anterior region outlined by red box in (E). (G) Wing from (A) rescaled, colored green and overlaid on wing (E) colored red. (H) Wing from (C) rescaled, colored green and overlaid on wing (E) colored red.

Mentions: The Ft/Ds pathway controls wing morphogenesis by determining the orientation of cell divisions and clonal growth [36] and it has been proposed that altered wing hair polarity associated with loss of Ft/Ds pathway activity might also be a consequence of abnormal cell division [37]. Our data show that altered Ft/Ds pathway activity can change wing morphology without affecting hair polarity across most of the wing (see Figure 3). In the context of our Bid-Bip model, this suggests that the role of the Ft/Ds pathway in wing morphogenesis is largely separable from its role in organizing the Late Fz(Pk) signal. However, we find that changes in Ft/Ds activity that alter wing shape consistently modify the pkpk mutant hair phenotype. This suggests that we have been unable to separate the role of Ft/Ds in wing morphogenesis from its role in organizing the Early Fz(Sple) signal. To attempt to unlink these activities, we controlled the timing of ds RNAi expression during the development of a pkpk mutant wing. Constitutive expression of ds RNAi in the developing pkpk wing (using the MS1096-Gal4 driver) alters wing morphology and changes pkpk wing hair polarity to a more distal orientation (see Figure 4G, 4H and 4I). We controlled the timing of ds RNAi expression in MS1096-Gal4; UAS-ds(IR) wings by constitutive expression of Gal80ts, a temperature-sensitive Gal4 inhibitor, that binds and inactivates Gal4 at 18°C, but not at 30°C [38]. Consequently, animals of the genotype MS1096-Gal4/+; pk30, ds(IR)/pk30, tubP-GAL80ts can be cultured at 18°C (when Gal80ts is active and inhibits Gal4) and then shifted to 30°C at specific times a.p.f. to induce ds RNAi expression in the wing. When flies of this genotype were cultivated continuously at 18°C, they showed a typical pkpk mutant wing phenotype (Figure 8A and 8B), indicating that Gal80ts effectively inhibited Gal4 at this temperature. In contrast, when flies of this genotype were cultivated continuously at 30°C, they displayed wing morphology typical of reduced ds activity (Figure 8C), combined with more distal hair polarity than a pkpk mutant (Figure 8D). Flies shifted from 18°C to 30°C during pupal development showed close to wild-type wing morphology (e.g. Figure 8E, 8G and 8H), but a hair phenotype that was dependent upon timing of the temperature shift. Flies shifted before 30 hours a.p.f. displayed the more distal hair polarity typical of continuous ds knockdown (e.g. Figure 8F) and we still observed significant modification of the pkpk hair phenotype when pupae were shifted at 36 hours a.p.f.. However, pupae shifted after 40 hours a.p.f. displayed hair polarity phenotypes within the range of normal pkpk mutant wings.


Two frizzled planar cell polarity signals in the Drosophila wing are differentially organized by the Fat/Dachsous pathway.

Hogan J, Valentine M, Cox C, Doyle K, Collier S - PLoS Genet. (2011)

Reduced ds expression can modify pkpk hair polarity without affecting wing morphogenesis.All micrographs show dorsal wing surface of MS1096-Gal4/+; pk30, UAS-ds(IR)/pk30, tubP-GAL80ts female flies. Black arrows indicate local hair polarity. (A) Wing from fly cultured at 18°C. (B) Enlargement of anterior region outlined by red box in (A). (C) Wing from fly cultured at 30°C. (D) Enlargement of anterior region outlined by red box in (C). (E) Wing from fly cultured at 18°C then shifted to 30°C at 30 hours a.p.f. (F) Enlargement of anterior region outlined by red box in (E). (G) Wing from (A) rescaled, colored green and overlaid on wing (E) colored red. (H) Wing from (C) rescaled, colored green and overlaid on wing (E) colored red.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040658&req=5

