rnai silencing
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PLoS Biology ◽  
2022 ◽  
Vol 20 (1) ◽  
pp. e3001515
Author(s):  
Maria L. Simões ◽  
Yuemei Dong ◽  
Godfree Mlambo ◽  
George Dimopoulos

Anopheles gambiae melanization-based refractoriness to the human malaria parasite Plasmodium falciparum has rarely been observed in either laboratory or natural conditions, in contrast to the rodent model malaria parasite Plasmodium berghei that can become completely melanized by a TEP1 complement-like system-dependent mechanism. Multiple studies have shown that the rodent parasite evades this defense by recruiting the C-type lectins CTL4 and CTLMA2, while permissiveness to the human malaria parasite was not affected by partial depletion of these factors by RNAi silencing. Using CRISPR/Cas9-based CTL4 knockout, we show that A. gambiae can mount melanization-based refractoriness to the human malaria parasite, which is independent of the TEP1 complement-like system and the major anti-Plasmodium immune pathway Imd. Our study indicates a hierarchical specificity in the control of Plasmodium melanization and proves CTL4 as an essential host factor for P. falciparum transmission and one of the most potent mosquito-encoded malaria transmission-blocking targets.


2021 ◽  
Vol 190 ◽  
pp. 112885
Author(s):  
Keyvan Dastmalchi ◽  
Oseloka Chira ◽  
Mathiu Perez Rodriguez ◽  
Barney Yoo ◽  
Olga Serra ◽  
...  

2021 ◽  
Vol 13 (10) ◽  
pp. 70
Author(s):  
Pietro E. Vicari ◽  
Esther S. Chang ◽  
André L. P. Perondini ◽  
Denise Selivon

In genera Anastrepha, Bactrocera and Ceratitis of the tephritid fruit flies the auto-regulatory function of gene transformer is assumed to be activated by maternal derived mRNA or the proteins of the gene transformer (tra-2 mat) and transformer-2 (tra-2mat). However, this maternal effect was not yet been demonstrated. The objective of the present study was to test the effect of absence of tra-2mat in the eggs on the sex determination of A. sp.1 affinis fraterculus. This was achieved by silencing gene tra-2 in the parental females via the pRNA interference. The data showed that tra-2 was transiently silenced in the female for three weeks period. The progenies sex ratio produced by these females during the silencing of tra-2 depart from 1:1 in favor of males. The excess of males was due to the transformation of a fraction of genotypical female XX embryos into XX males, the so-called pseudomales, Individual F1 males from the offspring of treated females crossed to females from the stock, revealed that majority of them showed regular mating behavior and were fertile. However, no offspring was produced in the crosses by a fraction of males that have produced sperms, showed regular mating behavior but did not transfer sperms to the females. The data allow the conclusion that the absence of tra-2mat in the eggs had impaired the self-regulation of the embryonic gene tra resulting in the transformation of XX embryos into pseudomales and also that these pseudomales are sterile. This effect may be useful improve more sustainable technologies for fruit fly control such as SIT.


Planta ◽  
2021 ◽  
Vol 254 (3) ◽  
Author(s):  
Bill Hendrix ◽  
Wei Zheng ◽  
Matthew J. Bauer ◽  
Ericka R. Havecker ◽  
Jennifer T. Mai ◽  
...  

Abstract Main conclusion 22 nt siRNAs applied to leaves induce production of transitive sRNAs for targeted genes and can enhance local silencing. Systemic silencing was only observed for a GFP transgene. Abstract RNA interference (RNAi) is a gene silencing mechanism important in regulating gene expression during plant development, response to the environment and defense. Better understanding of the molecular mechanisms of this pathway may lead to future strategies to improve crop traits of value. An abrasion method to deliver siRNAs into leaf cells of intact plants was used to investigate the activities of 21 and 22 nt siRNAs in silencing genes in Nicotiana benthamiana and Amaranthus cruentus. We confirmed that both 21 and 22 nt siRNAs were able to silence a green fluorescent protein (GFP) transgene in treated leaves of N. benthamiana, but systemic silencing of GFP occurred only when the guide strand contained 22 nt. Silencing in the treated leaves of N. benthamiana was demonstrated for three endogenous genes: magnesium cheletase subunit I (CHL-I), magnesium cheletase subunit H (CHL-H), and GENOMES UNCOUPLED4 (GUN4). However, systemic silencing of these endogenous genes was not observed. Very high levels of transitive siRNAs were produced for GFP in response to treatment with 22 nt siRNAs but only low levels were produced in response to a 21 nt siRNA. The endogenous genes tested also produced transitive siRNAs in response to 22 nt siRNAs. 22 nt siRNAs produced greater local silencing phenotypes than 21 nt siRNAs for three of the genes. These special properties of 22 nt siRNAs were also observed for the CHL-H gene in A. cruentus. These experiments suggest a functional role for transitive siRNAs in amplifying the RNAi response.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Eva Doleželová ◽  
Tomáš Klejch ◽  
Petr Špaček ◽  
Martina Slapničková ◽  
Luke Guddat ◽  
...  

