A nuclear localization signal binding protein in the nucleolus

U. Thomas Meier, Günter Blobel

Research output: Contribution to journalArticle

110 Citations (Scopus)

Abstract

We used functional wild-type and mutant synthetic nuclear localization signal peptides of SV-40 T antigen cross-linked to human serum albumin (peptide conjugates) to assay their binding to proteins of rat liver nuclei on Western blots. Proteins of 140 and 55 kD (p140 and p55) were exclusively recognized by wild-type peptide conjugates. Free wild-type peptides competed for the wild-type peptide conjugate binding to p140 and p55 whereas free mutant peptides, which differed by a single amino acid from the wild type, competed less efficiently. The two proteins were extractable from nuclei by either low or high ionic strength buffers. We purified p140 and raised polyclonal antibodies in chicken against the protein excised from polyacrylamide gels. The anti-p140 antibodies were monospecific as judged by their reactivity with a single nuclear protein band of 140 kD on Western blots of subcellular fractions of whole cells. Indirect immunofluorescence microscopy on fixed and permeabilized Buffalo rat liver (BRL) cells with anti-p140 antibodies exhibited a distinct punctate nucleolar staining. Rhodamine-labeled wild-type peptide conjugates also bound to nucleoli in a similar pattern on fixed and permeabilized BRL cells. Based on biochemical characterization, p140 is a novel nucleolar protein. It is possible that p140 shuttles between the nucleolus and the cytoplasm and functions as a nuclear import carrier.

Original languageEnglish (US)
Pages (from-to)2235-2245
Number of pages11
JournalJournal of Cell Biology
Volume111
Issue number6 PART 1
StatePublished - Dec 1990

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alpha Karyopherins
Peptides
Buffaloes
Nuclear Proteins
Anti-Idiotypic Antibodies
Liver
Western Blotting
Nuclear Localization Signals
Proteins
Rhodamines
Subcellular Fractions
Cell Nucleus Active Transport
Viral Tumor Antigens
Indirect Fluorescent Antibody Technique
Fluorescence Microscopy
Serum Albumin
Osmolar Concentration
Chickens
Carrier Proteins
Buffers

ASJC Scopus subject areas

  • Cell Biology

Cite this

A nuclear localization signal binding protein in the nucleolus. / Meier, U. Thomas; Blobel, Günter.

In: Journal of Cell Biology, Vol. 111, No. 6 PART 1, 12.1990, p. 2235-2245.

Research output: Contribution to journalArticle

Meier, UT & Blobel, G 1990, 'A nuclear localization signal binding protein in the nucleolus', Journal of Cell Biology, vol. 111, no. 6 PART 1, pp. 2235-2245.
Meier, U. Thomas ; Blobel, Günter. / A nuclear localization signal binding protein in the nucleolus. In: Journal of Cell Biology. 1990 ; Vol. 111, No. 6 PART 1. pp. 2235-2245.
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