ephA9, a novel avian receptor tyrosine kinase gene

Protein tyrosine kinase (TK) receptors are known to play crucial roles in various aspects of normal development and in tumorigenesis. Germ cells before their colonizing to the gonads during embryogenesis are called primordial germ cells (PGCs). To identify TK genes expressed in chicken PGCs, we purified these cells from the blood of 2.5-day-old embryos, extracted the polyA(+) RNA, and subjected it to reverse transcription-polymerase chain reaction (RT-PCR) amplification with TK gene-specific primers. As a result, we identified 13 receptor TK genes and 6 non-receptor TK genes. One of the receptor TK genes appeared to be novel, and thus the full-length DNA complementary to RNA (cDNA) sequence was retrieved by the rapid cloning of cDNA ends method. Sequence analysis of this cDNA indicated that it encoded a novel TK receptor of the Eph family. The putative amino-acid sequence of this TK was 63.0% identical to that of human EphA1; therefore, we designated our novel TK as EphA9. Northern blot hybridizations indicated that ephA9 RNA transcripts were present in the kidney, lung, testis, and thymus but not in the spleen, brain, or liver. Expression of a fusion protein containing the intracellular domain of EphA9 in bacterial cells showed that this domain had TK enzymatic activity. The EphA9 species produced in Cos-1 cells transfected with an expression vector were tyrosine-phosphorylated. These data suggest that EphA9 plays its biological role(s) in various organs of chickens as an active TK.