The hsr-omega locus forms one of the largest Drosophila heat-shock puffs and produces three major transcripts. These three transcripts are also produced constitutively, at lower levels, in almost all tissues and developmental stages. The amounts of the transcripts in nonstressed cells are modulated during development. The hormone ecdysone leads to increased levels of hsr-omega transcripts in cultured cells, suggesting that changing ecdysone titers may play a role in the developmental changes of hsr-omega transcript levels. By in situ hybridization to RNA in tissue sections, we detect only two cell types that lack hsr-omega transcripts--the preblastoderm embryo and the primary spermatocyte. There are no maternal transcripts of hsr-omega in the embryo. Transcripts appear abruptly at the time that the zygotic genome becomes transcriptionally active, shortly before the formation of the cellular blastoderm. No constitutive hsr-omega transcripts are found in primary spermatocytes. The spermatocytes cannot respond to heat shock by transcribing either hsr-omega or hsp70 RNA. Constitutive hsr-omega transcription is resumed later in spermatogenesis and hsr-omega RNA is detected in differentiating spermatids. These spermatids are also capable of mounting a heat-shock response, as measured by increases in hsr-omega and hsp70 RNA.