A Glycine-Rich Domain of hnRNP H/F Promotes Nucleocytoplasmic Shuttling and Nuclear Import through an Interaction with Transportin 1

Abstract

Heterogeneous nuclear ribonucleoprotein (hnRNP) H and F are members of a closely related subfamily of hnRNP proteins that are implicated in many aspects of RNA processing. hnRNP H and F are alternative splicing factors for numerous U2- and U12-dependent introns. The proteins have three RNA binding domains and two glycine-rich domains and localize to both the nucleus and cytoplasm, but little is known about which domains govern subcellular localization or splicing activity. We show here that the central glycine-tyrosine-arginine-rich (GYR) domain is responsible for nuclear localization, and a nonclassical nuclear localization signal (NLS) was mapped to a short, highly conserved sequence whose activity was compromised by point mutations. Glutathione S-transferase (GST) pulldown assays demonstrated that the hnRNP H NLS interacts with the import receptor transportin 1. Finally, we show that hnRNP H/F are transcription-dependent shuttling proteins. Collectively, the results suggest that hnRNP H and F are GYR domain-dependent shuttling proteins whose posttranslational modifications may alter nuclear localization and hence function.

Supplemental material for this article may be found at http://mcb.asm.org/.

We are grateful to S. Munroe, A. Hudson, and members of the McNally lab for helpful comments on the manuscript and to J. Steitz (Yale University) and G. Dreyfuss (University of Pennsylvania) for plasmids. We acknowledge the Cell Culture Center in Minneapolis, MN, which provided HeLa cells for some nuclear extract preparations.

This work was supported by Public Health Service grant R01 CA78709 from the National Cancer Institute to M.T.M.