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      Argonaute1 and Gawky Are Required for the Development and Reproduction of Melon fly, Zeugodacus cucurbitae

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          Abstract

          Argonaute family genes encode a highly conserved group of proteins that have been associated with RNA silencing in both animals and plants. This study investigates the importance of microRNA biogenesis key regulators Argonaute1 ( Ago1) and Gawky genes in the post-embryonic and ovarian development of the melon fly, Zeugodacus cucurbitae. The expression levels of these genes were mapped in all developmental stages and different adult tissues. Their roles in development were investigated using RNA interference (RNAi) via two different dsRNA delivery techniques. Embryo microinjection and oral feeding of third instar larvae successfully knocked down and greatly reduced the expression level of the target genes. Additionally, ex vivo essays revealed the stability of dsRNA in food was sufficient for gene silencing, although its integrity was affected in midgut. A wide range of phenotypes were observed on pupation, segmentation, pigmentation, and ovarian development. RNAi-mediated silencing of Gawky caused high mortality and loss of body segmentation, while Ago1 knockdown affected ovarian development and pigmentation. Developmental abnormalities and ovarian malformations caused by silencing these genes suggest that these genes are crucial for viability and reproductive capacity of Z. cucurbitae, and may be used as potential target genes in pest management.

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          Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

          Kenneth Livak, Thomas D. Schmittgen (2001)
          The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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            Origins and Mechanisms of miRNAs and siRNAs.

            Richard W. Carthew, Erik J. Sontheimer (2009)
            Over the last decade, approximately 20-30 nucleotide RNA molecules have emerged as critical regulators in the expression and function of eukaryotic genomes. Two primary categories of these small RNAs--short interfering RNAs (siRNAs) and microRNAs (miRNAs)--act in both somatic and germline lineages in a broad range of eukaryotic species to regulate endogenous genes and to defend the genome from invasive nucleic acids. Recent advances have revealed unexpected diversity in their biogenesis pathways and the regulatory mechanisms that they access. Our understanding of siRNA- and miRNA-based regulation has direct implications for fundamental biology as well as disease etiology and treatment.
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              Overview of MicroRNA Biogenesis, Mechanisms of Actions, and Circulation

              Jacob O'Brien, Heyam Hayder, Yara Zayed … (2018)
              MicroRNAs (miRNAs) are a class of non-coding RNAs that play important roles in regulating gene expression. The majority of miRNAs are transcribed from DNA sequences into primary miRNAs and processed into precursor miRNAs, and finally mature miRNAs. In most cases, miRNAs interact with the 3′ untranslated region (3′ UTR) of target mRNAs to induce mRNA degradation and translational repression. However, interaction of miRNAs with other regions, including the 5′ UTR, coding sequence, and gene promoters, have also been reported. Under certain conditions, miRNAs can also activate translation or regulate transcription. The interaction of miRNAs with their target genes is dynamic and dependent on many factors, such as subcellular location of miRNAs, the abundancy of miRNAs and target mRNAs, and the affinity of miRNA-mRNA interactions. miRNAs can be secreted into extracellular fluids and transported to target cells via vesicles, such as exosomes, or by binding to proteins, including Argonautes. Extracellular miRNAs function as chemical messengers to mediate cell-cell communication. In this review, we provide an update on canonical and non-canonical miRNA biogenesis pathways and various mechanisms underlying miRNA-mediated gene regulations. We also summarize the current knowledge of the dynamics of miRNA action and of the secretion, transfer, and uptake of extracellular miRNAs.
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                Author and article information

                Contributors
                Journal
                Journal ID (nlm-ta): Front Genet
                Journal ID (iso-abbrev): Front Genet
                Journal ID (publisher-id): Front. Genet.
                Title: Frontiers in Genetics
                Publisher: Frontiers Media S.A.
                ISSN (Electronic): 1664-8021
                Publication date (Electronic): 23 June 2022
                Publication date Collection: 2022
                Volume: 13
                Electronic Location Identifier: 880000
                Affiliations
                [1] 1 School of Life Sciences , Hainan University , Haikou, China
                [2] 2 School of Plant Protection , Hainan University , Haikou, China
                [3] 3 Key Laboratory of Green Prevention and Control of Tropical Plant Diseases and Pests, Ministry of Education, School of Plant Protection , Hainan University , Haikou, China
                [4] 4 Hainan Yazhou Bay Seed Lab , Sanya, China
                Author notes

                Edited by: Sufang Zhang, Chinese Academy of Forestry, China

                Reviewed by: Xun Zhang, Chinese Academy of Forestry, China

                Kang He, Zhejiang University, China

                Qing Chen, Chinese Academy of Tropical Agricultural Sciences, China

                Asad Ali, Abdul Wali Khan University, Pakistan

                *Correspondence: Xianwu Lin, linxianw@ 123456hainanu.edu.cn ; Rihui Yan, ryan1@ 123456hainanu.edu.cn

                This article was submitted to Epigenomics and Epigenetics, a section of the journal Frontiers in Genetics

                Article
                Publisher ID: 880000
                DOI: 10.3389/fgene.2022.880000
                PMC ID: 9260231
                SO-VID: 64244570-050b-4729-ad1f-030f1715062d
                Copyright © Copyright © 2022 Jamil, Ahmad, Ran, Ma, Cao, Lin and Yan.
                License:

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                Date received : 20 February 2022
                Date accepted : 04 May 2022
                Categories
                Subject: Genetics
                Subject: Original Research

                ScienceOpen disciplines: Genetics
                Keywords: zeugodacus cucurbitae,argonaute,functional analysis,rna interference (rnai),ovarian development,mortality
                Data availability:
                ScienceOpen disciplines: Genetics
                Keywords: zeugodacus cucurbitae, argonaute, functional analysis, rna interference (rnai), ovarian development, mortality

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