Changes in transcript abundance for cuticular proteins and other genes three hours after a blood meal in Anopheles gambiae

Abstract

Numerous studies have examined changes in transcript levels after Anopheles gambiae takes a blood meal. Marinotti et al. (2006) used microarrays and reported massive changes in transcript levels 3 h after feeding (BF3h) compared to non-blood fed (NBF). We were intrigued by the number of transcripts for structural cuticular proteins (CPs) that showed such major differences in levels and employed paired-end (50 bp) RNA-seq technology to compare whole body transcriptomes from 5-day-old females NBF and BF3h. We detected transcripts for the majority of CPs (164/243) but levels of only 12 were significantly altered by the blood meal. While relative transcript levels of NBF females were somewhat similar to the microarray data, there were major differences in BF3h animals, resulting in levels of many transcripts, both for CPs and other genes changing in the opposite direction. We compared our data also to other studies done with both microarrays and RNA-seq. Findings were consistent that a small number of CP genes have transcripts that persist even in 5-day-old adults. Some of these transcripts showed diurnal rhythms (Rund et al., 2013; Rinker et al., 2013). In situ hybridization revealed that transcripts for several of these CP genes were found exclusively or predominantly in the eye. Transcripts other than for CPs that changed in response to blood-feeding were predominantly expressed in midgut and Malpighian tubules. Even in these tissues, genes responsible for proteins with similar functions, such as immunity or digestion, responded differently, with transcript levels for some rising and others falling. These data demonstrate that genes coding for some CPs are dynamic in expression even in adults and that the response to a blood meal is rapid and precisely orchestrated.

Distribution of cuticular proteins in different structures of adult Anopheles gambiae

“…Vannini et al (2014) listed 10 CPs where in situ hybridization had revealed transcripts in the eye. Probes for two, CPR10 and CPR132, had been found exclusively in the eye.…”

“…From this we learned that a single structure such as the eyes, legs, or Johnston's organ could be the site of hybridization of numerous mRNAs for cuticular proteins. We have already reported on ten putative CP genes that had transcripts in the developing eye, most associated with cells that form the corneal lens and pseudocone (Vannini et al, 2014). We wondered if these transcripts were actually translated and exported into the cuticle.…”

The Evolution and Metamorphosis of Arthropod Proteomics and Genomics

“…gambiae following a blood meal (30). Vannini repeated a similar analysis using Illumina (RNA-Seq), and although our results on non-blood-fed females were similar to the James lab study, we had totally different results with females sampled 3 h after a blood meal (44). Our work was facilitated by collaborating with a former technician of mine, Augustine Dunn, who was then finishing off a bioinformatics degree in James's lab.…”

Immunolocalization of cuticular proteins in Johnston's organ and the corneal lens of Anopheles gambiae

“…They code for two of the proteins detected in this study and were found in cells beneath the corneal lens (Vannini et al, 2014a). …”

“…What was surprising was the large number of proteins we found in the corneal lens, first with in situ hybridization (Vannini et al, 2014a), next with MS/MS (Zhou et al, 2016) and now with EM immunolocalization.…”