The human keratinocyte two‐dimensional gel protein database (update 1995): Mapping components of signal transduction pathways

Celis, Julio E.; Rasmussen, Hanne H.; Gromov, Pavel; Olsen, Eydfinnur; Madsen, Peder; Leffers, Henrik; Honoré, Bent; Dejgaard, Kurt; Vorum, Henrik; Kristensen, Dan Bach

doi:10.1002/elps.11501601355

Cited by 118 publications

(74 citation statements)

References 36 publications

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“…p27 has previously been shown to resolve into a number of discrete spots with di erent isoelectric points upon 2D gel electrophoresis (Celis et al, 1995). In lysates from RAT1-MycER cells, p27 resolved into four major spots (labelled 1 to 4 in Figure 4a).…”

Section: Ipmentioning

confidence: 91%

Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes

Müller¹,

Bouchard²,

et al. 1997

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Activation of Myc triggers a rapid induction of cyclin E/ cdk2 kinase activity and degradation of p27. Overt degradation of p27 is preceded by a speci®c dissociation of p27 from cyclin E/cdk2, but not from cyclin D/cdk4 complexes. We now show that cyclin E/cdk2 phosphorylates p27 at a carboxy-terminal threonine residue (T187) in vitro; mutation of this residue to valine stabilises cyclin E/cdk2 complexes. This reaction is not signi®cantly inhibited by high concentrations of p27, suggesting that cdk2 bound to p27 is catalytically active. In vivo, p27 bound to cyclins E and A, but not to D-type cyclins is phosphorylated. Myc-induced release of p27 from cdk2 requires cdk2 kinase activity and is delayed in a T187V mutant of p27. After induction of Myc, p27 phosphorylated at threonine 187 transiently accumulates in a non cdk2 bound form. Our data suggest a mechanism in which p27 is released from cyclin E/cdk2 upon phosphorylation; in Myc-transformed cells, release is e cient as phosphorylated p27 is transiently bound in a non-cdk2 containing complex and subsequently degraded.

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Section: Ipmentioning

confidence: 91%

Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes

Müller¹,

Bouchard²,

et al. 1997

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“…Proteins of analytical gels were silver stained [IS]. For the calibration of data according to the twodimensional-gel protein database of human keratinocytes [19], cells and proteins were kindly supplied by Dr J. Celis Computer-assisted image analysis.…”

Section: Methodsmentioning

confidence: 99%

Identification of Human Common Nuclear‐Matrix Proteins as Heterogeneous Nuclear Ribonucleoproteins H and H′ by Sequencing and Mass Spectrometry

Holzmann

Korosec

Gerner

et al. 1997

European Journal of Biochemistry

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Nuclear-matrix proteins were prepared from different rat and human cells and separated by twodimensional gel electrophoresis. By computer-assisted analysis of the images, two of the proteins were identified as ubiquitously occurring (common) nuclear-matrix proteins, which appeared in tissue-dependent concentrations. The two proteins that originated from human blood mononuclear cells were analyzed further. Tryptic digests of the blotted proteins were analyzed by partial peptide sequencing and matrixassisted laser-desorption ionization-time-of-flight mass spectrometry. The two human common nuclearmatrix proteins were identified as heterogeneous nuclear ribonucleoproteins (hnRNP) H and H' or their variants. Furthermore, mass analysis revealed details on the N terminus of hnRNP H.Keywords: nuclear matrix ; common nuclear-matrix protein ; heterogeneous nuclear ribonucleoprotein ; tissue specificity ; peptide sequencing.The nuclear matrix is operationally defined as the residual nuclear structure that is yielded by sequential treatment of isolated nuclei with detergents, nucleases and buffers of high ionic strength 11 1. It is considered to represent the three-dimensional fibrillar protein structure constituting the framework of the interphase nucleus. In addition to its role in maintaining the nuclear architecture and the higher order structure of chromatin, the nuclear matrix has been reported as being involved in various nuclear activities, such as DNA replication, DNA transcription, RNA processing and steroid-hormone action. Studies on selected cell types and tissues have indicated that some nuclearmatrix proteins are cell and differentiation specific, while others are shared among various cell types [2]. Stuurman et al. [3] have presented evidence for the presence of a common set of polypeptides termed minimal matrix proteins in the nuclear matrices of mouse cells. Nakayasu and Berezney [4] have reported that eight proteins named nuclear matrins are major constituents of the internal nuclear matrix of rat liver cells. Belgrader et al. were the first to determine the sequence of a defined nuclear-matrix protein, matrin 3, by cloning and sequencing [5]. The detection of other common nuclear-matrix proteins, such as nuclear lamin B [3, 41, nucleolar protein B23 [3, 41, or the nuclear-mitotic apparatus protein [6, 71, was mainly based on immunological methods. However, the number and identity of common nuclearmatrix proteins have not been established unequivocally (reviewed in [8, 91).In systematic studies, we are investigating the nuclear-matrix proteins of various human and rat cells and tissues. ComputerCorrespondence to G. Sauermann,

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“…Jsing a simple expression cloning/screening procedure we ha ,e identified the human homolog of rheb, a novel member of ~;he ras-superfamily that is present in less than 20,000 molecuies per cell in normal human keratinocytes ( [24], unpublished ob ~ervation), and that exhibits interesting amino acid residues at 3ositions 15 (Arg) and 16 (Ser) that correspond to residues 12 and 13 in ras. Considering that the Glyl2--*Argl2 mutation ha, been implicated in ras-associated carcinogenesis [22,25,26], th~ occurrence of Arg at this position in rheb is intriguing.…”

Section: Discussionmentioning

confidence: 99%

A novel approach for expression cloning of small GTPases: identification, tissue distribution and chromosome mapping of the human homolog of rheb

et al. 1995

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We report a novel approach for identifying monomeric GTP-binding proteins that is based on probing cDNA expression libraries with [a-32P]GTP. In short, a nitrocellulose replica from a plated cDNA expression library is treated with 2% SDS to block the GTP-binding activity of various G proteins expressed b) E. coli, thus allowing the direct identification of positive clones. Using this procedure we have cloned several small GTP-binding proteins from human keratinocytes including the human homolog of rheb, a novel member of the ras-related GTP-hinding proteins. Human rheb cDNA shares 90% identity with the rat counterpart and it is highly upregulated in transformed human cells of various origin. Northern analysis showed that human rheb is ubiquitously expressed, with the highest levels observed in skeletal and cardiac muscle, and not in brain, as it is the case for rat rheb. The human RHEB gene was mapped to chromosome 10qll.

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The human keratinocyte two‐dimensional gel protein database (update 1995): Mapping components of signal transduction pathways

Cited by 118 publications

References 36 publications

Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes

Cdk2-dependent phosphorylation of p27 facilitates its Myc-induced release from cyclin E/cdk2 complexes

Identification of Human Common Nuclear‐Matrix Proteins as Heterogeneous Nuclear Ribonucleoproteins H and H′ by Sequencing and Mass Spectrometry

A novel approach for expression cloning of small GTPases: identification, tissue distribution and chromosome mapping of the human homolog of rheb

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