Yeast Shuttling SR Proteins Npl3p, Gbp2p, and Hrb1p Are Part of the Translating mRNPs, and Npl3p Can Function as a Translational Repressor

Abstract

A major challenge in current molecular biology is to understand how sequential steps in gene expression are coupled. Recently, much attention has been focused on the linkage of transcription, processing, and mRNA export. Here we describe the cytoplasmic rearrangement for shuttling mRNA binding proteins in Saccharomyces cerevisiae during translation. While the bulk of Hrp1p, Nab2p, or Mex67p is not associated with polysome containing mRNAs, significant amounts of the serine/arginine (SR)-type shuttling mRNA binding proteins Npl3p, Gbp2p, and Hrb1p remain associated with the mRNA-protein complex during translation. Interestingly, a prolonged association of Npl3p with polysome containing mRNAs results in translational defects, indicating that Npl3p can function as a negative translational regulator. Consistent with this idea, a mutation in NPL3 that slows down translation suppresses growth defects caused by the presence of translation inhibitors or a mutation in eIF5A. Moreover, using sucrose density gradient analysis, we provide evidence that the import receptor Mtr10p, but not the SR protein kinase Sky1p, is involved in the timely regulated release of Npl3p from polysome-associated mRNAs. Together, these data shed light onto the transformation of an exporting to a translating mRNP.

We are grateful to P. A. Silver for antibodies, strains, and plasmids and to C. N. Cole, C. Guthrie, E. Hurt, A. Jacobson, F. Stutz, and A. M. Tartakoff for strains and plasmids. We thank U. Beyer for technical assistance and R. Müller and B. Dobberstein for support.

This work was funded by grants from Kempkes and the Deutsche Forschungsgemeinschaft to H.K. (SFB397 and SFB593). C.I.G. is supported by grants from the National Institutes of Health (KO1 HL-04355-02 and GM008102-3052).