Abstract
Aedes aegypti PISCF-allatostatin or allatostatin-C (Ae-AS-C) was isolated using a combination of high performance liquid chromatography and enzyme-linked immunosorbent assay (ELISA). The matrix-assisted laser desorption/ionization time-of-flight (TOF) mass spectrum of positive ELISA fractions revealed a molecular mass of 1919.0 Da, in agreement with the sequence qIRYRQCYFNPISCF, with bridged cysteines. This sequence was confirmed by matrix-assisted laser desorption/ionization tandem TOF/TOF mass spectrometry analysis. The corresponding Ae-AS-C cDNA was amplified by PCR, and the sequence of the peptide was confirmed. An in vitro radiochemical assay was used to study the inhibitory effect of synthetic Ae-AS-C on juvenile hormone biosynthesis by the isolated corpora allata (CA) of adult female A. aegypti. The inhibitory action of synthetic Ae-AS-C was dose-dependent; with a maximum at 10(-9) m. Ae-AS-C showed no inhibitory activity in the presence of farnesoic acid, an immediate precursor of juvenile hormone, indicating that the Ae-AS-C target is located before the formation of farnesoic acid in the pathway. The sensitivity of the CA to inhibition by Ae-AS-C in the in vitro assay varied during the adult life; the CA was most sensitive during periods of low synthetic activity. In addition, the levels of Ae-AS-C in the brain were studied using ELISA and reached a maximum at 3 days after eclosion. These studies suggest that Ae-AS-C is an important regulator of CA activity in A. aegypti.
Highlights
Insects [1, 2]
Factors from the brain modulate corpora allata gland (CA) activity in mosquitoes; a significant reduction of Juvenile hormone (JH) synthesis is observed when CA are incubated with isolated brains, in medium in which brains have been maintained, or in medium with brain extract, suggesting that allatostatin-like factors are present in the mosquito brain [5]
We have shown that none of the five A. aegypti allatostatin-A had an effect on JH synthesis in female mosquito; on the other hand AS-C from A. gambiae showed a significant inhibitory response [5]
Summary
Insects [1, 2]. In adult female Aedes aegypti mosquitoes, JH levels are low at adult eclosion, elevated in sugar-fed females, and low again after a blood meal [3]. We describe the purification, biochemical, molecular, and functional characterization of A. aegypti PISCF-allatostatin (or Ae-AS-C). It is the first comprehensive report since the original description of a member of this peptide family as an inhibitor of JH synthesis in the lepidopteran Manduca sexta [8]. We describe the developmental stage-dependent Ae-AS-C inhibition of JH synthesis, as well as the changes in peptide levels in head samples. These studies suggest that AeAS-C is an important regulator of CA activity in A. aegypti
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