Variable Nuclear Cytoplasmic Distribution of the 11.5-kDa Zinc-binding Protein (Parathymosin-) and Identification of a Bipartite Nuclear Localization Signal

doi:10.1074/jbc.271.2.1187

Variable Nuclear Cytoplasmic Distribution of the 11.5-kDa Zinc-binding Protein (Parathymosin-) and Identification of a Bipartite Nuclear Localization Signal (*)

From the Abteilung Klinische Biochemie, Zentrum Innere Medizin, Universität Göttingen, Robert-Koch-Strasse 40, D-37075 Göttingen, Federal Republic of Germany

^§ To whom correspondence should be addressed. Tel.: 49-551-392920; Fax: 49-551-392953.

Abstract

The 11.5-kDa zinc-binding protein (ZnBP, parathymosin-α), a potent inactivator of 1-phosphofructokinase, is found only in the cytoplasm of most tissues despite the presence of the putative nuclear localization signal PKRQKT. Recent reports on nuclear uptake of ZnBP could not exclude the participation of unspecific diffusion. We show here that wild-type ZnBP overexpressed in COS cells accumulates exclusively in the nucleus but that ZnBP with a mutated or deleted PKRQKT motif appears both in the nucleus and in the cytoplasm. In contrast, fusion proteins between ZnBP and parts of the endoplasmic reticulum protein calreticulin required the intact PKRQKT motif for nuclear import. The motif RKR, located nine amino acids upstream of the PKRQKT motif, is also involved in the active nuclear import of ZnBP. In contrast to rat hepatocytes and kidney cells in situ, which have ZnBP almost exclusively in the cytosol, we find ZnBP in Reuber H35 hepatoma cells and normal rat kidney cells only in the nuclei. Freshly isolated rat hepatocytes translocate their ZnBP to the nucleus in <24 h during standard cell culture conditions.

Footnotes

↵* This work was supported by Grant So 43/45-3 from the Deutsche Forschungsgemeinschaft and by grants from the graduate college “Intracellular transport of proteins and vesicles” and the Fond der Chemischen Industrie (to H. D. S.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

ZnBP

11.5-kDa zinc-binding protein (parathymosin-α)

NLS

nuclear localization signal

ER

endoplasmic reticulum

bp

base pair(s)

WT

wild-type

PCR

polymerase chain reaction

FCS

fetal calf serum

NRK

normal rat kidney.
- Received July 31, 1995.
- Revision received October 24, 1995.