Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced and labeled by in vitro transcription.

Unr binding to msl-2 RNA is weak but strongly enhanced by the addition of a recombinant purified Sxl RNA-binding domains fragment.

Interaction in vitro; inhibitor produced as a recombinant fusion protein in bacterial system; enzyme target produced by in vitro transcription.

Sxl binds the msl-2 5\'UTR to promote translational initiation at the start codon of a short upstream open reading frame (uORF) in the 5\' UTR, instead of at the reporter\'s downstream initiation codon. Sxl-mediated translational inhibition requires both the Sxl binding site and the uORF. The presence of a short three codon uORF in the msl-2 5\' UTR about 24nt upstream of a Sxl motif is conserved in all 12 Drosophila species examined.

The msl-2 5\' UTR was fused to a luciferase reporter and translated using a Drosophila embryo translation extract in the presence or absence of recombinant Sxl. Translation was determined by measuring luciferase activity in the reaction after translation.

Positive control.

Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey derived from wild-type embryonic extract.

Interaction in vitro; protein produced as recombinant fusion protein in bacterial system; mRNA produced by in vitro transcription.

Structure of Sxl, Unr, msl-2 ternary complex shown. Captured as two protein-mRNA interactions since Sxl and Unr do not interact in the absence of mRNA.

Interaction in vitro; protein produced as recombinant fusion protein in bacterial system; mRNA produced by in vitro transcription.

Ternanry Sxl, Unr, msl-2 complex analyzed. Captured as two protein-mRNA interactions since Sxl and Unr do not interact in the absence of mRNA.

Recombinant GST-Sxl RNA binding domain 4 was added to RNA affinity column along with embryo extract.

Interaction in vitro; bait produced and labeled by in vitro transcription; prey produced as a recombinant fusion protein in bacterial system.

Source was embryos of wild-type flies; bait produced from endogenous gene; prey produced from endogenous gene.

Positive control.

Source was Kc cell line; bait produced from endogenous; prey produced from endogenous gene.

PAR-CLIP