Complex formation by the Drosophila MSL proteins: role of the MSL2 RING finger in protein complex assembly

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Abstract
Drosophila MSL proteins are thought to act within a complex to elevate transcription from the male X chromosome. We found that the MSL1, MSL2 and MSL3 proteins are associated in immunoprecipitations, chromatographic steps and in the yeast two‐hybrid system, but that the MLE protein is not tightly complexed in these assays. We focused our analysis on the MSL2–MSL1 interaction, which is postulated to play a critical role in MSL complex association with the X chromosome. Using a modified two‐hybrid assay, we isolated missense mutations in MSL2 that disrupt its interaction with MSL1. Eleven out of 12 mutated residues clustered around the first zinc‐binding site of the RING finger domain were conserved in a Drosophila virilis MSL2 homolog. Two pre‐existing msl2 alleles, which fail to support male viability in vivo , have lesions in the same region of the RING finger. We tested these in the two‐hybrid system and found that they are also defective in interaction with MSL1. Mutation of the second zinc‐binding site had little effect on MSL1 binding, suggesting that this portion of the RING finger may have a distinct function. Our data support a model in which MSL2–MSL1 interaction nucleates assembly of an MSL complex, with which MLE is weakly or transiently associated.