Abstract
Helicases are ubiquitous enzymes that unwind or remodel single or double-stranded nucleic acids, and that participate in a vast array of metabolic pathways. The ATP-dependent DEXH-box RNA/DNA helicase MLE was first identified as a core member of the chromatin remodeling MSL complex, responsible for dosage compensation in Drosophila males. Although this complex does not assemble in females, MLE is present. Given the multiplicity of functions attributed to its mammalian ortholog RNA helicase A, we have carried out an analysis for the purpose of determining whether MLE displays the same diversity. We have identified a number of different proteins that associate with MLE, implicating its role in specific pathways. We have documented this association in selected examples that include the spliceosome complex, heterogeneous Nuclear Ribonucleoproteins involved in RNA Processing and in Heterochromatin Protein 1 deposition, and the NuRD complex.
Highlights
As an initial approach we have carried out a mass spectrometry analysis of all of the proteins that coimmunoprecipitate with MLE in Drosophila S2 cells, when the MSL complex is present or when it is abrogated by RNA interference
MLE Interacts with a Variety of Factors Involved in Nucleic Acids Metabolism—In order to identify new interactors of MLE, we performed a series of immunoprecipitations from S2 cells expressing a FLAG-tagged MLE protein
In order to distinguish the interactions of MLE with unknown factors from its known interaction with the subunits of the MSL complex [7, 9, 10, 38] we obtained a FLAG-tagged MLE precipitate from S2 cells where the MSL complex was abrogated by RNA interference against the MSL2 subunit (MSL2kd)
Summary
RISC assembly [16, 17] to translation initiation (18 –20), or acting as a DNA-binding partner for EGFR-mediated transcription [21]. As an initial approach we have carried out a mass spectrometry analysis of all of the proteins that coimmunoprecipitate with MLE in Drosophila S2 cells, when the MSL complex is present or when it is abrogated by RNA interference. We used S2 cells in order to determine the possible relationship of the MLE fraction associated with the MSL complex with the fraction involved in other functions. We believe that the study of the role that MLE plays in selected pathways in Drosophila will be of major use in understanding the function of RNA helicase A in similar pathways in humans. We document the interaction of a selected group of proteins with MLE in order to provide preliminary evidence for its involvement in diverse regulatory pathways
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