A Novel Murine Staufen Isoform Modulates the RNA Content of Staufen Complexes

Abstract

Mouse Staufen (mStau) is a double-stranded RNA-binding protein associated with polysomes and the rough endoplasmic reticulum (RER). We describe a novel endogenous isoform of mStau (termed mStauⁱ) which has an insertion of six amino acids within dsRBD3, the major double-stranded RNA (dsRNA)-binding domain. With a structural change of the RNA-binding domain, this conserved and widely distributed isoform showed strongly impaired dsRNA-binding ability. In transfected cells, mStauⁱ exhibited the same tubulovesicular distribution (RER) as mStau when weakly expressed; however, when overexpressed, mStauⁱ was found in large cytoplasmic granules. Markers of the RER colocalized with mStauⁱ-containing granules, showing that overexpressed mStauⁱ could still be associated with the RER. Cotransfection of mStauⁱ with mStau relocalized overexpressed mStauⁱ to the reticular RER, suggesting that they can form a complex on the RER and that a balance between these isoforms is important to achieve proper localization. Coimmunoprecipitation demonstrated that the two mStau isoforms are components of the same complex in vivo. Analysis of the immunoprecipitates showed that mStau is a component of an RNA-protein complex and that the association with mStauⁱ drastically reduces the RNA content of the complex. We propose that this new isoform, by forming a multiple-isoform complex, regulates the amount of RNA in mStau complexes in mammalian cells.

ACKNOWLEDGMENTS

We thank Michael Kiebler and Gopal Subramaniam for critical reading of the manuscript, Judith Kashul for editing the manuscript, Frédéric Brizard for sharing unpublished data, Louise Cournoyer for help with tissue culture, and Danny Baranes for help with confocal microscopy. We also thank Michael Kiebler and Kavish Hemraj for help and support.

This work was supported by a Natural Sciences and Engineering Research Council of Canada (NSERC) grant to L.D. and a Medical Research Council of Canada grant to I.R.N. S.V.S. is a fellow of le Fonds de la Recherche en Santé du Québec. T.D. was supported by an NSERC studentship.