Identification and Characterization of Heat Shock 70 Genes inAedes aegypti(Diptera: Culicidae)

Abstract

Heat shock genes are highly evolutionarily conserved and are expressed to varying degrees in all organisms in response to stress. Heat shock 70 (hsp70) genes have been well characterized in a number of organisms, most notably Drosophila melanogaster, but not as yet for any of the major arthropod-borne viral mosquito vectors. To identify hsp70 genes in the yellow fever mosquito, Aedes aegypti (Diptera: Culicidae), basic local alignment searches of the Ae. aegypti genome were performed using D. melanogaster Hsp70 protein sequences as query. Two clusters of six previously unannotated AaHsp70 genes were identified and found to be organized into three pairs of nearly identical open reading frames, which mapped to two genomic scaffolds. Consistent with a designation as heat shock genes, no detectable level of expression of AaHsp70 genes was observed under normal rearing conditions (28°C), with robust expression observed with a heat shock of 37–39°C. Northern analysis showed heat-inducible expression of putative AaHsp70 genes at all life stages and in all tissues tested in a time- and temperature-dependent manner. Monitoring of AaHsp70 gene expression levels in field-caught Ae. aegypti may serve as a general marker for stress. In addition, promoter sequences from AaHsp70 genes may be used to control the expression of transgenes in an inducible manner.

Functional characterization of Aedes aegypti alkaline phosphatase ALP1 involved in the toxicity of Cry toxins from Bacillus thuringiensis subsp. israelensis and jegathesan

“…This sequence is quite distinct from that of the other ALPs. A highly heat-inducible heat shock 70A promoter (Gross et al 2009) was used to drive the hairpin dsRNA expression in case Aedes mosquitoes are not tolerant to the AaeALP1 gene knockdown (Fig. 4A).…”

“…To obtain the 3′-UTR sequences of AaeALP1 gene for making hairpin dsRNA, 3′-RACE was done and the PCR product was cloned into the TA vector, pCR2.1 Topo, and eventually sequenced. The hairpin dsRNA construct of AaeALP1 was made by using a heat-inducible Aedes heat shock promoter 70A (Gross et al 2009), an AaeALP invert repeat with an APN intron sequence as a stem loop (140 bp, AAEL008155), and SV40 terminator (250 bp). The synthesized dsRNA construct was then cloned into pBAC[3xP3-EGFP afm] transformation vector.…”

Identification of heat shock protein genes hsp70s and hsc70 and their associated mRNA expression under heat stress in insecticide-resistant and susceptible diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae)

“…In D. melanogaster, eight hsp70 genes are identified through the bioinformatic analysis of the whole genome, and each gene could generate the varying amounts of mRNAs upon alternative splicing (Mou et al, 2011). In the yellow fever mosquito Aedes aegypti (Diptera: Culicidae), two clusters of six hsp70 genes are found through the basic local alignment searches of the genome (Gross et al, 2009). The human Hsp70 family contains at least eight proteins with distinct amino acid sequences, expression levels and subcellular localization (Tavaria et al, 1996).…”

Mosquito/microbiota interactions: from complex relationships to biotechnological perspectives

“…Recognitions of microbial molecules triggers various immune responses: (1) killing of microbes by hemocytes (insect blood cells) that engulf (phagocytosis) or surround (encapsulation) the pathogen, (2) activation of a serine protease cascade to activate melanization (humoral melanotic encapsulation), or (3) production of antimicrobial peptides and other effector genes [40]. The ability to mount an adapted immune response relies on the existence of various insect pattern recognition receptors (PRRs) capable of recognizing different microorganisms-associated molecular patterns (MAMPs) [41,40]. Phagocytosis involves the engulfment of bacteria and other small pathogen particles by cells.…”