Interaction in vitro; bait produced as a recombinant fusion protein in baculovirus and Sf9 cell system; prey produced and labeled by in vitro transcription.

mle binds double-stranded RNA more strongly than single-stranded RNA. Deletion of the C-terminal G domain of mle increases its interaction with single-stranded RNA.

RNA-protein interactions were characterized by GRNA chromatography. roX2 RNA was tagged with the boxB RNA element, and bound to glutathione beads by virtue of the interaction between boxB and a GST fusion to the λ[[N22]] peptide. This resin was incubated with nuclear extracts, bound proteins were eluted by RNaseA treatment, and characterized by western blot.

Interaction in vitro; bait produced by in vitro transcription; prey derived from wild-type embryonic nuclear extract.

In this assay, roX2 RNA pulled down mle but not msl-2.

Binding modes of mle to roX2 differ for different roX2 regions: binding of mle to the roX2 region containing R2H1 and R2H2/3 is ATP-independent, but binding to the roX2 region containing R2H4, R2H5 and R2H6 stem loops does require ATP.

RNA-protein interactions were characterized by GRNA chromatography. roX2 RNA was tagged with the boxB RNA element, and bound to glutathione beads by virtue of the interaction between boxB and a GST fusion to the λ[[N22]] peptide. This resin was incubated with nuclear extracts, bound proteins were eluted by RNaseA treatment, and characterized by western blot.

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

In this assay, roX2 RNA pulled down mle but not msl-2.

RNA-protein interactions were characterized by GRNA chromatography. roX2 RNA was tagged with the boxB RNA element, and bound to glutathione beads by virtue of the interaction between boxB and a GST fusion to the λ[[N22]] peptide. This resin was incubated with nuclear extracts, bound proteins were eluted by RNaseA treatment, and characterized by western blot.

Interaction in vitro; bait produced by in vitro transcription; prey derived from transgenic embryonic nuclear extract.

Binding modes of mle to roX2 differ for different roX2 regions: binding of mle to the roX2 region containing R2H1 and R2H2/3 is ATP-independent, but binding to the roX2 region containing R2H4, R2H5 and R2H6 stem loops does require ATP.

RNA-protein interactions were characterized using the iCLIP protcol (cross-linking and immunoprecipitation). RNA was UV-crosslinked to protein, isolated by immunoprecipitation using mle antibody, and characterized by deep sequencing analysis.

Source was cell extract of clone8 cell line; bait produced from endogenous gene; prey produced from endogenous gene.

Though mle bound to 2,447 sites, roX1 and roX2 sites scored considerably higher, representing over three quarters of all observed cross-linking events.

RNA bait was tagged with MS2 stem loops, and immobilized on amylose beads using a fusion of MS2-binding protein and maltose binding protein (MBP).

mle binds to a conserved stem loop in roX1 and roX2.

ATP improves the binding affinity and specificity of mle for the conserved roX1/roX2 stem loop.

Interaction in vitro; bait produced by in vitro transcription; prey derived from L2-4 (SL2 cell line derivative) nuclear extract.

RNA bait was tagged with MS2 stem loops, and immobilized on amylose beads using a fusion of MS2-binding protein and maltose binding protein (MBP).

Interaction in vitro; bait produced by in vitro transcription; prey produced as a recombinant fusion protein in baculovirus-infected Sf21 cell system.

Binding and specificity of mle for roX2 is stimulated by ATP.

Radiolabeled roX2 RNA was probed by chemical and enzymatic footprinting in the presence or absence of mle.

Interaction in vitro; bait produced by in vitro transcription; prey produced as a recombinant fusion protein in baculovirus-infected Sf21 cell system.

The roX2 stem is disrupted by mle in the presence of ATP.

mle binds and remodels many regions of the roX2 RNA, in addition to the SLroX2 stem loop.

Interaction in vitro; bait produced by in vitro transcription; prey produced as a recombinant fusion protein in baculovirus-infected Sf21 cell system.

In the presence of ATP, mle forms higher order complexes with roX2.

Interaction in vitro; bait produced as a recombinant fusion protein in baculovirus-infected Sf21 cell system; prey produced by in vitro transcription.

In the presence of ATP, mle forms higher order complexes with roX2.

Interaction in vitro; bait produced by in vitro transcription; prey produced as a recombinant fusion protein in baculovirus-infected Sf21 cell system.

The roX2-mle interaction is stimulated by the addition of Unr.

Factors were detected on polytene chromosomes using antibodies (to proteins) or biotinylated antisense probes (to ncRNA). The PLA signal develops only if probes to the two factors are in close proximity to each other (within tens of nanometers).

Source was larval salivary gland of wild-type fly line; proteins produced from endogenous gene.

Interaction in vitro; bait produced by in vitro transcription; prey derived from extract of transfected HEK293 cells.

mle interacts more strongly with the 3-prime end of roX2 than the 5-prime end in the presence of ATP.

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

Kd = 0.20uM

mle dsRBD1,2 bound RNA with higher affinity than either individual RBD.

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

In presence of ATP, mle significantly enriched lncRNA:roX2 from the S2 transcriptome

Interaction in vitro. Source was cell extract of a male S2 cell line; bait produced as a recombinant fusion protein in bacterial system; prey produced from endogenous gene.

In presence of ATP, mle significantly enriched lncRNA:roX2 from the S2 transcriptome

Interaction in vitro. Source was cell extract of a clone 8 cell line; bait produced as a recombinant fusion protein in bacterial system; prey produced from endogenous gene.