pgen-1001305-g008: Reduced ds expression can modify pkpk hair polarity without affecting wing morphogenesis.All micrographs show dorsal wing surface of MS1096-Gal4/+; pk30, UAS-ds(IR)/pk30, tubP-GAL80ts female flies. Black arrows indicate local hair polarity. (A) Wing from fly cultured at 18°C. (B) Enlargement of anterior region outlined by red box in (A). (C) Wing from fly cultured at 30°C. (D) Enlargement of anterior region outlined by red box in (C). (E) Wing from fly cultured at 18°C then shifted to 30°C at 30 hours a.p.f. (F) Enlargement of anterior region outlined by red box in (E). (G) Wing from (A) rescaled, colored green and overlaid on wing (E) colored red. (H) Wing from (C) rescaled, colored green and overlaid on wing (E) colored red.
Mentions: The Ft/Ds pathway controls wing morphogenesis by determining the orientation of cell divisions and clonal growth [36] and it has been proposed that altered wing hair polarity associated with loss of Ft/Ds pathway activity might also be a consequence of abnormal cell division [37]. Our data show that altered Ft/Ds pathway activity can change wing morphology without affecting hair polarity across most of the wing (see Figure 3). In the context of our Bid-Bip model, this suggests that the role of the Ft/Ds pathway in wing morphogenesis is largely separable from its role in organizing the Late Fz(Pk) signal. However, we find that changes in Ft/Ds activity that alter wing shape consistently modify the pkpk mutant hair phenotype. This suggests that we have been unable to separate the role of Ft/Ds in wing morphogenesis from its role in organizing the Early Fz(Sple) signal. To attempt to unlink these activities, we controlled the timing of ds RNAi expression during the development of a pkpk mutant wing. Constitutive expression of ds RNAi in the developing pkpk wing (using the MS1096-Gal4 driver) alters wing morphology and changes pkpk wing hair polarity to a more distal orientation (see Figure 4G, 4H and 4I). We controlled the timing of ds RNAi expression in MS1096-Gal4; UAS-ds(IR) wings by constitutive expression of Gal80ts, a temperature-sensitive Gal4 inhibitor, that binds and inactivates Gal4 at 18°C, but not at 30°C [38]. Consequently, animals of the genotype MS1096-Gal4/+; pk30, ds(IR)/pk30, tubP-GAL80ts can be cultured at 18°C (when Gal80ts is active and inhibits Gal4) and then shifted to 30°C at specific times a.p.f. to induce ds RNAi expression in the wing. When flies of this genotype were cultivated continuously at 18°C, they showed a typical pkpk mutant wing phenotype (Figure 8A and 8B), indicating that Gal80ts effectively inhibited Gal4 at this temperature. In contrast, when flies of this genotype were cultivated continuously at 30°C, they displayed wing morphology typical of reduced ds activity (Figure 8C), combined with more distal hair polarity than a pkpk mutant (Figure 8D). Flies shifted from 18°C to 30°C during pupal development showed close to wild-type wing morphology (e.g. Figure 8E, 8G and 8H), but a hair phenotype that was dependent upon timing of the temperature shift. Flies shifted before 30 hours a.p.f. displayed the more distal hair polarity typical of continuous ds knockdown (e.g. Figure 8F) and we still observed significant modification of the pkpk hair phenotype when pupae were shifted at 36 hours a.p.f.. However, pupae shifted after 40 hours a.p.f. displayed hair polarity phenotypes within the range of normal pkpk mutant wings.

Bottom Line: There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein.The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model.Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Marshall University, Huntington, West Virginia, United States of America.

ABSTRACT
The regular array of distally pointing hairs on the mature Drosophila wing is evidence for the fine control of Planar Cell Polarity (PCP) during wing development. Normal wing PCP requires both the Frizzled (Fz) PCP pathway and the Fat/Dachsous (Ft/Ds) pathway, although the functional relationship between these pathways remains under debate. There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein. The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model. Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.

Show MeSH
Related in: MedlinePlus