The bxd pre from bithorax complex of Drosophila melanogaster has weak insulator activity

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Abstract

The autonomous functioning of regulatory domains in the Bithorax complex (BX-C) of Drosophila melanogaster is maintained by boundaries (insulators) that prevent inappropriate enhancer–promoter interactions. Polycomb response elements (PREs) maintain epigenetic memory within regulatory domains and are often located near insulators, enhancing their blocking activity. The bxd PRE is a well-characterized silencer that regulates the bxd/pbx domain of the Ubx gene in the first abdominal segment. We used a boundary replacement strategy to assess the insulator activity of the bxd PRE. Substituting the bxd PRE for the Fab-7 boundary, which separates the Abd-B regulatory domains in the BX-C, revealed that the bxd PRE has weak insulator activity.

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About the authors

А. N. Ibragimov

Institute of gene biology, Russian academy of science

Author for correspondence.
Email: airat.ibra@gmail.com
Russian Federation, Moscow

O. V. Kyrchanova

Institute of gene biology, Russian academy of science

Email: airat.ibra@gmail.com
Russian Federation, Moscow

Y. Е. Vorontsova

Institute of gene biology, Russian academy of science

Email: airat.ibra@gmail.com
Russian Federation, Moscow

E. N. Kozlov

Institute of gene biology, Russian academy of science

Email: airat.ibra@gmail.com
Russian Federation, Moscow

V. O. Dubrovskaya

Institute of gene biology, Russian academy of science

Email: airat.ibra@gmail.com
Russian Federation, Moscow

P. G. Georgiev

Institute of gene biology, Russian academy of science

Email: airat.ibra@gmail.com

Academician of the RAS

Russian Federation, Moscow

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Supplementary files

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2. Fig. 1. Scheme of the Bithorax complex. The curly brackets of different colors indicate the regulatory domains, the gray vertical lines are the domain boundaries, the arrows indicate homeotic genes. The pink rectangles indicate the embryonic enhancers in the abx/bx and bxd/pbx domains, the green rectangles indicate the PRE. The region containing the bxd PRE is shown on the left below. The blue ovals indicate the binding sites of the Pho protein, the yellow ones indicate the GAF protein. The scheme of the deletion at the Fab-7 boundary in the Fab-7attP50 line is shown on the right. The lilac rectangles indicate the sites of hypersensitivity to DNase I. The gray arrow indicates the attP site for integration of the studied sequence, the triangles indicate the lox and frt sites for excision of the plasmid body and the rosy marker.

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3. Fig. 2. Results of replacing the Fab-7 boundary with bxd PRE derivatives. The bxd PRE fragments used for integration into the Fab-7attP50 platform are shown at the top. The green rectangles indicate the DNase I hypersensitive sites (HS1, HS2), and the magenta rectangles indicate the regions of the bxd s1 and s2 enhancers. The binding sites of the Pho and GAF ​​proteins are indicated by yellow and blue ovals, respectively. Cuticular preparations of abdominal segments of wild-type (wt), Fab-7attP50, bxd PRE1090, bxd PREprox, bxd PREdist, bxd PREdist mGAGA, and bxd PREdist mPho flies. The upper panel shows the morphology of abdominal segments A3–A6 (numbered) in the light field. Red arrows indicate deviations from the normal phenotype. Lower panel: dark-field morphology of tergites A5 and A6.

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