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The single Drosophila ZO-1 protein Polychaetoid regulates embryonic morphogenesis in coordination with Canoe/afadin and Enabled.

Choi W, Jung KC, Nelson KS, Bhat MA, Beitel GJ, Peifer M, Fanning AS - Mol. Biol. Cell (2011)

Bottom Line: Pyd loss does not dramatically affect AJ protein localization or initial localization of actin and myosin during dorsal closure.The defects, which include segmental grooves that fail to retract, a disrupted leading edge actin cable, and reduced zippering as leading edges meet, closely resemble defects in canoe zygotic mutants and in embryos lacking the actin regulator Enabled (Ena), suggesting that these proteins act together.Canoe (Cno) and Pyd are required for proper Ena localization during dorsal closure, and strong genetic interactions suggest that Cno, Pyd, and Ena act together in regulating or anchoring the actin cytoskeleton during dorsal closure.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, USA.

ABSTRACT
Adherens and tight junctions play key roles in assembling epithelia and maintaining barriers. In cell culture zonula occludens (ZO)-family proteins are important for assembly/maturation of both tight and adherens junctions (AJs). Genetic studies suggest that ZO proteins are important during normal development, but interpretation of mouse and fly studies is limited by genetic redundancy and/or a lack of alleles. We generated alleles of the single Drosophila ZO protein Polychaetoid (Pyd). Most embryos lacking Pyd die with striking defects in morphogenesis of embryonic epithelia including the epidermis, segmental grooves, and tracheal system. Pyd loss does not dramatically affect AJ protein localization or initial localization of actin and myosin during dorsal closure. However, Pyd loss does affect several cell behaviors that drive dorsal closure. The defects, which include segmental grooves that fail to retract, a disrupted leading edge actin cable, and reduced zippering as leading edges meet, closely resemble defects in canoe zygotic mutants and in embryos lacking the actin regulator Enabled (Ena), suggesting that these proteins act together. Canoe (Cno) and Pyd are required for proper Ena localization during dorsal closure, and strong genetic interactions suggest that Cno, Pyd, and Ena act together in regulating or anchoring the actin cytoskeleton during dorsal closure.

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pydMZ mutants have defects in LE cell shape and epithelial zippering. Stage 13–15 wild-type (WT) or pydMZ mutants, anterior left, dorsal view, antigens and embryonic stages indicated. (A, B) pydMZ mutants are largely normal through the onset of dorsal closure. (C–E, G, H) pydMZ mutants have apparent defects in zippering at the canthi (yellow arrows), leading to more oval dorsal openings, and also have a wavy leading edge. (F) pydMZ mutant in which dorsal closure failed, leading to exposure of the underlying gut and muscle (arrow) after amnioserosal cell apoptosis. (I, J) Many pydMZ mutants eventually close. (K–R) Close-ups of leading edge and amnioserosa at successively later stages. In pydMZ mutants LE cells retain the ability to elongate (K–O, green arrows), and more lateral epidermal cells can also elongate (M–O, red arrows). However, pydMZ mutants exhibit several defects, including a dorsal opening that is not “eye shaped,” a wavy margin of the sheet of epidermal cells, and broadened (L, N, O, and Q, blue arrows) or narrowed (L, N, O, and Q, white arrows) leading edges. In addition, some groups of epidermal cells fail to change shape entirely (O, bracket). Scale bars, 10 μm.
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Figure 4: pydMZ mutants have defects in LE cell shape and epithelial zippering. Stage 13–15 wild-type (WT) or pydMZ mutants, anterior left, dorsal view, antigens and embryonic stages indicated. (A, B) pydMZ mutants are largely normal through the onset of dorsal closure. (C–E, G, H) pydMZ mutants have apparent defects in zippering at the canthi (yellow arrows), leading to more oval dorsal openings, and also have a wavy leading edge. (F) pydMZ mutant in which dorsal closure failed, leading to exposure of the underlying gut and muscle (arrow) after amnioserosal cell apoptosis. (I, J) Many pydMZ mutants eventually close. (K–R) Close-ups of leading edge and amnioserosa at successively later stages. In pydMZ mutants LE cells retain the ability to elongate (K–O, green arrows), and more lateral epidermal cells can also elongate (M–O, red arrows). However, pydMZ mutants exhibit several defects, including a dorsal opening that is not “eye shaped,” a wavy margin of the sheet of epidermal cells, and broadened (L, N, O, and Q, blue arrows) or narrowed (L, N, O, and Q, white arrows) leading edges. In addition, some groups of epidermal cells fail to change shape entirely (O, bracket). Scale bars, 10 μm.

