Identification of nuclear-import and cell-cycle regulatory proteins that bind to prothymosin alpha.
Prothymosin alpha (ProT alpha) is a nuclear protein that is widely distributed in mammalian tissues, and is thought to play a role in cell proliferation. In an attempt to shed light on this role, affinity chromatography on ProT alpha-Sepharose columns was used to identify proteins in subcellular extracts of transformed human lymphocytes (NC37 cells) that interact with ProT alpha in vitro, and thus may interact with ProT alpha in vivo. Immunoblotting techniques were used to screen the ProT alpha-binding fractions for histones and other proteins involved in nuclear transport and cell-cycle control. The most abundant ProT alpha-binding proteins were histones H2A, H2B, H3, and H4. Of the nuclear-transport proteins, karyopherin beta1, Rch-1, Ran, and RCC1 were detected at high concentrations; NTF2, nucleoporin p62, and Hsp70 were detected at low concentrations; while tranportin, CAS, and Ran BPI were not detected. Of the cell-cycle control proteins, PCNA, Cdk2, and cyclin A were detected at high concentrations; cdc2, Cdk4, and cyclin B were detected at very low concentrations; while cyclin D1, cyclin D3, Cip1, and Kip1 were not detected. These results suggest (i) that ProT alpha is transported into the nucleus by the karyopherin beta1-Rch-1 complex, and (ii) that ProT alpha may interact in the nucleus with proteins involved in DNA metabolism and cell-cycle control.
Freire
J
J
Departamento de Bioquímica y Biología Molecular, Facultad de Biología, Universidad de Santiago, Santiago de Compostela, Spain.
Covelo
G
G
Sarandeses
C
C
Díaz-Jullien
C
C
Freire
M
M
D021581:Active Transport, Cell Nucleus / Q000502:physiology; D042846:CDC2-CDC28 Kinases; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004261:DNA Replication / Q000502:physiology; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry / Q000378:metabolism*; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism; D013947:Thymosin / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism*; D028901:alpha Karyopherins; D028961:beta Karyopherins
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D009687:Nuclear Proteins; D018809:Proliferating Cell Nuclear Antigen; D011498:Protein Precursors; D011506:Proteins; D028901:alpha Karyopherins; D028961:beta Karyopherins; C087978:karyopherin alpha 2; C040782:prothymosin alpha; D013947:Thymosin; D004247:DNA; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases