The primary sequence of the PFK‐1 inactivating zinc‐binding protein as deduced from cDNA sequencing Identity of the zinc‐binding protein with rat parathymosin

Trompeter, Hans-Ingo; Brand, Ingeborg A.; Söling, Hans‐Dieter

doi:10.1016/0014-5793(89)80930-5

Cited by 28 publications

(21 citation statements)

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“…This suggests either weak overlap between the FBP11 WW and FBP30 WW binding motifs, which seems unlikely based on the peptide library data in Table I (41). WBP13 represents an out-of-frame fusion with the zinc-binding protein ZnB (42), in which this frame-shifted fusion results in a proline-rich protein. The four remaining sequences (WBP14, WBP15, WBP16, and WBP17) are novel sequences, showing no significant similarity to any known protein in the data base.…”

Section: Resultsmentioning

confidence: 99%

A Novel Pro-Arg Motif Recognized by WW Domains

Bedford¹,

Sarbassova²,

Xu³

et al. 2000

Journal of Biological Chemistry

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WW domains mediate protein-protein interactions through binding to short proline-rich sequences. Two distinct sequence motifs, PPXY and PPLP, are recognized by different classes of WW domains, and another class binds to phospho-Ser-Pro sequences. We now describe a novel Pro-Arg sequence motif recognized by a different class of WW domains using data from oriented peptide library screening, expression cloning, and in vitro binding experiments. Many signaling events in eukaryotic cells involve the assembly of large protein-protein complexes. These diverse associations are mediated through interactions of a limited number of modular signaling domains such as the SH2, 1 SH3, PDZ, and PTB domains and their respective ligands (1, 2). Particular families of domains occasionally share affinity for related ligands. For example, both SH2 and PTB domains can bind phosphotyrosine-containing substrates, while FHA domains, 14-3-3 proteins, and certain WW domains bind phosphoserine/ phosphothreonine-containing proteins (3). Recently, several unrelated signaling modules have been identified that specifically recognize proline-rich ligands. These include WW and EVH1 domains in addition to the canonical member of this group, the SH3 domain (4 -7). WW domains are encoded as 38 -40-amino acid long modules in single copy or tandem repeats in over 25 signaling proteins. WW domain-containing proteins include those implicated directly or indirectly in a variety of human diseases such as Liddle's syndrome, Duchenne muscular dystrophy, Huntington's disease, and Alzheimer's disease (6,8,9). Named after two highly conserved tryptophan residues characteristically spaced either 22 or 23 residues apart, WW domains participate in a variety of cellular processes, including ubiquitin-mediated protein degradation (10, 11), viral budding (12, 13), RNA splicing (14, 15), transcriptional co-activation (16 -18), and mitotic regulation (19,20). Recently another WW domain-containing protein, FBP30, was identified as one of the major genes up-regulated by p53 in thymocytes undergoing programmed cell death following ␥-irradiation (21). Despite their general importance in normal and disease states, most of the molecular targets recognized by different WW domains in vivo remain unidentified.Sudol and colleagues (22) have proposed a classification of WW domains based on their proline-based ligand specificity. In this scheme, Group I WW domains recognize and bind the motif PPXY, as typified by the prototype WW domain in Yes-associated protein (YAP), whereas Group II WW domains bind a PPLP motif, as observed for the WW domains in formin-binding protein (FBP)-11. A third class of WW domains bind sequences rich in Pro, Gly, and Met residues, although the exact amino acid sequence motif they recognize is not known. A fourth class of WW domain-containing proteins has recently been found that appears to bind Pro residues preceded by phosphoserine (23).In the present study we use oriented peptide library screening, phage-expression cloning, peptide binding studies, ...

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Section: Resultsmentioning

confidence: 99%

A Novel Pro-Arg Motif Recognized by WW Domains

Bedford¹,

Sarbassova²,

Xu³

et al. 2000

Journal of Biological Chemistry

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“…Down-regulation of prothymosin mRNA was also observed in an HL-60 myeloid leukemia cell line induced to differentiate with either dimethyl sulfoxide or phorbol 12-myristate 13-acetate (36). In other reports, prothymosin has been shown to be covalently linked to a small cytoplasmic RNA (17), and parathymosin has been identified as the Zn2+-binding protein that reversibly inactivates phosphofructokinase (18).Recent evidence has also raised the possibility of a nuclear site of action for both prothymosin and parathymosin. On the basis of the presence of a "nuclear localization signal" near the COOH terminus ofprothymosin and superficial structural similarities between prothymosin and a variety of nuclear proteins, proposed that it might function as a nuclear protein.…”

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confidence: 98%

Evidence for nuclear targeting of prothymosin and parathymosin synthesized in situ.

Clinton

Graeve

El-Dorry

et al. 1991

Proc. Natl. Acad. Sci. U.S.A.

