In one of the simplest metazoan organisms, the sponge Spongilla lacustris, at least four different src-related kinase genes (srk1-4) are expressed, all of which show a high degree of similarity to the c-src genes of vertebrates. Whereas srk2 and srk3 are clearly unrelated at the nucleic acid level, srk1 and srk4 share identical sequences in the 5' parts of their cDNAs. The cloning of several primer extension clones and genomic polymerase chain reaction experiments confirmed the hypothesis of an alternative splicing of tandemly arranged carboxy-terminal parts of srk1 and srk4. The genomic sequence encoding both proteins was found to be interrupted at the splice point by an intron which is located in the same position as one of the introns in the chicken src gene, which is the only gene conserved in invertebrates and vertebrates. All four srk genes are expressed in adult sponges as mRNA transcripts of about 2.2 kb. Tyrosine kinase activity of a src-related kinase could be detected in adult sponges but not in their resting form (gemmulae), and may reflect the activity of the srk protein products. Spongilla lacustris is the simplest organism from which a protein tyrosine kinase gene has been isolated. The presence of at least four such genes in the evolutionary ancient and primitive phylum Porifera suggests that tyrosine kinase genes arose concomitantly with or shortly after the appearance of multicellular organisms and that their activity may be involved in aggregation and cell-cell recognition.