Protein and cDNA sequence of a glycine-rich, dimethylarginine-containing region located near the carboxyl-terminal end of nucleolin (C23 and 100 kDa).

Lapeyre, Bruno; Amalríc, François; Ghaffari, Seyed H.; Rao, Shengbin; Dumbar, Tamba S.; Olson, Matthew S.

doi:10.1016/s0021-9258(18)67634-3

Cited by 103 publications

(15 citation statements)

References 42 publications

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“…The amino acid composition of the 29-kDa CNBr fragment was determined on a Beckman 119CL amino acid analyzer. For sequencing studies, rat nucleolin was digested with CNBr and fractionated by reverse-phase HPLC as previously described (Lapeyre et al, 1986). Protein sequencing was done on an Applied Biosystems 470A sequencer using the 03RPTH program.…”

Section: Methodsmentioning

confidence: 99%

Phosphorylation of nucleolin by a nucleolar type NII protein kinase

Caizergues‐Ferrer

Bélenguer²,

Lapeyre³

et al. 1987

Biochemistry

151

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Nucleolin [C23 or 100 kilodaltons (kDa)] is the major nucleolar phosphorylated protein in exponentially growing Chinese hamster ovary cells. A nucleolar cyclic nucleotide independent protein kinase copurified with nucleolin in a complex which could be dissociated by hydroxyapatite chromatography. The kinase was stimulated by spermine and inhibited by heparin and presented most of the properties of nuclear casein kinase NII. Kinetic analyses showed the apparent Km value for nucleolin (7 X 10(-4) mg/mL) to be lower than those for other casein kinase II substrates such as nuclear protein HMG 14 (0.15 mg/mL), topoisomerase I (0.025 mg/mL), or topoisomerase II (0.04 mg/mL). Similarly, Vmax values were higher for nucleolin than for other substrates. Nucleolin thus appears to be a natural preferential substrate of nucleolar casein kinase NII. The kinase phosphorylated nucleolin in vitro at serine residues in a 29-kDa CNBr fragment located near the amino terminus of the molecule. The enzyme labeled typical casein kinase II sites. These sites were found predominantly in two highly acidic tryptic fragments designated A (residues 21-49) and C (residues 180-221) which contained serines having at least two acidic residues on their carboxyl-terminal sides. These results demonstrate the existence in the nucleolus of a type of NII protein kinase that uses a protein involved in ribosome assembly as preferential substrate.

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Section: Methodsmentioning

confidence: 99%

Phosphorylation of nucleolin by a nucleolar type NII protein kinase

Caizergues‐Ferrer

Bélenguer²,

Lapeyre³

et al. 1987

Biochemistry

151

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“…Many nuclear bodies have specific marker proteins, for example nucleolin in the nucleoli or coilin for Cajal bodies [28,29]. We used immunofluorescence and co-localization analysis to investigate the spatial relationship between HIF-2a and other nuclear proteins with a view to determining whether HIF-2a is localizing at known nuclear bodies.…”

Section: Hif-2a Speckles Co-localize With Active Transcription Sitesmentioning

confidence: 99%

Differential sub-nuclear distribution of hypoxia-inducible factors (HIF)-1 and -2 alpha impacts on their stability and mobility

Taylor¹,

Bagnall²,

Mason³

et al. 2016

Open Biol.

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Cellular adaptation to hypoxia occurs via a complex programme of gene expression mediated by the hypoxia-inducible factor (HIF). The oxygen labile alpha subunits, HIF-1α/-2α, form a heterodimeric transcription factor with HIF-1β and modulate gene expression. HIF-1α and HIF-2α possess similar domain structure and bind to the same consensus sequence. However, they have different oxygen-dependent stability and activate distinct genes. To better understand these differences, we used fluorescent microscopy to determine precise localization and dynamics. We observed a homogeneous distribution of HIF-1α in the nucleus, while HIF-2α localized into speckles. We demonstrated that the number, size and mobility of HIF-2α speckles were independent of cellular oxygenation and that HIF-2α molecules were capable of exchanging between the speckles and nucleoplasm in an oxygen-independent manner. The concentration of HIF-2α into speckles may explain its increased stability compared with HIF-1α and its slower mobility may offer a mechanism for gene specificity.

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“…A number of RNA binding proteins, including the hnRNP proteins (Chung & Wooley, 1986; Cobianchi et al, 1988), the U1 RNA binding protein (Theissen et al, 1986), or the poly(A) binding protein (Adam et al, 1986;Sachs et al, 1986) and nucleolin (Lapeyre et al, 1986(Lapeyre et al, , 1987, share sequence elements in common.…”

mentioning

confidence: 99%

Interaction of nucleolar phosphoprotein B23 with nucleic acids

Dumbar¹,

Gentry²,

Olson³

1989

Biochemistry

153

111

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The interaction of eukaryotic nucleolar phosphoprotein B23 with nucleic acids was examined by gel retardation and filter binding assays, by fluorescence techniques, and by circular dichroism. All studies utilized protein prepared under native conditions by a newly developed purification procedure. Electrophoretic gel mobility shift assays with phage M13 DNA suggested that protein B23 is a single-stranded nucleic acid binding protein. This was confirmed in competition binding assays with native or heat-denatured linearized plasmid pUC18 DNA where the protein showed a marked preference for the denatured form. In other competition assays, there was no apparent preference for single-stranded synthetic ribo- versus deoxyribonucleotides. Equilibrium binding with poly(riboethenoadenylic acid) indicated cooperative ligand binding with a protein binding site size of 11 nucleotides and an apparent binding constant (K omega) of 5 x 10(7) M-1 which includes an intrinsic binding constant (K) of 6.3 x 10(4) M-1 and a cooperativity factor (omega) of 800. In circular dichroism (CD) studies, protein B23, when combined with the single-stranded synthetic nucleic acids poly(rA) and poly(rC), effected a decrease in ellipticity and a shift of the positive peak at 260-270 nm toward higher wavelengths, indicating helix destabilizing activity. No CD changes were seen with double-stranded poly(dA.dT). The change in ellipticity of poly(rA) was sigmoidal upon addition of protein, confirming the cooperative behavior seen with fluorescence methods. These studies indicate that protein B23 binds cooperatively with high affinity for single-stranded nucleic acids and exhibits RNA helix destabilizing activity. These features may be related to its role in ribosome assembly.

show abstract

Protein and cDNA sequence of a glycine-rich, dimethylarginine-containing region located near the carboxyl-terminal end of nucleolin (C23 and 100 kDa).

Cited by 103 publications

References 42 publications

Phosphorylation of nucleolin by a nucleolar type NII protein kinase

Phosphorylation of nucleolin by a nucleolar type NII protein kinase

Differential sub-nuclear distribution of hypoxia-inducible factors (HIF)-1 and -2 alpha impacts on their stability and mobility

Interaction of nucleolar phosphoprotein B23 with nucleic acids

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