Mechanisms of transcriptional regulation inAnopheles gambiaerevealed by allele specific expression

Abstract

Malaria control relies on insecticides targeting the mosquito vector, but this is increasingly compromised by insecticide resistance, which can be achieved by elevated expression of detoxifying enzymes that metabolize the insecticide. In diploid organisms, gene expression is regulated both incis, by regulatory sequences on the same chromosome, and bytransacting factors, affecting both alleles equally. Differing levels of transcription can be caused by mutations incis-regulatory modules (CRM), but few of these have been identified in mosquitoes. We crossed bendiocarb resistant and susceptibleAnopheles gambiaestrains to identifycis-regulated genes that might be responsible for the resistant phenotype using RNAseq, andcis-regulatory module sequences controlling gene expression in insecticide resistance relevant tissues were predicted using machine learning. We found 115 genes showing allele specific expression in hybrids of insecticide susceptible and resistant strains, suggestingcisregulation is an important mechanism of gene expression regulation inAnopheles gambiae. The genes showing allele specific expression included a higher proportion ofAnophelesspecific genes on average younger than genes those with balanced allelic expression.Author SummaryThe evolution of insecticide resistance, including resistance that is due to changes in the expression levels of certain resistance associated genes is threatening progress in malaria control. We investigated how the expression of genes in the malaria vectorAnopheles gambiaeis controlled, by implementing a method for the first time in this species. Each mosquito inherits a set of chromosomes from both parents, so has a maternal and paternal copy of most genes. When a gene is expressed, the DNA encoding that gene is transcribed into messenger RNA. This process is controlled by the cellular environment and by other DNA sequences on the same chromosome as each gene. We crossed mosquitoes from insecticide resistant and susceptible strains to equalize the cellular environment and then measured the levels of messenger RNA from both gene copies. 115 genes showed consistently different messenger RNA levels between gene copies in most crosses, suggesting these genes are regulated by factors on the same chromosome. There were relatively more Anopheles specific genes with imbalanced expression. Using machine learning we identified DNA sequences that may be responsible for controlling gene expression in mosquito tissues; several of these sequences were close to genes with imbalanced expression.

Copy number variants underlie the major selective sweeps in insecticide resistance genes inAnopheles arabiensisfrom Tanzania

“…In the Cyp12f cluster, Cyp12f2 and Cyp12f3 showed allelic imbalance in gene expression in F1 crosses between resistant and susceptible colonies of An. gambiae , suggesting differential cis regulation of expression linked to resistance (Dyer et al, 2023). Furthermore, the presence of Cyp12f1 CNVs in both An.…”

Parallel Evolution in Mosquito Vectors—A Duplicated Esterase Locus is Associated With Resistance to Pirimiphos-methyl in Anopheles gambiae

“…More recently, whole-genome sequencing combined with population genetics has revealed the full extent of positive selection acting upon the wild genomes of Anopheles populations ( Miles et al 2017 ; Clarkson et al 2020 ). These data have revealed a complex genetic architecture of insecticide resistance, including the involvement of amino acid mutations, copy number variants (CNVs), and cis-regulatory variation ( Lucas et al 2019 ; Clarkson et al 2021 ; Dyer et al 2023 ; Lucas et al 2023 ; Kengne-Ouafo et al 2024 ).…”