IMMUNOLOCALIZATION OF THE OOSTATIC AND PROTHORACICOSTATIC PEPTIDE, NEB-TMOF, IN ADULTS OF THE FLESHFLY, NEOBELLIERIA-BULLATA

The hexapeptide Neb-TMOF (H-NPTNLH-OH, trypsin modulating oostatic fac tor of the gray fleshfly, Neobellieria bullata)(2) occurs in vitelloge nic ovaries and is involved in negative feedback regulation of trypsin biosynthesis in the gut of late vitellogenic females. Polyclonal anti sera were raised against the synthetic peptide and were used to identi fy and immunolocalize Neb-TMOF epitopes in different fleshfly tissues. Neb-TMOF-immunoreactive material first appears in the cortical layer of young vitellogenic oocytes and later spreads over the yolk granules . This suggests a pinocytosis with the three yolk polypeptides (vitell ogenins). Controls treated with the preimmune sera or with anti-Neb-TM OF antiserum preadsorbed to Neb-TMOF peptide coupled to a solid phase support did not stain. There was no immunostaining in the central nerv ous system (brain and ventral nerve cord), the retrocerebral complex, the fat body, or the testes. Western blot analysis showed that the ant i-Neb-TMOF antisera specifically recognize a putative hormone precurso r polypeptide (M(r) 75 kDa) from vitellogenic ovaries. This protein is virtually absent from the hemolymph. It is not immunologically relate d to the three yolk polypeptides, since it is not recognized by yolk p olypeptides antisera. In adult females the ovary appears to be the onl y site of synthesis of Neb-TMOF and of its precursor. Immunopositive s taining is found in the apical areas of ovarian follicle cells, sugges ting these cells as a site of hormone precursor biosynthesis. This is the first demonstration that a protein colocalized with yolk proteins might act as a precursor for a folliculostatic hormone. (C) 1996 Acade mic Press, Inc.