Ecdysteroid t i t e r s measured i n t h e haemolymph o f t h i r d i n s t a r l a r v a e o f Drosophila revealed t h e existence o f a small peak (40 ng/ml) preceding t h e major peak a t p u p a r i a t i o n . Under experimental c o n d i t i o n s o f delayed p u p a r i a t i o n , t h e release o f b o t h e c d y s t e r o i d peaks i s retarded. t h e programming o f t h e t i m e o f p u p a r i a t i o n i s r a i s e d .
The p o s s i b i l i t y t h a t t h e e a r l i e r pulse i s i m p l i c a t e d i n I t i s known t h a t p u p a r i a t i o n i n t h e D i p t e r a i s induced by ecdysteroids (Fraenkel, '34; Butenandt and Karlson, '54). Several authors have measured e c d y s t e r o i d l e v e l s i n Drosophila melanogaster (Borst e t a l . , '74; De Reggie t a l . , '75; Hodgetts e t a l . , ' 7 7 ) . We are i n v e s t i g a t i n g t h e programming o f p u p a r i a t i o n i n t h i s animal and our s t u d i e s have l e d us t o examine w i t h g r e a t e r p r e c i s i o n m o l t i n g hormone t i t e r s throughout t h e t h i r d l a r v a l i n s t a r and e a r l y pupal period. whole animals.(Porcheron e t a l . , '76) we have been a b l e t o measure t h e e c d y s t e r o i d l e v e l d i r e c t l y i n small volumes of haemolymph. E a r l i e r measures were performed on homogenates o f
A library of Calliphora vicina genomic DNA was constructed in the λEMBL3 vector and screened for recombinant phages containing chromosomal segments encoding calliphorin, the major larval serum protein (LSP) of Calliphora. A large series of recombinants hybridizing with in vitro labelled poly(A)+ RNA from Calliphora larval fat bodies and with specific probes derived from the LSP‐1 genes of Drosophila melanogaster was isolated. Five of these phages, chosen at random, were shown by hybrid selection to retain calliphorin mRNA specifically. Eleven calliphorin mRNA‐homologous regions were located on restriction maps of these phages by hybridization with 5′ end‐labelled poly(A)+ RNA from Calliphora larval fat bodies. Each phage contains at least two calliphorin genes arranged in direct repeat orientation and seperated by 3.5–5 kb intergenic regions. The genes display similar but not identical restriction patterns. Filter hybridization and heteroduplex analysis indicate that they share a detectable homology with the LSP‐1β gene of D. melanogaster. Whole genome Southern analysis showed that these genes belong to a large family of closely related calliphorin genes which were found by in situ hybridization to polytene chromosomes of trichogen cells to be clustered in region 4a of chromosome 2 of Calliphora vicina.
Glycoprotein hormones LH, FSH, TSH and chorionic gonadotrophin are heterodimers composed of two non-covalently associated subunits, a common alpha- and a specific beta-subunit. A recombinant baculovirus containing a cDNA encoding the alpha-subunit of rat glycoprotein hormones was constructed. Viral-infected cells expressed, 48 h post infection, 7-10 mg immunoreactive alpha-glycopolypeptide/6 x 10(8) cells, of which 65-6% was able to associate with native LH beta and formed a biologically active heterodimeric hormone that bound to testicular receptors. The treatment with specific glycanases showed that the recombinant alpha-subunit was produced as two differently glycosylated forms; an M(r) 23 000 form which contained exclusively N-linked carbohydrate units and another of M(r) 25 000 which appeared to contain additional 0-linked carbohydrate. Data demonstrated that the alpha-subunit was expressed by insect cells in a manner similar to that by mammalian pituitary gonadotropes producing both the N- and O-glycosylated forms although only the N-glycosylated alpha-subunit is known to be capable of associating with the beta-subunit.
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