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Trafficking through COPII stabilises cell polarity and drives secretion during Drosophila epidermal differentiation.

Norum M, Tång E, Chavoshi T, Schwarz H, Linke D, Uv A, Moussian B - PLoS ONE (2010)

Bottom Line: We are studying the molecular mechanisms that Drosophila tracheal and epidermal cells deploy to form their specific apical ECM during differentiation.In this work we demonstrate that the two genetically identified factors haunted and ghost are essential for polarity maintenance, membrane topology as well as for secretion of the tracheal luminal matrix and the cuticle.Taken together, epithelial differentiation during Drosophila embryogenesis is a concerted action of ECM formation, plasma membrane remodelling and maintenance of cell polarity that all three rely mainly, if not absolutely, on the canonical secretory pathway from the ER over the Golgi apparatus to the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Interfaculty Institute for Cell Biology, University of Tübingen, Tübingen, Germany.

ABSTRACT

Background: The differentiation of an extracellular matrix (ECM) at the apical side of epithelial cells implies massive polarised secretion and membrane trafficking. An epithelial cell is hence engaged in coordinating secretion and cell polarity for a correct and efficient ECM formation.

Principal findings: We are studying the molecular mechanisms that Drosophila tracheal and epidermal cells deploy to form their specific apical ECM during differentiation. In this work we demonstrate that the two genetically identified factors haunted and ghost are essential for polarity maintenance, membrane topology as well as for secretion of the tracheal luminal matrix and the cuticle. We show that they code for the Drosophila COPII vesicle-coating components Sec23 and Sec24, respectively, that organise vesicle transport from the ER to the Golgi apparatus.

Conclusion: Taken together, epithelial differentiation during Drosophila embryogenesis is a concerted action of ECM formation, plasma membrane remodelling and maintenance of cell polarity that all three rely mainly, if not absolutely, on the canonical secretory pathway from the ER over the Golgi apparatus to the plasma membrane. Our results indicate that COPII vesicles constitute a central hub for these processes.

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Molecular identification of hau.The hau mutations were mapped to the deficiency Df(3R)ED5187 that uncovers two genes: sec23 and elm (A). Sec23 is a component of the COPII complex. The elm gene plays a role in memory formation and ethanol sensitivity, and mutations in this gene are not lethal [66]. A nonsense mutation was identified in the sec23-coding region of the 9G14 allele, and a frame shift mutation was identified in the same coding region of the CK allele (B). Sec23 is characterised by five motifs. From the N-terminus to the C-terminus, these are the Zn binding domain, the Sec23/24 domain, which belongs to the von Willebrand factor type A (vWFA) domain family, a β-sandwich domain, a helical domain and finally a Gelsolin domain. Molecular identification of gho. The gho mutations were mapped to the interval framed by the break points of the deficiencies Df(2L)BSC688 and Df(2L)Excel7010 (C). Among the 16 genes in this region, one codes for a Sec24-like protein, CG10882 (D). In the coding region of this gene, we identified one early nonsense mutation in each allele, IB104 and IP107.
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pone-0010802-g011: Molecular identification of hau.The hau mutations were mapped to the deficiency Df(3R)ED5187 that uncovers two genes: sec23 and elm (A). Sec23 is a component of the COPII complex. The elm gene plays a role in memory formation and ethanol sensitivity, and mutations in this gene are not lethal [66]. A nonsense mutation was identified in the sec23-coding region of the 9G14 allele, and a frame shift mutation was identified in the same coding region of the CK allele (B). Sec23 is characterised by five motifs. From the N-terminus to the C-terminus, these are the Zn binding domain, the Sec23/24 domain, which belongs to the von Willebrand factor type A (vWFA) domain family, a β-sandwich domain, a helical domain and finally a Gelsolin domain. Molecular identification of gho. The gho mutations were mapped to the interval framed by the break points of the deficiencies Df(2L)BSC688 and Df(2L)Excel7010 (C). Among the 16 genes in this region, one codes for a Sec24-like protein, CG10882 (D). In the coding region of this gene, we identified one early nonsense mutation in each allele, IB104 and IP107.

Mentions: To understand the molecular roles of hau and gho in cuticle differentiation, we identified the genomic location and the sequence of both genes. Mutations in hau had previously been mapped to the right arm of chromosome 3 (cytological location 85D) [35], and mutations in gho had been localised to the right arm of chromosome 2 (recombination map position 68) [24]. By deficiency mapping, we localised hau to the cytological interval between 83B7 and 83B8 uncovered by the deficiency Df(3R)ED5187 on the right arm of chromosome 3 (Figure 11A), and gho to the cytological interval between 22D4 and 22D6 defined by the overlapping region of the deficiencies Df(2L)Excel7010 and Df(2L)BSC688 on the left arm of chromosome 2 (Figure 11C). Thus, the mapping data in [24] and [35] are inaccurate. The hau-containing interval harbours two genes, one of which is CG1250 that encodes the only Drosophila Sec23 ortholog, that, as a COPII component, is involved in vesicle budding from the ER [36]. We sequenced the sec23 genomic DNA of embryos homozygous mutant for hau and detected a point mutation in each of our alleles (Figure 11B). A transition of the C643 to T resulting in a nonsense mutation changing Gln215 to amber (TAG) was detected in the hau9G14 allele. This mutation disrupts the Sec23/24 or von Willebrand factor type A (vWFA)-like domain in the first half of the protein and deletes all consecutive domains. The P-element induced allele [35] has a deletion of G2076 of the coding sequence causing a frame shift that changes the peptide sequence after Lys692. This mutation leads to the elimination of Arg722, which is essential for the interaction of Sec23 with Sar1, the GTPase that triggers vesicle budding from the ER [37], [38]. The Drosophila Sec23 protein is over its entire length 73% identical and 85% similar to the human Sec23 protein (isoform A).


Trafficking through COPII stabilises cell polarity and drives secretion during Drosophila epidermal differentiation.

Norum M, Tång E, Chavoshi T, Schwarz H, Linke D, Uv A, Moussian B - PLoS ONE (2010)

Molecular identification of hau.The hau mutations were mapped to the deficiency Df(3R)ED5187 that uncovers two genes: sec23 and elm (A). Sec23 is a component of the COPII complex. The elm gene plays a role in memory formation and ethanol sensitivity, and mutations in this gene are not lethal [66]. A nonsense mutation was identified in the sec23-coding region of the 9G14 allele, and a frame shift mutation was identified in the same coding region of the CK allele (B). Sec23 is characterised by five motifs. From the N-terminus to the C-terminus, these are the Zn binding domain, the Sec23/24 domain, which belongs to the von Willebrand factor type A (vWFA) domain family, a β-sandwich domain, a helical domain and finally a Gelsolin domain. Molecular identification of gho. The gho mutations were mapped to the interval framed by the break points of the deficiencies Df(2L)BSC688 and Df(2L)Excel7010 (C). Among the 16 genes in this region, one codes for a Sec24-like protein, CG10882 (D). In the coding region of this gene, we identified one early nonsense mutation in each allele, IB104 and IP107.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2875407&req=5

