Genome-wide analyses of ATP-Binding Cassette (ABC) transporter gene family and its expression profile related to deltamethrin tolerance in non-biting midge Propsilocerus akamusi

Highlights

• The chironomid ABC transporter gene family are firstly described in Propsilocerus akamusi on genome-scale.

• The ABCC, ABCG, and ABCH subfamilies identified some members formed tandem repeats.

• The PaABCG17 gene plays an important role with deltamethrin detoxification.

• The ABC transporter genes of other chironomids with assembled genome are first described.

Abstract

Non-biting midges are dominant species in aquatic systems and often used for studying the toxicological researches of insecticides. ATP-binding cassette (ABC) transporters represent the largest known members in detoxification genes but is little known about their function in non-biting midges. Here, we selected Propsilocerus akamusi, widespread in urban streams, to first uncover the gene structure, location, characteristics, and phylogenetics of chironomid ABC transporters at genome-scale. Fifty-seven ABC transporter genes are located on four chromosomes, including eight subfamilies (ABCA-H). The ABCC, ABCG, and ABCH subfamilies experienced the duplication events to different degrees. The study showed that expression of the PaABCG17 gene is uniquely significantly elevated, with deltamethrin concentration increasing (1, 4, and 20 ug/L) both in RNA-seq and qPCR results. Additionally, the ABC transporter members of other six chironomids with assembled genomes are first described and used to investigate the characteristic of those living in the different adverse habitats. The ABC transporter frame for Propsilocerus akamusi and its transcriptomic results lay an important foundation for providing valuable resources for understanding the ABC transporter function in insecticide toxification of this species as well as those of other non-biting midges. The PaABCG17 gene is shown to play an important role in deltamethrin detoxification, and it functions need to be further investigated and might be used in the management of insecticide-resistance in chironomid adults.

Introduction

ATP-binding cassette (ABC) transporters, which are transmembrane proteins, are widespread in all living organisms and bind ATP to transport substrates (mainly metabolites and xenobiotics) (Dassa & Bouige, 2001; Dean et al., 2001; Holland et al., 2003). The gene structure of ABC transporters is highly conserved among insects, generally composed of cytosolic nucleotide-binding domain (NBD) and less conserved transmembrane domain (TMD) (Wu et al., 2019). The complete and functional ABC transporter (full transporter) consists of two NBDs and two TMDs that exports various substrates across the membrane by conformational changes with binding and hydrolyzing ATP as a pump (Rees et al., 2009). In eukaryotes, some members of ABC transporter are half-transporters only containing one TMD and one NBD domains, which are later assembled as either homodimers or heterodimers to perform function with four domains (Hollenstein et al., 2007). Generally, the NBD includes several highly conserved elements such as Walker A and B motifs, Q-loop, H motif, and LSGGQ motif as signature sequence (Higgins and Linton, 2004) While for TMD, it is a prominent quaternary structure with 12 transmembrane helices that acts as a switch to transport substrate in all ABC exporters (Higgins, 1991) Most known present-day ABC transporters are classified into eight subfamilies, named ABCA-ABCH, based on the homology of the NBD sequences (Merzendorfer, 2014). With much genome-level resources published in insects, a large number of studies focused on the function of different superfamilies of ABC transporter in xenobiotics resistance (Kang et al., 2016; Denecke et al., 2017; He et al., 2019; Mastrantonio et al., 2019; Pan et al., 2020).

In freshwater systems, Chironomidae larvae are dominant species with tube-dwelling life. Additionally, the lifespans of those are short and their high sensitivity to xenobiotics, the toxicological impacts of chiromomids are used for monitoring water quality (Anderson, 1997). As potential bioindicators, a series of investigations of toxicological studies in chironomids have been reported, especially for insecticides or pesticides such as DDT, aldrin, tributyltin, nonylphenol, bisphenol A, and neonicotinoid insecticides (Li et al., 2009; Dermanuw & Van Leeuwen, 2014; Chen, Li & You, 2015; Martínez-Paz et al., 2012, 2013, Martínez-Paz, 2017). Among those toxicants, the pyrethroid insecticides are widely used in mosquito control and the molecular mechanisms for pyrethroid resistance of those are well-studied involving in overexpression of detoxification enzymes of mosquitoes (WHO, 2006). However, few studies associate the larvae of chironomids with pyrethroid exposure, which is highly toxic in urban water. Additionally, the massive swarms of several species of non-biting midges cause trouble to humans, especially for respiratory problems (Cranston, 1995; Hirabayashi et al., 1997). A better understanding of the molecular mechanism of pyrethroid resistance to chironomids could enable investigators to seek feasible strategies for pest management.

To date, only a handful of genes such as antioxidant enzymes, cytochrome P450s, acetylcholine receptors (AChRs), and glutathione-S-transferases (GSTs), have been demonstrated to be correlated with the tolerance/resistance to insecticides in chironomids (Martínez-Paz et al., 2012; Monterio et al., 2019; (Ballesteros et al., 2020). Recently, the ABC transporters have gained more attention as efflux-pumps for studying the impact of pyrethroid resistance/tolerance in some agricultural pests and disease vectors (Rault et al., 2019; Xu et al, 2020). The overall detoxification process is generally divided into three phases (phase I-III). Enzymes such as P450s and carboxylesterases (CCEs), activate the toxic molecules in phase I, and the reactive toxicants are often coupled of phase II conjugating enzymes such as GSTs or UGTs. Finally, in phase III the polar compounds or conjugates might be transported out of the cell by ABC transporters(Dermauw and Van Leeuwen, 2014) Sometimes ABC transporters directly transport toxicants out of the cell without enzymatic modifications to prevent toxic molecules accumulating in the cells or organism, which is referred to as phase 0 (Kennedy & Tierney, 2013). With the development of genome sequencing technology, genome-wide ABC transporters are identified in many insects, and members of those are found to play a part in the detoxification process of numerous model insects and agricultural pests, including Drosophila melanogaster, Plutella xylostella, Anopheles gambiae, and Helicoverpa armigera (Dean et al., 2001; Roth et al., 2003; Qi et al., 2016; Jin et al., 2019). However, the expression of ABC transporter genes involved in pyrethroid resistance has not been explored in chironomids.

Propsilocerus akamusi (Chironomidae: Prodiamesinae) is a ubiquitous species in many lakes of East Asia. In Asian aquatic ecosystems, this species is widely distributed in urban streams and is of great significance for fishery production (Vander Zanden & Vadeboncoeur, 2002). Except for a good indicator of ongoing eutrophication, this species is highly tolerant and sensitive to a range of insecticides. Our early study published a high-quality genome assembly of this species on chromosome-level (Sun et al., 2021). Therefore, we selected a common insecticide, deltamethrin, to explore the expression pattern of all ABC transporter genes of P. akamusi using genomic resources. Here, the ABC transporter genes of this species were identified and classified at whole-genome level, and we described its genomic distribution, gene structure, and phylogenetic analyses. Based on RNA-seq analysis and RT-qPCR validation, we examined how deltamethrin exposure influenced gene expression of ABC transporters in this midge with different concentrations. We also compared ABC transporters with those of other chironomids to explore its potential function or common traits in non-biting midges. ABC transporter gene families of several chironmids are provided in our work, and lays the foundation for gene function verification on the toxicological researches on P. akamusi.