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First published online 11 June 2008
doi: 10.1242/dev.023275

Development 135, 2383-2390 (2008)
Published by The Company of Biologists 2008
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Association of trxG and PcG proteins with the bxd maintenance element depends on transcriptional activity

Svetlana Petruk1, Sheryl T. Smith2, Yurii Sedkov1 and Alexander Mazo1,*

1 Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA.
2 Department of Biology, Arcadia University, Glenside, PA 19038, USA.


Figure 1
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  		Fig. 1. Multiple trxG genes are essential for functioning of the bxd TRE/PRE maintenance element. (A) The effects of ash122 and Asx3 on the eye color of {Delta}C1 transgenic flies. (B) Map of trxG and PcG response elements in the bxd region of Ubx based on the results of the genetic white tests shown in Tables 1 and 2. Data from Table 1: genes that interact or do not interact with the bxd N constructs are shown in red and black, respectively, above the map. Data from Table 2: a scheme of the {Delta}C constructs used in these experiments is shown at the bottom. The B TRE has been deleted in the constructs because of redundancy with the C TRE (Tillib et al., 1999Go). Mapped Trx, Asx, Ash1 and Brm response elements are in red. Previously mapped PcG response elements are in green (Tillib et al., 1999Go).

 

Figure 2
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  		Fig. 2. In any one cell in a Drosophila salivary gland, trxG and PcG proteins alternatively associate with the bxd ME. (A) Variegating phenotypes of the white reporter gene of the 18-15 transgenic line in the salivary gland. DAPI staining of the same glands is shown in the lower panels. (B) Binding of trxG proteins Trx, Asx, Ash1 and Kis to chromosome 3 of the salivary gland polytene chromosomes of the wild type and 18-15 transgenic line. The site of insertion of the N construct is at the very tip of chromosome 3 and is indicated by arrowheads. Trx, Asx and Ash1, but not Kis, bind simultaneously to the site of insertion of the N construct in ~50% of nuclei from the salivary glands prepared from the same larvae. (C) PcG proteins Ph, Pc and E(z) bind to the site of insertion of the N construct in those nuclei where Trx protein is not associated with this site.

 

Figure 3
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  		Fig. 3. Association of trxG or PcG proteins with the bxd ME in individual cells correlates with an activated or repressed white reporter gene, respectively. (A) Pol II phoshorylated at Ser5 is associated with the site of insertion of the N construct (arrowheads) in the same Drosophila salivary gland cells as Trx (left), but not Pc (right). (B) H3-meK27 is not associated with the site of insertion of the N construct in the same cells as Trx.

 

Figure 4
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  		Fig. 4. Effect of trx RNAi on association of trxG and PcG proteins with their target genes. Salivary gland polytene chromosomes were prepared from wild-type third instar Drosophila larvae and from the line expressing trx RNAi as described (Petruk et al., 2006Go). The overall structure of these chromosomes is indistinguishable (Petruk et al., 2006Go). (A) Trx is essential for association of Ash1 and Asx. Binding of Trx (green), Ash1 (red, column 2) and Asx (red, column 3) is completely abolished in the trx RNAi line, whereas binding of the control protein, Ecdysone receptor (EcR, green) is unaffected (column 1). (B) Trx is not essential for association of the trxG BRM complex and Kis. Components of the BRM complex, Brm and Osa, and trxG protein Kis (red) bind to their sites on polytene chromosomes in a line expressing trx RNAi. Trx binding is in green. (C) Association of the PcG complexes PRC1 and PRC2 is not affected by induction of trx RNAi. The intensities and the number of bands for the components of PRC1, Pc and Ph, and the component of the PRC2, E(z) (all in red), are not significantly affected in the trx RNAi line.

 

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