Cloning and characterization of a cDNA encoding a novel heterogeneous nuclear ribonucleoprotein-like protein and its expression in myeloid leukemia cells.
We isolated a cDNA encoding a novel heterogeneous nuclear ribonucleoprotein (hnRNP)-like protein on DNA affinity screening of a K562 cDNA expression library with an oligodeoxynucleotide (JKT41) derived from intron 9 of the human myeloperoxidase gene. The cDNA has a 1,305 bp sequence that encodes a polypeptide of 301 amino acid residues. The protein, named JKTBP, contains two repeats of a putative RNA binding domain (RBD), each composed of canonical RNP-2 and RNP-1 motifs, and a glycine- and tyrosine-rich carboxyl terminus. The sequences of these two repeats are highly homologous with those of the 2 x RBD-Gly rich group of hnRNPs. Northern blotting showed that two mRNAs of approximately 1.4 and 2.8 kb were present in most cultured cells examined. The recombinant protein expressed in Escherichia coli interacted with the double-stranded form of JKT41 as well as with its single-stranded form. This interaction was competitively inhibited by the same unlabeled JKT41 and to nearly the same extent by unrelated oligonucleotides. Moreover, the recombinant protein interacted with poly(G) and poly(A), but not with poly(U) or poly(C). Transient expression of the protein in SKM-1 cells repressed the expression of chloramphenicol acetyltransferase reporter genes located downstream of the intron 9 element of JKT41 or intron 7 element of FERE27. The implications of the protein in the biogenesis of mRNA are discussed.
Tsuchiya
N
N
Graduate School of Integrated Science, Yokohama City University, 22-2 Seto, Kanazawa-ku, Yokohama 236-0027, Japan.
Kamei
D
D
Takano
A
A
Matsui
T
T
Yamada
M
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary; D015972:Gene Expression Regulation, Neoplastic; D015723:Gene Library; D017930:Genes, Reporter; D006801:Humans; D007438:Introns; D007951:Leukemia, Myeloid / Q000235:genetics*; D008969:Molecular Sequence Data; D009195:Peroxidase / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; C111986:HNRPDL protein, human; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D004247:DNA; D009195:Peroxidase; D015500:Chloramphenicol O-Acetyltransferase