The Drosophila genome contains a number of genes related to the major heat-shock-inducible (Hsp70) genes of Drosophila. The structure and expression of three of these heat-shock cognate (Hsc) genes have been studied; Hsc1 (previously described), Hsc2, and Hsc4 are dispersed on chromosome 3 at cytological loci 70C, 87D, and 88E, respectively. RNA homologous to Hsc4 was shown by hybridization selection translation experiments to encode a protein of approximately 70kd, with similar, but distinguishable, mobility to that of the heat-shock-inducible 70K protein. Because of variance in size of the 5' nonprotein coding region, it was possible to identify in adult flies distinct transcripts from each gene. Unlike transcripts from Hsp genes, the abundance of the Hsc transcripts was not increased after heat shock. Transcription of these Hsc genes is regulated during development. While transcripts from Hsc4 were equally abundant in RNA isolated from embryos, larvae and adults, Hsc1 and Hsc2 transcripts were not detected in embryo and larval RNA, and are therefore at least 20 times less abundant in these stages than in adults. The primary DNA sequences of the regions homologous to that encoding the amino terminus of hsp70 were compared. Approximately 76% homology of the base sequence and 78% homology of the deduced amino acid sequence were found among the four genes. The Hsp70 genes contain no intervening sequences; Hsc4 contains no insertions relative to Hsp70 in the region encoding the first 101 aa. However, in the codon specifying aa66 Hsc1 is interrupted by 1.7 kb of DNA; in the codon for aa55 Hsc2 contains an insertion of about 650 b. The sequences at the 5' and 3' junctions of the insertions in both Hsc1 and Hsc2 are similar to those found in intervening sequences of Drosophila and of many other organisms.