Identification of the Balbiani ring 2 chromomere and determination of the content and compaction of its DNA.

Abstract

The giänt puff Balbiani ring 2 (BR 2) in the salivary glands of Chironomus tentants is known to contain transcriptionally active 75S RNA genes. The corresponding chromosomal superstructure, the BR 2 chromomere, has been identified and characterized in the present study. It was demonstrated by cytological methods that BR 2 originates from the broad band IV-3B10, since all the bands except the 3B10 band could be detected as intact bands in the vicinity of BR 2. In situ hybridization of BR 2 RNA to squashed Malpighian tubule chromosomes lacking BR 2, showed that all or almost all the DNA sequences complementary to BR 2 RNA are located in the 3B10 band. The DNA amount in the 3B10 band, designated the Br 2 band, was measured in relation to the DNA content of the whole chromosome set by both direct and indirect microspectrophotometry of Feulgen-stained Malpighian tubule chromosomes. From a determination of the haploid DNA content in sperm cells (0.25 pg), the amount of DNA in at BR 2 chromomere could then be calculated to be 5.1 x 10-4 pg DNA or 470 kb DNA. A minimum value of the DNA compaction within the BR 2 chromomere was estimated to 380 from the B-form length of BR 2 DNA (160 mum) and the thickness of the BR 2 band (0.42 mum). Since there are only between 1 and 4 75S RNA genes, 37 kb in size, in a BR 2 chromomere, most of the BR 2 DNA must consist of DNA not coding for 75S RNA. The nature of this DNA is discussed in relation to the high average AT-content of Chironomus DNA. The orgnization of the chromosome fiber in the BR 2 chromomere is considered in relation to the gene activation process in the salivary glands, i.e. the formation of a 75S RNA transcription loop from a protion of the tightly packed chromosome fiber in the BR 2 chromomere.