Ultrastructural analysis of Balbiani ring genes of Chironomus pallidivittatus in different states of Balbiani ring activity

Patterns of gene activity on Balbiani ring (BR) transcription units (TUs) of polytene chromosome IV of Chironomus pallidivittatus larvae were ultrastructurally characterized by electron microscopy. In Miller chromatin spreads, the 9 micrometer long TUs of Balbiani ring 1 and 2 (BR1 and BR2), respectively, were unambiguously identified on the basis of the size and structure of their stalked ribonucleoprotein (RNP) granules. Because of the mild spreading procedure the morphology of these 50 nm BR RNP particles remained quite reminiscent of their in situ configuration. BR TUs were found as clusters of sister copies, looking almost homogeneous in structure and organization, a number of them appearing as loops. TUs of normal and hyperactive BRs (the latter resulting from stimulation of the larvae with pilocarpine) were compared. The patterns of transcriptional activity of BR TUs were rather similar but not absolutely identical on all homologous sister chromatin strands. In normal as well as in hyperactive BRs, the great majority of TUs were highly transcriptionally active, i.e. densely packed with RNP fibres all along the deoxyribonucleoprotein (DNP) axis. Mean RNP fibre frequency of BR TUs of normal larvae was 30/micrometer while hyperactive BRs showed a fibre frequency of 31/micrometer. The average number of RNP fibres per BR TU is in the range of 200 to 300. Assuming a transcription time of 20 min per BR mRNA molecule, the number of RNA polymerase molecules starting transcription of the 37 kb gene could be estimated to be 10 to 15 per minute. A number of BR genes were only partially active, exhibiting short, middle or long arrays of RNP fibres. Such arrays occurred in any segment of the genes showing normal RNP fibre density and a nucleosomal configuration in the inactive part of the gene. The variations in RNP fibre frequencies suggest that transcription can be modulated at the level of the individual BR gene. However, the large increase in the rate of BR mRNA synthesis upon pilocarpine stimulation may mainly be due to activation of hitherto inactive sister copies of the BR gene.