Characterization of a carboxypeptidase A gene from the mosquito, Aedes aegypti.

A gut-specific carboxypeptidase A gene (AeCPA) from the mosquito, Aedes aegypti, was cloned and characterized. The gene has an open reading frame that predicts a protein of 427 amino acids, 61% of which are identical to an Anopheles gambiae carboxypeptidase A sequence. AeCPA messenger RNA (mRNA) was not detected during larval and pupal development. In situ hybridization experiments indicated that AeCPA mRNA is expressed by posterior midgut epithelial cells. In sharp contrast to An. gambiae carboxypeptidase A gene expression, AeCPA mRNA accumulates to high levels only late ( approximately 16-24 h) after ingestion of a blood meal. The temporal profile of AeCPA gene induction is similar to that of Ae. aegypti late trypsin, suggesting the existence of common regulatory elements.

Carboxypeptidase B in Anopheles gambiae (Diptera: Culicidae): effects of abdominal distention and blood ingestion.

Carboxypeptidase B (CPB) activity was detected in the guts of strain G-3 of Anopheles gambiae (Giles) and Aedes aegypti (L.). Mosquitoes were examined 3-5 d after emergence following exposure to 20% sucrose, from 0 to 4 h after feeding on a meal of latex beads in saline, and from 0 to 96 h after blood feeding. CPB activity was assayed in whole-gut homogenates, including lumenal contents and peritrophic matrix, by following the hydrolysis of a substrate specific for CPB-[3H]-benzoyl-L-Phe-L-Arg. Homogenates were divided into cytosolic plus lumenal components and membrane-associated components. Activity levels changed in response to feeding, decreasing in response to distention by saline plus latex and increasing only in response to blood. Overall, CPB activity was higher in unfed An. gambiae than in unfed Ae. aegypti. Detection of CPB activity in the peritrophic matrix of both species indicated that this enzyme was secreted actively into the gut lumen. In An. gambiae, CPB activity was optimal at pH 8, and thiol-type CPB was the predominant form detected. The data indicated that CPB in An. gambiae was regulated by both physical and chemical factors.

The molecular biology of hair cell regeneration in the avian cochlea.

The sensory cells of the ear, the hair cells, are damaged by loud noise or certain types of drugs. In the bird cochlea, new hair cells are produced to replace those that are lost. Regeneration also occurs in the vestibular epithelia of birds, fish, and mammals but does not occur in the mammalian cochlea. In order to further our understanding of the regeneration process in the bird cochlea, we have begun to identify the genes that are involved. However, the small size of this organ has made it difficult to use traditional molecular biology methods to address these problems. Recently, many molecular techniques have been adapted for use with small amounts of tissue. Northern blot analysis, the ribonuclease protection assay, semiquantitative PCR and differential display of mRNA are all techniques that are being used to greatly improve our understanding of hair cell regeneration and may eventually provide the information necessary to induce regeneration in hearing-impaired humans.

Peritrophic matrix proteins of Anopheles gambiae and Aedes aegypti.

Little is known about the composition and function of the mosquito peritrophic matrix (PM), a physical barrier that pathogens must traverse to complete their life cycles. Anopheles gambiae and Aedes aegypti PM proteins induced by blood or by a protein-free meal have been characterized by the use of 2-D gel electrophoresis and lectin-binding affinity assays. More than forty proteins have been identified in both species. Over half of the PM proteins of both mosquitoes migrate identically. Many PM proteins appear to be glycosylated, primarily by high mannose N-linked glycosyl groups.