Mentions: To explore the cellular roles of Pyd, we explored the characteristic cell shape changes occurring during dorsal closure, and how they shape tissue rearrangements. Dorsal closure begins immediately after germband retraction. During the extended germband stage (stages 9–11), epidermal cells are roughly columnar, with polygonal apical cross sections. During germband retraction (stage 12), the leading edge (LE) cells (those that abut the amnioserosa) become square in apical profile, forming a straight line along the edge of the amnioserosa. As germband retraction is completed, LE cells begin to elongate along the dorsal–ventral (DV) axis (Figure 4, A and K, green arrow) and assemble at their leading edge a cable of actin and nonmuscle myosin II (myosin; Figure 3, C–G). At the same time, amnioserosal cells begin periodic apical constrictions, gradually reducing their apical areas. As constriction proceeds more ventral epidermal cells elongate (Figure 4, C and M, red arrow). Elegant laser surgery experiments demonstrate that apical constriction of amnioserosal cells and constriction of the actomyosin cable both provide the driving force for closure, whereas the epidermis resists elongation, slowing the process (Kiehart et al., 2000). The two leading edges meet first at the anterior and posterior ends of the amnioserosa, at structures termed the canthi (Figure 4C, yellow arrows), forming an “eye-shaped” dorsal opening. As the edges meet, cells meet adjoining neighbors from the other side, and the two sheets zip closed (Figure 4, G, M, and P, yellow arrows). Protrusive behavior at the leading edge is required for fully effective zippering (e.g., Woolner et al., 2005; Gates et al., 2007; Millard and Martin, 2008).


The single Drosophila ZO-1 protein Polychaetoid regulates embryonic morphogenesis in coordination with Canoe/afadin and Enabled.

Choi W, Jung KC, Nelson KS, Bhat MA, Beitel GJ, Peifer M, Fanning AS - Mol. Biol. Cell (2011)

