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Essential roles of the Tap42-regulated protein phosphatase 2A (PP2A) family in wing imaginal disc development of Drosophila melanogaster.

Wang N, Leung HT, Mazalouskas MD, Watkins GR, Gomez RJ, Wadzinski BE - PLoS ONE (2012)

Bottom Line: RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults.The Tap42(RNAi)-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects.The experimental platform described herein identifies crucial roles for Tap42•phosphatase complexes in governing imaginal disc and fly development.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee, United States of America.

ABSTRACT
Protein ser/thr phosphatase 2A family members (PP2A, PP4, and PP6) are implicated in the control of numerous biological processes, but our understanding of the in vivo function and regulation of these enzymes is limited. In this study, we investigated the role of Tap42, a common regulatory subunit for all three PP2A family members, in the development of Drosophila melanogaster wing imaginal discs. RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults. Silencing of Tap42 also altered multiple signaling pathways (HH, JNK and DPP) and triggered apoptosis in wing imaginal discs. The Tap42(RNAi)-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects. The experimental platform described herein identifies crucial roles for Tap42•phosphatase complexes in governing imaginal disc and fly development.

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Suppression of Tap42 expression in wing imaginal discs interrupts HH signaling, hampers mitosis, and triggers apoptosis.Panel A: Isolated wing imaginal discs were immunostained with antibodies recognizing Tap42 (green) and multiple components in the HH signaling pathway, including Ptc, Smo, and Ci (red). Control wing discs displayed strong Tap42 (A1) expression and the expected expression pattern for Ptc (B1), Smo (C1), and Ci (D1). Suppression of Tap42 with the pnr-Gal4 or ap-Gal4 driver effectively reduced Tap42 levels in wing discs (A2 & A3). While the levels of the HH receptor Ptc were unaffected by Tap42 silencing (B3), the expression of other downstream components of HH signaling, Smo (C3) and Ci (D3), were abrogated. Suppression of Tap42 with the pnr-Gal4 driver did not alter the expression pattern of HH signaling as shown in B2 (Ptc), C2 (Smo) and D2 (Ci). Genotypes: (A1, B1, C1, & D1) UAS-Tap42RNAi/+ as control. (A2, B2, C2, & D2) UAS-Tap42RNAi/+; pnr-Gal4/+. (A3, B3, C3, & D3) ap-Gal4/UAS-Tap42RNAi; +/+.
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pone-0038569-g004: Suppression of Tap42 expression in wing imaginal discs interrupts HH signaling, hampers mitosis, and triggers apoptosis.Panel A: Isolated wing imaginal discs were immunostained with antibodies recognizing Tap42 (green) and multiple components in the HH signaling pathway, including Ptc, Smo, and Ci (red). Control wing discs displayed strong Tap42 (A1) expression and the expected expression pattern for Ptc (B1), Smo (C1), and Ci (D1). Suppression of Tap42 with the pnr-Gal4 or ap-Gal4 driver effectively reduced Tap42 levels in wing discs (A2 & A3). While the levels of the HH receptor Ptc were unaffected by Tap42 silencing (B3), the expression of other downstream components of HH signaling, Smo (C3) and Ci (D3), were abrogated. Suppression of Tap42 with the pnr-Gal4 driver did not alter the expression pattern of HH signaling as shown in B2 (Ptc), C2 (Smo) and D2 (Ci). Genotypes: (A1, B1, C1, & D1) UAS-Tap42RNAi/+ as control. (A2, B2, C2, & D2) UAS-Tap42RNAi/+; pnr-Gal4/+. (A3, B3, C3, & D3) ap-Gal4/UAS-Tap42RNAi; +/+.

Mentions: The morphological changes seen in the ap-Gal4>UAS-Tap42RNAi wing discs could also be due to alterations in the HH signaling pathway, which has been shown to modulate DPP activity and plays a crucial role in regulation and patterning of the discs during development [28]. Given that both PP2A/Mts and PP4 have been implicated in the control of HH signaling and wing development [33]-[35], we examined the effects of Tap42RNAi on various components of this pathway. Silencing of Tap42 using the ap-Gal4 driver did not have any noticeable effects on the levels or expression pattern of Ptc (HH receptor) (Fig. 4-B3), but led to suppressed expression of the downstream effectors of HH signaling, Smoothened (Smo) and Cubitus interruptus (Ci) (Fig. 4-C3 & D3). In contrast to the ap-Gal4>UAS-Tap42RNAi wing discs, silencing of Tap42 in pnr gene domain did not alter the expression pattern of HH components (Fig. 4-B2, C2, & D2). Our cumulative analyses of ap-Gal4>UAS-Tap42RNAi wing discs indicate that Tap42's modulation of HH, DPP, and JNK signaling is required for normal wing imaginal disc development.


