Identification of the Aedes aegypti Peritrophic Matrix Protein AeIMUCI as a Heme-Binding Protein

Abstract

The gene Aedes aegypti intestinal mucin 1 (AeIMUC1) encodes a putative peritrophic matrix (PM) protein that is expressed in the midgut of mosquito larvae and adults and is upregulated in response to exposure to heavy metals. The AeIMUC1 protein has a predicted secretory signal peptide and three putative chitin-binding domains (CBDs) with an intervening mucin-like domain. Immunofluorescence and immunoelectron microscopy experiments established that AeIMUC1 is a bona fide PM protein, and binding of the recombinant protein to chitin was demonstrated in vitro. Previous experiments suggested that the Ae. aegypti PM can bind toxic heme molecules generated during blood digestion. However, the identity of the binding molecule(s) was unknown. Using of heme-agarose beads and spectrophotometric and microcalorimetric titrations, we show that recombinant AeIMUC1 can bind large amounts of heme in vitro, suggesting for the first time a role for a PM protein in heme detoxification during blood digestion. Binding of heme to AeIMUC1 was accompanied by an altered circular dichroism spectrum indicating a change in protein conformation, consistent with an increase in secondary structure. Heme-binding activity was mapped to the AeIMUC1 CBDs, suggesting that these domains possess dual chitin- and heme-binding activity.

Modulation of Midgut Peritrophins' Expression during <i>Plasmodium Infection</i> in <i>Anopheles stephensi</i> (Diptera:Culicidae)

“…Based on the sequence similarity with A. gambiae peritrophins, presence of secretory signal peptide, chitin-binding domain (s) and glycosylation sites, eight putative peritrophins were identified in A. stephensi. The absence of serine/proline/threonine-rich domains and the heme-binding domains further confirm that these proteins are different from the mucin-like peritrophins 3 , which are also reported under Class III peritrophic matrix proteins 4,14 . The presence of a varied number of chitin-binding domains in A. stephensi peritrophins might arose from a common ancestor protein harbouring a single chitin-binding domain (CBD) as revealed earlier 20 .…”

“…These proteins possess water retaining capability and hence protect the tissue from dehydration and provide lubrication for the food passage 3 . 13 and Aedes aegypti 14 . The PM of hematophagous insects, particularly mosquitoes, has been demonstrated to have an added role of limiting Plasmodium establishment in the mosquito midgut.…”

Induction of Nitric Oxide Synthase and Activation of Signaling Proteins inAnophelesMosquitoes by the Malaria Pigment, Hemozoin

“…Up to 18 nmol of heme (equivalent to 2 l of blood containing 10 mM heme) was found to bind to the PM 48 h after feeding in Aedes aegypti (12,37). Although it is not known whether Hz binds to PM as heme does, our findings demonstrate that PfHz could function in vivo as a proinflammatory signal to mosquito midgut cells in a way that is fundamentally different from the way that heme that is released during blood digestion functions (Fig.…”

Disruption of the peritrophic matrix by exogenous chitinase feeding reduces fecundity in Lutzomyia longipalpis females

“…Alterations in early blood digestion in the absence of a PM may have additional effects on the female reproductive cycle. The direct contact of the gut epithelium with intermediates of haemoglobin digestion could induce oxidative stress in sandfly females, which was observed previously in Lutzomyia spinicrassa (Walters et al 1995) and A. aegypti (Pascoa et al 2002, Devenport et al 2006 leading to compromised ability to produce eggs. In addition, the premature direct contact of the L. longipalpis gut microbiota with the midgut epithelium may cause an additional challenge to the BCC-fed insects because potentially pathogenic bacteria, such as Serratia marcescens, are present in the L. longipalpis midgut (Gouveia et al 2008).…”