Cloning and characterization of a cDNA for rat CD30 homolog and chromosomal assignment of the genomic gene.
CD30 is a member of the tumor necrosis factor receptor superfamily, which is expressed on some activated lymphocytes, virus-infected cells and transformed lymphocytes. To facilitate our understanding of biological functions and functional domains, we isolated rat cDNA clones encoding the rat homolog of human CD30 from a cDNA library of a rat T-cell line, TARL-2. The nucleotide sequence of the cDNA showed 73% homology with that of human CD30. The deduced rat CD30 protein consisted of 493 amino acids with an M(r) of 59 160 and contained a single transmembrane domain. It lacked the second repeat of the cysteine-rich motif in the extracellular domain found in human CD30. The amino acid sequence showed 51.8 and 61.2% identity with the cysteine-rich and the cytoplasmic domains, respectively. In the cytoplasmic domain, however, the amino acid sequence was highly conserved in about 100 residues near the C-terminus showing 77.7% identity, whereas the rest of the cytoplasmic domain showed 45.2% identity. This conservation suggests the functional importance of this region. Comparison with the recently reported mouse CD30 revealed 83.7% conservation of the amino acid sequence and a common structure of the extracellular domain which lacks the second cysteine-rich motif. Northern blots revealed a 3.4-kb mRNA in the PHA-activated spleen cells and human T-cell leukemia virus type 1 (HTLV-1)-infected rat T-cell lines, whereas smaller transcripts of 2.3 kb were found in the lung. A rabbit polyclonal antibody raised against GST-fusion protein of the cytoplasmic domain detected bands with an apparent M(r) of 80 kDa and 100- 110 kDa expressed in TARL-2 and spleen cells. Transient overexpression of rat CD30 in TARL-2 cells activated HIV LTR in a NF-kappa B site-dependent manner, indicating that CD30 signals activate NF-kappa B. The chromosomal location of the gene was identified by fluorescence in situ hybridisation at 5q36.2, and appeared to correspond to human 1p36, where human CD30 has been mapped. The identification and characterization of the rat counterpart of human CD30 will facilitate studies of the biological function of this molecule.
Aizawa
S
S
Department of Pathology, Institute of Medical Science, University of Tokyo, Japan.
Satoh
H
H
Horie
R
R
Ito
K
K
Choi
S H
SH
Takeuchi
H
H
Watanabe
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D003545:Cysteine / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry*; D005786:Gene Expression Regulation / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence; D017730:Ki-1 Antigen / Q000737:chemistry*; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D014162:Transfection / Q000235:genetics
D018076:DNA, Complementary; D017730:Ki-1 Antigen; D016328:NF-kappa B; D011993:Recombinant Fusion Proteins; D003545:Cysteine