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To test the hypothesis that prothymosin and parathymosin contain amino acid sequences that cause them to be targeted to the cell nucleus, expression vectors were constructed containing a simian virus 40 promoter and cDNAs that would code for chimeric proteins composed of truncated human growth hormone (hGH) linked to the NH2 terminus of prothymosin or parathymosin. The truncated hGH lacked the signal peptide sequence required for its secretion. After transfection of these constructs into HeLa S3 cells, which do not normally synthesize hGH, the use of indirect immunofluorescence staining to follow the localization of the hGH chimeras demonstrated that both prothymosin and parathymosin caused targeting to the cell nucleus. Controls with a construct coding for native hGH only, and one coding for the truncated hGH lacking the signal peptide, revealed secretion into culture medium and staining in the endoplasmic reticulum and Golgi apparatus in the first case, and diffuse staining throughout the cytoplasm in the second. The results provide direct evidence, with proteins synthesized in situ, for the presence of nuclear localization signals in both prothymosin and parathymosin.Interest in proteins of thymic origin was stimulated by the observation that a crude preparation from calf thymus, designated thymosin fraction 5, showed immunomodulatory properties in a number of in vitro test systems (reviewed in ref. 1). Many of these properties were attributed to a single peptide purified from this fraction, named thymosin a, (2), which was proposed to function as a "thymic hormone" influencing lymphocyte maturation (3). Based on evidence suggesting that this thymosin a, might be a proteolytic artifact that arose during the preparation ofthymosin fraction 5 (4, 5), a search was initiated for the native protein or polypeptide precursor. A procedure designed to minimize endogenous proteolysis led to the isolation of two homologous polypeptides, named prothymosin (6) and parathymosin (7), the first of which contained the 28-amino acid sequence of thymosin a, at its NH2 terminus. The proposed role of thymosin a, as a thymic hormone was brought into question when the putative precursor, prothymosin, and the mRNA coding for this polypeptide were shown to be present in all mammalian tissues examined (8,9). In addition, the cDNAs coding for both human (10, 11) and rat (12) prothymosins were found to lack sequences coding for the signal peptides expected in secretory proteins. An extracellular function was also rendered unlikely by the observation that the mRNA for prothymosin was localized exclusively on free polysomes (13).The experimental findings relating to parathymosin have closely mimicked those for prothymosin, with quantitative differences in tissue polypeptide and mRNA content (7,9,12,14).More recent studies have suggested a relationship between prothymosin and cell growth. Increased prothymosin mRNA in response to a variety of stimulators of mammalian cell division has been reported (11), and elevated mRNA was also ...

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“…Early studies have identified ParaT as a zinc-binding protein, which interacts with several enzymes involved in carbohydrate metabolism (30,31). Recently, ParaT was found to inhibit the binding of the activated glucocorticoid receptor to nuclei, suggesting its involvement in the regulation of glucocorticoid steroids action (32).…”

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confidence: 99%

Parathymosin Affects the Binding of Linker Histone H1 to Nucleosomes and Remodels Chromatin Structure

Martic

Karetsou

Kefala

et al. 2005

Journal of Biological Chemistry

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Linker histone H1 is the major factor that stabilizes higher order chromatin structure and modulates the action of chromatin-remodeling enzymes. We have previously shown that parathymosin, an acidic, nuclear protein binds to histone H1 in vitro and in vivo. Confocal laser scanning microscopy reveals a nuclear punctuate staining of the endogenous protein in interphase cells, which is excluded from dense heterochromatic regions. Using an in vitro chromatin reconstitution system under physiological conditions, we show here that parathymosin (ParaT) inhibits the binding of H1 to chromatin in a dose-dependent manner. Consistent with these findings, H1-containing chromatin assembled in the presence of ParaT has reduced nucleosome spacing. These observations suggest that interaction of the two proteins might result in a conformational change of H1. Fluorescence spectroscopy and circular dichroism-based measurements on mixtures of H1 and ParaT confirm this hypothesis. Human sperm nuclei challenged with ParaT become highly decondensed, whereas overexpression of green fluorescent protein-or FLAG-tagged protein in HeLa cells induces global chromatin decondensation and increases the accessibility of chromatin to micrococcal nuclease digestion. Our data suggest a role of parathymosin in the remodeling of higher order chromatin structure through modulation of H1 interaction with nucleosomes and point to its involvement in chromatindependent functions.Eukaryotic DNA is packaged into chromatin (1), a structural framework that plays a key role in the regulation of the transcriptional status of genes and genetic loci (2-4). The basic level of chromatin organization is the nucleosome that consists of DNA wrapped around an octamer of histones (5). Linker histones (e.g. H1 and H5) seal the entry/exit of DNA and stabilize the folding of 10-nm nucleosomal arrays into higher order chromatin structures (6, 7). Binding of histone H1 to nucleosomal arrays affects the spacing of nucleosomes (8), restricts their mobility (9), and reduces the transient exposure of DNA on the nucleosome surface (10). Consistent with its structural role as a major determinant of chromatin folding, histone H1 was found to inhibit the action of enzymes that enhance the accessibility of DNA, such as histone acetyltransferases and ATP-dependent remodeling complexes. For example, changes in linker histone stoichiometry modulate the levels of core histone acetylation (11) and regulate the activity of all classes of ATP-dependent remodeling enzymes (12, 13). Collectively, these findings support the notion that H1 "locks down" chromatin regions and acts as a general inhibitor of transcription (14, 15). However, genetic studies have shown that, rather than being global repressors of transcription, linker histones affect the expression of specific genes (16). Multiple isoforms of linker histone H1 exist in mammalian somatic cells (17) and play differential roles in gene expression (18). Recently, two independent studies highlight the important role of H1 subtypes in dev...

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The primary sequence of the PFK‐1 inactivating zinc‐binding protein as deduced from cDNA sequencing Identity of the zinc‐binding protein with rat parathymosin

Abstract: We have recently described the sequence of the Zn2+-binding domain (43 amino acid residues) of a newly detected Zn 2+-binding protein which reversibly inactivates phosphofructokinase-I in a Zn2+-dependent manner [(1986

Cited by 28 publications

References 14 publications

A Novel Pro-Arg Motif Recognized by WW Domains

A Novel Pro-Arg Motif Recognized by WW Domains

Evidence for nuclear targeting of prothymosin and parathymosin synthesized in situ.

Parathymosin Affects the Binding of Linker Histone H1 to Nucleosomes and Remodels Chromatin Structure

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