Prothymosin alpha is not a nuclear polypeptide

Tsitsiloni, O.E.; Yialouris, P.P.; Sekeri‐Pataryas, Kalliope E.; Haritos, A.A.

doi:10.1007/bf01957467

Cited by 28 publications

(18 citation statements)

References 18 publications

(1 reference statement)

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“…From the literature (16,17,20) it seems obvious that prothymosin, at least, is also found in the cytoplasm. This could be the result of specific removal of prothymosin from the nucleus by linkage to a small RNA, as described by Makarova et al (17).…”

Section: Discussionmentioning

confidence: 99%

“…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, Gomez-Marquez and Segade (19) proposed that it might function as a nuclear protein. Although a conventional cell-fractionation procedure yielded detectable levels of prothymosin only in the cytoplasmic fraction of calf thymus or liver (20), an elegant series ofexperiments by Watts et al (21,22) demonstrated that both prothymosin and parathymosin injected into Xenopus oocytes accumulated in the nucleus, in contrast to thymosin f84, which remained in the cytoplasm.…”

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confidence: 99%

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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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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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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“…In contrast, polyvalent antibodies to thymosin ␣ 1 highlighted immunoreactive material in the nuclei of bile duct cells but not hepatocytes (Fraga et al 1993), and distinguished "thymosin immunoreactive peptides" primarily in the nuclei of IEC-6 cells, a line derived from the small intestine of the rat (Conteas et al 1990). In disrupted cells, prothymosin ␣ was almost invariably cytoplasmic (Tsitsiloni et al 1989;Sburlati et al 1990). Therefore, these studies, too, failed to dispel the uncertainty surrounding the in vivo location of prothymosin ␣ .…”

Section: S U M M a R Ymentioning

confidence: 99%

Mobility Within the Nucleus and Neighboring Cytosol Is a Key Feature of Prothymosin-α

Enkemann

Ward

Berger

2000

J Histochem Cytochem.

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S U M M A R YProthymosin ␣ is a small, unfolded, negatively charged, poorly antigenic mammalian protein with a potent nuclear localization signal. Although it is apparently essential for growth, its precise function is unknown. We examined the location and behavior of the protein bearing different epitope tags using in situ immunolocalization in COS-1 and NIH3T3 cells. Tagged prothymosin ␣ appeared to be punctate and widely dispersed throughout the nucleus, with the exception of the nucleolus. A tiny cytoplasmic component, which persisted in the presence of cycloheximide and actinomycin D during interphase, became pronounced immediately before, during, and after mitosis. When nuclear uptake was abrogated, small tagged prothymosin ␣ molecules, but not prothymosin ␣ fused to ␤ -galactosidase, accumulated significantly in the cytoplasm. Tagged prothymosin ␣ shared domains with mobile proteins such as Ran, transportin, and karyopherin ␤ , which also traverse the nuclear membrane, and co-localized with active RNA polymerase II. Mild digitonin treatment resulted in nuclei devoid of prothymosin ␣ . The data do not support tight binding to any nuclear component. Therefore, we propose that prothymosin ␣ is a highly diffusible bolus of salt and infer that it facilitates movement of charged molecules in highly charged environments within and near the nucleus. T he function of prothymosin ␣ remains unclear despite many attempts to determine both its binding partners and its cellular location. Although the levels of the protein and its mRNA unquestionably increase in proliferating mammalian cells (Eschenfeldt and Berger 1986;Gómez-Márquez et al. 1989;Bustelo et al. 1991;Tsitsiloni et al. 1993), precisely what prothymosin ␣ does, where it does it, and what other macromolecules participate are questions with multiple unsatisfactory answers. The difficulties stem from three attributes: (a) Prothymosin ␣ is extremely acidic, with a net negative charge of ‫ف‬ 40 units (Eschenfeldt and Berger 1986;Goodall et al. 1986;Frangou-Lazaridis et al. 1988;Panneerselvam et al. 1988;Schmidt and Werner 1991). The large number of contiguous acidic amino acids in a protein of only ‫ف‬ 12 kD (Haritos et al. 1985), an unfolded conformation (Gast et al. 1995), and an abundance approaching that of histone cores (Palvimo and Linnala-Kankkunen 1990;Sburlati et al. 1990Sburlati et al. ,1993 make prothymosin ␣ an easy target for basic proteins in binding studies performed in vitro. The relevance of binding data is, accordingly, problematic. (b) Antibodies raised to prothymosin ␣ or peptides derived from it are invariably low in titer and broad in specificity. Such antibodies, particularly those directed against the N-terminus, crossreact with materials from crabs, insects, protozoans, fungi, and bacteria (Oates and Erdos 1989). However, the fully sequenced yeast genome does not code for prothymosin ␣ or related proteins, and homologous proteins in bacteria and homologous genes in a broad swath of the evolutionary spectrum up to and including reptiles h...

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“…Nevertheless, several investigations have questioned such immunological role based on its wide tissue distribution [2], its broad phylogenetic presence from yeast to man [3] and the absence of a signal peptide [4]. Moreover, since we postulated a nuclear location for ProTa [5], several contradictory data on its subcellular distribution have been reported, indicating either nuclear [6,7] or cytoplasmic localization [3,8]. However, microinjection of ProTa into the cytoplasm of Xenopus oocytes showed that this protein indeed migrates to the nucleus [6].…”

Section: Introductionmentioning

confidence: 99%

Tissue‐specific and differential expression of prothymosin α gene during rat development

1990

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We have analyzed the RNA expression of prothymosin a (ProTa) gene during rat development in several tissues and compared it to that of two proteins related to cell proliferation: proliferating cell nuclear antigen (PCNA)/cyclin and histone H3 (H3). The expression of ProTa gene was found to be regulated in a developmental and tissue-specific manner. The mRNA levels of ProTa followed a similar time-course in liver, brain, kidney, and testis, being highly increased in the early periods of postnatal development. However, in thymus ProTa mRNA showed only moderate changes throughout development. Our findings suggest that ProTa participates in developmental processes like cell proliferation and/or differentiation.

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Prothymosin alpha is not a nuclear polypeptide

Cited by 28 publications

References 18 publications

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

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

Mobility Within the Nucleus and Neighboring Cytosol Is a Key Feature of Prothymosin-α

Tissue‐specific and differential expression of prothymosin α gene during rat development

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