3-Hydroxy-3-methyl glutaryl coenzyme A reductase is required for ovarian development in the oriental fruit fly Bactrocera dorsalis (Hendel)

Highlights

• A HMGR gene was identified in Bactrocera dorsalis.

• BdHMGR was highly expressed in the pupal and adult stages.

• BdHMGR was mainly distributed in the ovary.

• BdHMGR is potentially involved in ovarian development.

Abstract

3-Hydroxy-3-methyl glutaryl coenzyme A reductase (HMGR) is an important rate-limiting enzyme in the mevalonate pathway, which is critical for the synthesis of juvenile hormone and insect development. In this study, a full-length cDNA encoding HMGR (BdHMGR) was cloned from the oriental fruit fly, Bactrocera dorsalis (Hendel). The open reading frame of BdHMGR encoded 928 amino acids and shared a high degree of identity with other known HMGRs. The expression levels of BdHMGR were high during the early and lately pupal stages, and were low at the larval and mid pupal stages. In the adult stages, the highest level of BdHMGR was detected on day-10 of the adult lifecycle, and the expression levels in females were greater than those in males. The highest tissue-specific expression was in the ovary, followed by fat body and other tissues. Injection of double-stranded RNA (dsRNA) of BdHMGR into adult females significantly reduced BdHMGR transcript levels, and inhibited ovarian development and affected the expression of two yolk protein genes. After dissection, we found that one or two of the spheroids of ovaries from the BdHMGR dsRNA-injected flies were amorphous and reduced in size as compared with the controls. The results suggest that BdHMGR is required for ovarian development in B. dorsalis and may be a potential target for pest control.

Introduction

Juvenile hormone (JH) is a sesquiterpenoid hormone that plays crucial roles in the regulation of growth, development, metamorphosis and reproduction in insects (Wyatt and Davey, 1996; Riddiford, 2012). JH is synthesized in the corpora allata of insects and secreted into the hemolymph to prevent metamorphosis and regulate reproductive maturation (Gilbert et al., 2000; Li et al., 2003; Riddiford, 2012). Two distinct metabolic parts for JH biosynthesis have been suggested: the early part comprises the mevalonate pathway, which utilizes acetyl-CoA to form farnesyl diphosphate (FPP) (Bellés et al., 2005). The later part is specific to insects and crustaceans where FPP is converted to JH through the intermediates of farnesol and farnesoic acid (Cheng et al., 2014). The mevalonate pathway is composed of several reactions that are catalyzed by eight enzymes and is conserved in both vertebrates and invertebrates (Noriega, 2014). Generally, the mevalonate pathway begins with reactions catalyzed by acetyl coenzyme A followed by 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA), HMG-CoA synthase (HMGS), HMG-CoA reductase (HMGR), and FPP synthetase to form FPP ((Noriega, 2014). The insect mevalonate pathway primarily functions to produce many essential molecules, such as JH, monoterpene pheromones, and defensive secretions, which are associated with insect growth, reproduction, chemical communication, and defense (Hall et al., 2002a,b; Bellés et al., 2005; Taban et al., 2006; Vranova and Coman, 2013; Zhang et al., 2017).

HMGR (EC 1.1.1.88) is an important rate-limiting enzyme that converts HMG-CoA to mevalonate (Bellés et al., 2005). To date, HMGR homologues have been identified in many insect species, including Dendroctonus armandi (Yu et al., 2015), Drosophila melanogaster (Gertler et al., 1988), Bombyx mori (Kinjoh et al., 2007), Ips pini (Keeling et al., 2004), Helicoverpa armigera (Wang et al., 2013), Agrotis ipsilon (Duportets et al., 2000), Diploptera punctata (Huang et al., 2015), and Blattela germanica (Martínez-González et al., 1993). The copy numbers of the HMGR gene vary, and two genes occur in Leptinotarsa decemlineata (Li et al., 2016) and Tribolium castaneum (Cheng et al., 2014). Previous studies have demonstrated that HMGR might be involved in the biosynthesis of various pheromones and regulation of development and reproduction in insects. Functional characterization of HMGR genes in D. punctata and H. armigera through RNA interference (RNAi) has revealed their critical roles in regulation of embryonic development and vitellogenin synthesis (Wang et al., 2013; Huang et al., 2015). In B. mori and Spodoptera litura, the inhibitors of HMGR inhibited sex pheromone biosynthesis (Fonagy et al., 1992; Bjostad and Roelofs, 1984). Further, interference with expression of HMGR in adults of E. chinensis resulted in the decrease of cantharidin production, while application of exogenous JH in males raised the amount of cantharidin (Lü et al., 2016).

The oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), is one of the most destructive insects being responsible for significant economic losses in a wide range of vegetables and fruits (Clarke et al., 2005, Stephens et al., 2007). Currently, chemical control of the fruit flies using a variety of insecticides is still regarded as the most effectively method, but does result in resistance development and environmental pollution (Hsu et al., 2012, Wang et al., 2015). Exploring the vital genes of destructive insects will aid in the development of gene switches to control insect pests (Yu et al., 2014; Kola et al., 2015). It has been demonstrated that ovarian development is critical for modulating reproductive progress in insects (Storto, 1994). Earlier studies in B. dorsalis revealed that many genes such as insulin receptor substrate (Xu et al., 2015), target of rapamycin (Suganya et al., 2010), S6 kinase (Suganya et al., 2011), double-sex proteins (Chen et al., 2008), vitellogenin receptor (Cong et al., 2015), Krüppel-homolog 1 and Methoprene-tolerant (Yue et al., 2018) were essential for reproductive system development, and had been considered as potential targets for pest management. In this study, we identify and characterize the full-length cDNA of BdHMGR from B. dorsalis. Quantitative real-time PCR (qRT-PCR) was used to analyze the expression profiles of BdHMGR in different developmental stages and different tissues. Moreover, the function of BdHMGR was explored using RNAi. These analyses elucidate the important role of BdHMGR in ovarian development, and this gene will be useful for developing new pest control strategies.