Features of in vitro puffing and RNA synthesis in polytene chromosomes of Chironomus

In order to elucidate the causal relation between RNA synthesis and puffing in polytene chromosomes, salivary gland nuclei and chromosomes from Chironomus tentans and C. pallidivittatus (Diptera, Chironomidae) were isolated in the native state under controlled and optimized conditions. The behavior of the polytene chromatin in vitro as compared to that in vivo was studied morphologically and autoradiographically. It was shown that the absence of cytologically visible puffing does not necessarily indicate transcriptional quiescence. Under suitable conditions all potential puff sites incorporate the radioactive precursor simultaneously. This can be seen in vivo and even more clearly in chromosomes decondensed by 340-400 mM NaCl in vitro. Potential puff sites therefore appear to be permanently equipped with RNA-polymerases, and there is always a basic low level of transcriptive activity. Experimental shrinking of Balbiani rings and other puffs by dimethylsulfoxide (DMSO) or heat shock in vivo, leaves the genes in the collapsed puffs still transcriptionally active, although to a reduced extent. Light optically visible puffing is therefore not a necessary prerequisite for RNA synthesis, while, conversely, RNA synthesis is required for puff formation.