pydMZ mutants have defects in LE cell shape and epithelial zippering. Stage 13–15 wild-type (WT) or pydMZ mutants, anterior left, dorsal view, antigens and embryonic stages indicated. (A, B) pydMZ mutants are largely normal through the onset of dorsal closure. (C–E, G, H) pydMZ mutants have apparent defects in zippering at the canthi (yellow arrows), leading to more oval dorsal openings, and also have a wavy leading edge. (F) pydMZ mutant in which dorsal closure failed, leading to exposure of the underlying gut and muscle (arrow) after amnioserosal cell apoptosis. (I, J) Many pydMZ mutants eventually close. (K–R) Close-ups of leading edge and amnioserosa at successively later stages. In pydMZ mutants LE cells retain the ability to elongate (K–O, green arrows), and more lateral epidermal cells can also elongate (M–O, red arrows). However, pydMZ mutants exhibit several defects, including a dorsal opening that is not “eye shaped,” a wavy margin of the sheet of epidermal cells, and broadened (L, N, O, and Q, blue arrows) or narrowed (L, N, O, and Q, white arrows) leading edges. In addition, some groups of epidermal cells fail to change shape entirely (O, bracket). Scale bars, 10 μm.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 4: pydMZ mutants have defects in LE cell shape and epithelial zippering. Stage 13–15 wild-type (WT) or pydMZ mutants, anterior left, dorsal view, antigens and embryonic stages indicated. (A, B) pydMZ mutants are largely normal through the onset of dorsal closure. (C–E, G, H) pydMZ mutants have apparent defects in zippering at the canthi (yellow arrows), leading to more oval dorsal openings, and also have a wavy leading edge. (F) pydMZ mutant in which dorsal closure failed, leading to exposure of the underlying gut and muscle (arrow) after amnioserosal cell apoptosis. (I, J) Many pydMZ mutants eventually close. (K–R) Close-ups of leading edge and amnioserosa at successively later stages. In pydMZ mutants LE cells retain the ability to elongate (K–O, green arrows), and more lateral epidermal cells can also elongate (M–O, red arrows). However, pydMZ mutants exhibit several defects, including a dorsal opening that is not “eye shaped,” a wavy margin of the sheet of epidermal cells, and broadened (L, N, O, and Q, blue arrows) or narrowed (L, N, O, and Q, white arrows) leading edges. In addition, some groups of epidermal cells fail to change shape entirely (O, bracket). Scale bars, 10 μm.
Mentions: To explore the cellular roles of Pyd, we explored the characteristic cell shape changes occurring during dorsal closure, and how they shape tissue rearrangements. Dorsal closure begins immediately after germband retraction. During the extended germband stage (stages 9–11), epidermal cells are roughly columnar, with polygonal apical cross sections. During germband retraction (stage 12), the leading edge (LE) cells (those that abut the amnioserosa) become square in apical profile, forming a straight line along the edge of the amnioserosa. As germband retraction is completed, LE cells begin to elongate along the dorsal–ventral (DV) axis (Figure 4, A and K, green arrow) and assemble at their leading edge a cable of actin and nonmuscle myosin II (myosin; Figure 3, C–G). At the same time, amnioserosal cells begin periodic apical constrictions, gradually reducing their apical areas. As constriction proceeds more ventral epidermal cells elongate (Figure 4, C and M, red arrow). Elegant laser surgery experiments demonstrate that apical constriction of amnioserosal cells and constriction of the actomyosin cable both provide the driving force for closure, whereas the epidermis resists elongation, slowing the process (Kiehart et al., 2000). The two leading edges meet first at the anterior and posterior ends of the amnioserosa, at structures termed the canthi (Figure 4C, yellow arrows), forming an “eye-shaped” dorsal opening. As the edges meet, cells meet adjoining neighbors from the other side, and the two sheets zip closed (Figure 4, G, M, and P, yellow arrows). Protrusive behavior at the leading edge is required for fully effective zippering (e.g., Woolner et al., 2005; Gates et al., 2007; Millard and Martin, 2008).

Bottom Line: Pyd loss does not dramatically affect AJ protein localization or initial localization of actin and myosin during dorsal closure.The defects, which include segmental grooves that fail to retract, a disrupted leading edge actin cable, and reduced zippering as leading edges meet, closely resemble defects in canoe zygotic mutants and in embryos lacking the actin regulator Enabled (Ena), suggesting that these proteins act together.Canoe (Cno) and Pyd are required for proper Ena localization during dorsal closure, and strong genetic interactions suggest that Cno, Pyd, and Ena act together in regulating or anchoring the actin cytoskeleton during dorsal closure.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina at Chapel Hill, USA.

ABSTRACT
Adherens and tight junctions play key roles in assembling epithelia and maintaining barriers. In cell culture zonula occludens (ZO)-family proteins are important for assembly/maturation of both tight and adherens junctions (AJs). Genetic studies suggest that ZO proteins are important during normal development, but interpretation of mouse and fly studies is limited by genetic redundancy and/or a lack of alleles. We generated alleles of the single Drosophila ZO protein Polychaetoid (Pyd). Most embryos lacking Pyd die with striking defects in morphogenesis of embryonic epithelia including the epidermis, segmental grooves, and tracheal system. Pyd loss does not dramatically affect AJ protein localization or initial localization of actin and myosin during dorsal closure. However, Pyd loss does affect several cell behaviors that drive dorsal closure. The defects, which include segmental grooves that fail to retract, a disrupted leading edge actin cable, and reduced zippering as leading edges meet, closely resemble defects in canoe zygotic mutants and in embryos lacking the actin regulator Enabled (Ena), suggesting that these proteins act together. Canoe (Cno) and Pyd are required for proper Ena localization during dorsal closure, and strong genetic interactions suggest that Cno, Pyd, and Ena act together in regulating or anchoring the actin cytoskeleton during dorsal closure.

Show MeSH
Related in: MedlinePlus