A peptide-reactive antibody to a Balbiani ring gene product: immunological evidence that a 6.5-kb RNA in Chironomus tentans salivary glands is mRNA for a 180-kDa nonfibrous component of larval secretion

Abstract

An immunological approach was utilized to demonstrate that a tissue-specific Balbiani ring (BR) transcript in Chironomus tentons is the mRNA for a secreted 180-kDa polypeptide. Balbiani ring 1 (BR1) on the polytene chromosome IV of larval salivary glands contains a gene comprised of tandemly duplicated nucleotide sequences that are transcribed into a salivary gland-specific, 6.5-kb poly(A) + RNA for which a partial cDNA sequence exists [Dreesen et al., J. Biol. Chem. 260 (1985) 11824–11830]. A nonapeptide was synthesized so that its amino acid sequence corresponded to an open reading frame in the cDNA. This peptide was used to raise rabbit polyclonal antisera and to purify the peptide-reactive antibody by affinity chromatography. The affinity-purified antibody bound specifically to a 180-kDa polypeptide on Western blots containing extracts of total salivary gland protein. Western blot analysis of microdissected cellular vs. lumenal fractions of salivary glands indicated that this 180-kDa polypeptide was primarily localized in the lumen. Consequently, this polypeptide was designated a secretory polypeptide (sp180). Finally, the peptide-reactive antibody was used to localize sp180 in a nonfibrous component of salivary gland secretion by indirect immunofluorescence microscopy.