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Molecular and Cellular Biology, March 1999, p. 2212-2219, Vol. 19, No. 3
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

A Human Sequence Homologue of Staufen Is an RNA-Binding Protein That Is Associated with Polysomes and Localizes to the Rough Endoplasmic Reticulum

Rosa María Marión,1 Puri Fortes,1,dagger Ana Beloso,1 Carlos Dotti,2 and Juan Ortín1,*

Centro Nacional de Biotecnología (CSIC), Cantoblanco, 28049 Madrid, Spain,1 and European Molecular Biology Laboratory, 69012 Heidelberg, Germany2

Received 27 May 1998/Returned for modification 7 July 1998/Accepted 11 September 1998

In the course of a two-hybrid screen with the NS1 protein of influenza virus, a human clone capable of coding for a protein with high homology to the Staufen protein from Drosophila melanogaster (dmStaufen) was identified. With these sequences used as a probe, cDNAs were isolated from a lambda  cDNA library. The encoded protein (hStaufen-like) contained four double-stranded RNA (dsRNA)-binding domains with 55% similarity and 38% identity to those of dmStaufen, including identity at all residues involved in RNA binding. A recombinant protein containing all dsRNA-binding domains was expressed in Escherichia coli as a His-tagged polypeptide. It showed dsRNA binding activity in vitro, with an apparent Kd of 10-9 M. Using a specific antibody, we detected in human cells a major form of the hStaufen-like protein with an apparent molecular mass of 60 to 65 kDa. The intracellular localization of hStaufen-like protein was investigated by immunofluorescence using a series of markers for the cell compartments. Colocalization was observed with the rough endoplasmic reticulum but not with endosomes, cytoskeleton, or Golgi apparatus. Furthermore, sedimentation analyses indicated that hStaufen-like protein associates with polysomes. These results are discussed in relation to the possible functions of the protein.

* Corresponding author. Mailing address: Centro Nacional de Biotecnología (CSIC), Cantoblanco, 28049 Madrid, Spain. Phone: 3491 585 4557. Fax: 3491 585 4506. E-mail jortin{at}cnb.uam.es.

dagger Present address: European Molecular Biology Laboratory, 69012 Heidelberg, Germany.

Molecular and Cellular Biology, March 1999, p. 2212-2219, Vol. 19, No. 3
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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