Sequences of two cDNAs and expression of the genes encoding methionine-rich storage proteins of Manduca sexta

In Manduca sexta, storage proteins accumulate during the final larval stadium for utilization during subsequent larval-pupal-adult transformations. Two cDNA clones (designated clone 119 and clone 201), that represent two distinct but related genes (42% sequence identity), were isolated from a cDNA library prepared from day 7 fifth instar larval fat body and found to encode two different storage proteins synthesized during the last larval instar. Northern blot analyses revealed that the two clones hybridize to 2.4 kb transcripts that are translated to 79 kDa protein products during in vitro translation experiments. Clone 119 encodes a methionine-rich storage protein, designated as SP1A, that shares 37% sequence identity with the Bombyx mori sex-specific storage protein SP1. Clone 201, on the other hand encodes a storage protein, designated as SP1B, that is more closely related to B. mori SP1 (63% identity), and is probably identical to the Manduca female-specific storage protein (FSP). Insert DNA from clone 201, but not clone 119, cross-hybridizes to that of FSP cDNA (Webb and Riddiford, Dev. Biol.130, 671–691, 1988a). Both storage proteins are synthesized only in the fat body and only during the fifth larval stadium, indicating tissue- and stage-specific expression of the two genes. Both genes exhibit sex-specific differences in expression. In the fifth larval stadium, the mRNAs for the SP1A and SP1B proteins begin to accumulate at about day 2 in the female fat body but appear 2 or 3 days later in the male fat body. In both sexes SP1A mRNA remains relatively high beyond the time when SP1B mRNA has already declined to low levels, suggesting differences in mRNA stability or expression. Injection of 20-hydroxyecdysone into ligated fifth instar abdomens causes substantial increases in the levels of both mRNAs, whereas topical application of the juvenile hormone mimc, fenoxycarb, to feeding fifth instar larvae produces substantial declines in the mRNA levels, indicating hormonal effects at the transcriptional level. The data support the hypothesis that the expression of these M. sexta methionine-rich storage protein genes is stimulated by ecdysteroid and inhibited by juvenile hormone.