Essential roles of the Tap42-regulated protein phosphatase 2A (PP2A) family in wing imaginal disc development of Drosophila melanogaster.

Wang N, Leung HT, Mazalouskas MD, Watkins GR, Gomez RJ, Wadzinski BE - PLoS ONE (2012)

Suppression of Tap42 expression in wing imaginal discs interrupts HH signaling, hampers mitosis, and triggers apoptosis.Panel A: Isolated wing imaginal discs were immunostained with antibodies recognizing Tap42 (green) and multiple components in the HH signaling pathway, including Ptc, Smo, and Ci (red). Control wing discs displayed strong Tap42 (A1) expression and the expected expression pattern for Ptc (B1), Smo (C1), and Ci (D1). Suppression of Tap42 with the pnr-Gal4 or ap-Gal4 driver effectively reduced Tap42 levels in wing discs (A2 & A3). While the levels of the HH receptor Ptc were unaffected by Tap42 silencing (B3), the expression of other downstream components of HH signaling, Smo (C3) and Ci (D3), were abrogated. Suppression of Tap42 with the pnr-Gal4 driver did not alter the expression pattern of HH signaling as shown in B2 (Ptc), C2 (Smo) and D2 (Ci). Genotypes: (A1, B1, C1, & D1) UAS-Tap42RNAi/+ as control. (A2, B2, C2, & D2) UAS-Tap42RNAi/+; pnr-Gal4/+. (A3, B3, C3, & D3) ap-Gal4/UAS-Tap42RNAi; +/+.
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Related In: Results  -  Collection

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pone-0038569-g004: Suppression of Tap42 expression in wing imaginal discs interrupts HH signaling, hampers mitosis, and triggers apoptosis.Panel A: Isolated wing imaginal discs were immunostained with antibodies recognizing Tap42 (green) and multiple components in the HH signaling pathway, including Ptc, Smo, and Ci (red). Control wing discs displayed strong Tap42 (A1) expression and the expected expression pattern for Ptc (B1), Smo (C1), and Ci (D1). Suppression of Tap42 with the pnr-Gal4 or ap-Gal4 driver effectively reduced Tap42 levels in wing discs (A2 & A3). While the levels of the HH receptor Ptc were unaffected by Tap42 silencing (B3), the expression of other downstream components of HH signaling, Smo (C3) and Ci (D3), were abrogated. Suppression of Tap42 with the pnr-Gal4 driver did not alter the expression pattern of HH signaling as shown in B2 (Ptc), C2 (Smo) and D2 (Ci). Genotypes: (A1, B1, C1, & D1) UAS-Tap42RNAi/+ as control. (A2, B2, C2, & D2) UAS-Tap42RNAi/+; pnr-Gal4/+. (A3, B3, C3, & D3) ap-Gal4/UAS-Tap42RNAi; +/+.
Mentions: The morphological changes seen in the ap-Gal4>UAS-Tap42RNAi wing discs could also be due to alterations in the HH signaling pathway, which has been shown to modulate DPP activity and plays a crucial role in regulation and patterning of the discs during development [28]. Given that both PP2A/Mts and PP4 have been implicated in the control of HH signaling and wing development [33]-[35], we examined the effects of Tap42RNAi on various components of this pathway. Silencing of Tap42 using the ap-Gal4 driver did not have any noticeable effects on the levels or expression pattern of Ptc (HH receptor) (Fig. 4-B3), but led to suppressed expression of the downstream effectors of HH signaling, Smoothened (Smo) and Cubitus interruptus (Ci) (Fig. 4-C3 & D3). In contrast to the ap-Gal4>UAS-Tap42RNAi wing discs, silencing of Tap42 in pnr gene domain did not alter the expression pattern of HH components (Fig. 4-B2, C2, & D2). Our cumulative analyses of ap-Gal4>UAS-Tap42RNAi wing discs indicate that Tap42's modulation of HH, DPP, and JNK signaling is required for normal wing imaginal disc development.

Bottom Line: RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults.The Tap42(RNAi)-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects.The experimental platform described herein identifies crucial roles for Tap42•phosphatase complexes in governing imaginal disc and fly development.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee, United States of America.

ABSTRACT
Protein ser/thr phosphatase 2A family members (PP2A, PP4, and PP6) are implicated in the control of numerous biological processes, but our understanding of the in vivo function and regulation of these enzymes is limited. In this study, we investigated the role of Tap42, a common regulatory subunit for all three PP2A family members, in the development of Drosophila melanogaster wing imaginal discs. RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults. Silencing of Tap42 also altered multiple signaling pathways (HH, JNK and DPP) and triggered apoptosis in wing imaginal discs. The Tap42(RNAi)-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects. The experimental platform described herein identifies crucial roles for Tap42•phosphatase complexes in governing imaginal disc and fly development.

Show MeSH
Related in: MedlinePlus