pone-0010802-g011: Molecular identification of hau.The hau mutations were mapped to the deficiency Df(3R)ED5187 that uncovers two genes: sec23 and elm (A). Sec23 is a component of the COPII complex. The elm gene plays a role in memory formation and ethanol sensitivity, and mutations in this gene are not lethal [66]. A nonsense mutation was identified in the sec23-coding region of the 9G14 allele, and a frame shift mutation was identified in the same coding region of the CK allele (B). Sec23 is characterised by five motifs. From the N-terminus to the C-terminus, these are the Zn binding domain, the Sec23/24 domain, which belongs to the von Willebrand factor type A (vWFA) domain family, a β-sandwich domain, a helical domain and finally a Gelsolin domain. Molecular identification of gho. The gho mutations were mapped to the interval framed by the break points of the deficiencies Df(2L)BSC688 and Df(2L)Excel7010 (C). Among the 16 genes in this region, one codes for a Sec24-like protein, CG10882 (D). In the coding region of this gene, we identified one early nonsense mutation in each allele, IB104 and IP107.
Mentions: To understand the molecular roles of hau and gho in cuticle differentiation, we identified the genomic location and the sequence of both genes. Mutations in hau had previously been mapped to the right arm of chromosome 3 (cytological location 85D) [35], and mutations in gho had been localised to the right arm of chromosome 2 (recombination map position 68) [24]. By deficiency mapping, we localised hau to the cytological interval between 83B7 and 83B8 uncovered by the deficiency Df(3R)ED5187 on the right arm of chromosome 3 (Figure 11A), and gho to the cytological interval between 22D4 and 22D6 defined by the overlapping region of the deficiencies Df(2L)Excel7010 and Df(2L)BSC688 on the left arm of chromosome 2 (Figure 11C). Thus, the mapping data in [24] and [35] are inaccurate. The hau-containing interval harbours two genes, one of which is CG1250 that encodes the only Drosophila Sec23 ortholog, that, as a COPII component, is involved in vesicle budding from the ER [36]. We sequenced the sec23 genomic DNA of embryos homozygous mutant for hau and detected a point mutation in each of our alleles (Figure 11B). A transition of the C643 to T resulting in a nonsense mutation changing Gln215 to amber (TAG) was detected in the hau9G14 allele. This mutation disrupts the Sec23/24 or von Willebrand factor type A (vWFA)-like domain in the first half of the protein and deletes all consecutive domains. The P-element induced allele [35] has a deletion of G2076 of the coding sequence causing a frame shift that changes the peptide sequence after Lys692. This mutation leads to the elimination of Arg722, which is essential for the interaction of Sec23 with Sar1, the GTPase that triggers vesicle budding from the ER [37], [38]. The Drosophila Sec23 protein is over its entire length 73% identical and 85% similar to the human Sec23 protein (isoform A).

Bottom Line: We are studying the molecular mechanisms that Drosophila tracheal and epidermal cells deploy to form their specific apical ECM during differentiation.In this work we demonstrate that the two genetically identified factors haunted and ghost are essential for polarity maintenance, membrane topology as well as for secretion of the tracheal luminal matrix and the cuticle.Taken together, epithelial differentiation during Drosophila embryogenesis is a concerted action of ECM formation, plasma membrane remodelling and maintenance of cell polarity that all three rely mainly, if not absolutely, on the canonical secretory pathway from the ER over the Golgi apparatus to the plasma membrane.

View Article: PubMed Central - PubMed

Affiliation: Interfaculty Institute for Cell Biology, University of Tübingen, Tübingen, Germany.

ABSTRACT

Background: The differentiation of an extracellular matrix (ECM) at the apical side of epithelial cells implies massive polarised secretion and membrane trafficking. An epithelial cell is hence engaged in coordinating secretion and cell polarity for a correct and efficient ECM formation.

Principal findings: We are studying the molecular mechanisms that Drosophila tracheal and epidermal cells deploy to form their specific apical ECM during differentiation. In this work we demonstrate that the two genetically identified factors haunted and ghost are essential for polarity maintenance, membrane topology as well as for secretion of the tracheal luminal matrix and the cuticle. We show that they code for the Drosophila COPII vesicle-coating components Sec23 and Sec24, respectively, that organise vesicle transport from the ER to the Golgi apparatus.

Conclusion: Taken together, epithelial differentiation during Drosophila embryogenesis is a concerted action of ECM formation, plasma membrane remodelling and maintenance of cell polarity that all three rely mainly, if not absolutely, on the canonical secretory pathway from the ER over the Golgi apparatus to the plasma membrane. Our results indicate that COPII vesicles constitute a central hub for these processes.

Show MeSH
Related in: MedlinePlus