A 180-kDa Protein Kinase Seems to Be Responsible for the Phosphorylation of Prothymosin α Observed in Proliferating Cells*

Prothymosin α (ProTα) is an acidic protein involved in cell proliferation. Its phosphorylation status is correlated with proliferative activity. Here we report the isolation and characterization of a ProTα-phosphorylating kinase (ProTαK) from mouse splenocytes that seems to be responsible for the in vivo phosphorylation of ProTα and that differs from other protein kinases reported to date. This enzyme, mainly located in the cytosol, has an molecular mass of 180 kDa and appears to be made up of two proteins of 64 and 60 kDa. Its activity was markedly enhanced by mitogenic activation of cells. The ProTα residues phosphorylated by the enzyme in vitro are a Thr at position 7 and another Thr at positions 12 or 13, both located within casein kinase 2 (CK-2) consensus motifs; these are the same residues as are phosphorylatedin vivo. The new enzyme shows a number of clear structural and catalytic differences from CK-2. It phosphorylates histones H2B and H3, although with weaker activity than ProTα. An enzyme with the same characteristics was also found in other murine tissues and cell lines.

[1]  G. Covelo,et al.  Prothymosin alpha binds histones in vitro and shows activity in nucleosome assembly assay. , 1996, Biochimica et biophysica acta.

[2]  C. Dang,et al.  Do products of the myc proto-oncogene play a role in transcriptional regulation of the prothymosin alpha gene? , 1995, Molecular and cellular biology.

[3]  G. Damaschun,et al.  Prothymosin alpha: a biologically active protein with random coil conformation. , 1995, Biochemistry.

[4]  M. Smith.  Prothymosin alpha: in search of a function. , 1995, Leukemia & lymphoma.

[5]  S Gaubatz,et al.  An E-box element localized in the first intron mediates regulation of the prothymosin alpha gene by c-myc , 1994, Molecular and cellular biology.

[6]  K. Ahmed,et al.  Association of casein kinase 2 with nuclear matrix. Possible role in nuclear matrix protein phosphorylation. , 1994, The Journal of biological chemistry.

[7]  L. Pannell,et al.  Phosphorylation of human and bovine prothymosin alpha in vivo. , 1993, Biochemistry.

[8]  M. Freire,et al.  Prothymosin alpha is phosphorylated in proliferating stimulated cells. , 1993, The Journal of biological chemistry.

[9]  M. Freire,et al.  Prothymosin α is phosphorylated by casein kinase‐2 , 1992 .

[10]  A. Philpott,et al.  Nucleoplasmin remodels sperm chromatin in Xenopus egg extracts , 1992, Cell.

[11]  M. Freire,et al.  Thymosin α1 is a native peptide in several tissues , 1992 .

[12]  D. Beach,et al.  p34cdc2-mediated phosphorylation at T124 inhibits nuclear import of SV- 40 T antigen proteins , 1991, The Journal of cell biology.

[13]  R. Petryshyn,et al.  Phosphorylation and proteolytic degradation of nucleolin from 3T3-F442A cells. , 1991, Biochemical and biophysical research communications.

[14]  E. Rodriguez-Boulan,et al.  Evidence for nuclear targeting of prothymosin and parathymosin synthesized in situ. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[15]  R. Peters,et al.  The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T‐antigen. , 1991, The EMBO journal.

[16]  S. L. Berger,et al.  Nuclear targeting of prothymosin alpha. , 1991, The Journal of biological chemistry.

[17]  X. Bustelo,et al.  Expression of the rat prothymosin alpha gene during T-lymphocyte proliferation and liver regeneration. , 1991, The Journal of biological chemistry.

[18]  J. Bishop,et al.  The MYC protein activates transcription of the alpha‐prothymosin gene. , 1991, The EMBO journal.

[19]  E. Chambaz,et al.  Cytoplasmic and nuclear distribution of casein kinase II: characterization of the enzyme uptake by bovine adrenocortical nuclear preparation. , 1990, Biochemistry.

[20]  L. Pinna.  Casein kinase 2: an 'eminence grise' in cellular regulation? , 1990, Biochimica et biophysica acta.

[21]  M. Erdos,et al.  Cellular levels of thymosin immunoreactive peptides are linked to proliferative events: evidence for a nuclear site of action. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[22]  C. Hill,et al.  Phosphorylation at clustered ‐Ser‐Pro‐X‐Lys/Arg‐ motifs in sperm‐specific histones H1 and H2B. , 1990, The EMBO journal.

[23]  G. Maelandsmo,et al.  Phosphorylation of the high-mobility-group-like protein P1 by casein kinase-2. , 1989, European journal of biochemistry.

[24]  X. Bustelo,et al.  The expression of prothymosin alpha gene in T lymphocytes and leukemic lymphoid cells is tied to lymphocyte proliferation. , 1989, The Journal of biological chemistry.

[25]  S. L. Berger,et al.  Isolation and partial sequencing of the human prothymosin alpha gene family. Evidence against export of the gene products. , 1989, The Journal of biological chemistry.

[26]  M. Rosemeyer,et al.  Evidence for the monomeric nature of thymosins , 1989, FEBS letters.

[27]  M. Clinton,et al.  Prothymosin α and parathymosin: mRNA and polypeptide levels in rodent tissues , 1989 .

[28]  M. Frangou-Lazaridis,et al.  Isolation and partial characterization of prothymosin α from porcine tissues , 1988 .

[29]  F. Segade,et al.  Prothymosin α is a nuclear protein , 1988 .

[30]  W. Earnshaw,et al.  Anionic regions in nuclear proteins , 1987, The Journal of cell biology.

[31]  S. L. Berger,et al.  The human prothymosin alpha gene is polymorphic and induced upon growth stimulation: evidence using a cloned cDNA. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[32]  G. Goodall,et al.  Molecular cloning of cDNA for human prothymosin alpha. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[33]  G. M. Walton,et al.  Phosphorylation of high mobility group protein 14 by casein kinase II. , 1985, The Journal of biological chemistry.

[34]  A. Inoue,et al.  Heterogeneity of protein kinase NII from rat liver nuclei. , 1984, Biochemical and biophysical research communications.

[35]  G. Goodall,et al.  Prothymosin alpha: isolation and properties of the major immunoreactive form of thymosin alpha 1 in rat thymus. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[36]  E. D. De Robertis,et al.  An acidic protein which assembles nucleosomes in vitro is the most abundant protein in Xenopus oocyte nuclei. , 1980, Journal of molecular biology.

[37]  M. M. Bradford.  A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. , 1976, Analytical biochemistry.

[38]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[39]  M. Mori,et al.  Prothymosin-α mRNA expression correlates with that of c-myc in human colon cancer , 1993 .

[40]  R. Pearson,et al.  Protein kinase phosphorylation site sequences and consensus specificity motifs: tabulations. , 1991, Methods in enzymology.

[41]  S. L. Berger,et al.  Prothymosin alpha antisense oligomers inhibit myeloma cell division. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[42]  B. Horecker,et al.  Primary structure of rat thymus prothymosin alpha. , 1985, Proceedings of the National Academy of Sciences of the United States of America.