AbstractAll medically important unicellular protozoans cannot synthesize purines de novo and they entirely rely on the purine salvage pathway (PSP) for their nucleotide generation. Therefore, purine derivatives have been considered as a promising source of anti-parasitic compounds since they can act as inhibitors of the PSP enzymes or as toxic products upon their activation inside of the cell. Here, we characterized a Trypanosoma brucei enzyme involved in the salvage of adenine, the adenine phosphoribosyl transferase (APRT). We showed that its two isoforms (APRT1 and APRT2) localize partly in the cytosol and partly in the glycosomes of the bloodstream form (BSF) of the parasite. RNAi silencing of both APRT enzymes showed no major effect on the growth of BSF parasites unless grown in artificial medium with adenine as sole purine source. To add into the portfolio of inhibitors for various PSP enzymes, we designed three types of acyclic nucleotide analogs as potential APRT inhibitors. Out of fifteen inhibitors, four compounds inhibited the activity of the recombinant APRT1 with Ki in single µM values. The ANP phosphoramidate membrane-permeable prodrugs showed pronounced anti-trypanosomal activity in a cell-based assay, despite the fact that APRT enzymes are dispensable for T. brucei growth in vitro. While this suggests that the tested ANP prodrugs exert their toxicity by other means in T. brucei, the newly designed inhibitors can be further improved and explored to identify their actual target(s).


2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Arun Kumaran Anguraj Vadivel ◽  
Tim McDowell ◽  
Justin B. Renaud ◽  
Sangeeta Dhaubhadel

AbstractGmMYB176 is an R1 MYB transcription factor that regulates multiple genes in the isoflavonoid biosynthetic pathway, thereby affecting their levels in soybean roots. While GmMYB176 is important for isoflavonoid synthesis, it is not sufficient for the function and requires additional cofactor(s). The aim of this study was to identify the GmMYB176 interactome for the regulation of isoflavonoid biosynthesis in soybean. Here, we demonstrate that a bZIP transcription factor GmbZIP5 co-immunoprecipitates with GmMYB176 and shows protein–protein interaction in planta. RNAi silencing of GmbZIP5 reduced the isoflavonoid level in soybean hairy roots. Furthermore, co-overexpression of GmMYB176 and GmbZIP5 enhanced the level of multiple isoflavonoid phytoallexins including glyceollin, isowighteone and a unique O-methylhydroxy isoflavone in soybean hairy roots. These findings could be utilized to develop biotechnological strategies to manipulate the metabolite levels either to enhance plant defense mechanisms or for human health benefits in soybean or other economically important crops.


Genes ◽  
2021 ◽  
Vol 12 (3) ◽  
pp. 355
Author(s):  
Maria-Eleni Gregoriou ◽  
Martin Reczko ◽  
Evdoxia G. Kakani ◽  
Konstantina T. Tsoumani ◽  
Kostas D. Mathiopoulos

In most diploid organisms, mating is a prerequisite for reproduction and, thus, critical to the maintenance of their population and the perpetuation of the species. Besides the importance of understanding the fundamentals of reproduction, targeting the reproductive success of a pest insect is also a promising method for its control, as a possible manipulation of the reproductive system could affect its destructive activity. Here, we used an integrated approach for the elucidation of the reproductive system and mating procedures of the olive fruit fly, Bactrocera oleae. Initially, we performed a RNAseq analysis in reproductive tissues of virgin and mated insects. A comparison of the transcriptomes resulted in the identification of genes that are differentially expressed after mating. Functional annotation of the genes showed an alteration in the metabolic, catalytic, and cellular processes after mating. Moreover, a functional analysis through RNAi silencing of two differentially expressed genes, yellow-g and troponin C, resulted in a significantly reduced oviposition rate. This study provided a foundation for future investigations into the olive fruit fly’s reproductive biology to the development of new exploitable tools for its control.


2021 ◽  
Author(s):  
Lev A. Elkonin ◽  
Valery M. Panin ◽  
Odissey A. Kenzhegulov ◽  
Saule Kh. Sarsenova

Modification of the composition of grain storage proteins is an intensively developing area of plant biotechnology, which is of particular importance for sorghum – high-yielding drought tolerant crop. Compared to other cereals, the majority of sorghum cultivars and hybrids are characterized by reduced nutritional value that is caused by a low content of essential amino acids in the seed storage proteins (kafirins), and resistance of kafirins to protease digestion. RNA interference (RNAi) by suppressing synthesis of individual kafirin subclasses may be an effective approach to solve this problem. In this chapter, we review published reports on RNAi silencing of the kafirin-encoding genes. In addition, we present new experimental data on phenotypic effects of RNAi-silencing of γ-KAFIRIN-1 gene in sorghum cv. Avans. To obtain RNAi mutants with γ-KAFIRIN-1 gene silencing we used Agrobacterium-mediated genetic transformation. Transgenic kernels had modified endosperm type with reduced vitreous layer and significantly improved in vitro protein digestibility (93% vs. 57%, according to the densitometry of SDS-PAGE patterns). SDS-PAGE of transgenic kernels showed lowered level of kafirins and appearance of globulin proteins, which were not observed in the original cultivar. For the first time, the cases of instability of inserted genetic construct were identified: elimination of ubi1-intron that is a constituent part of the genetic construct for RNAi silencing, or nos-promotor governing expression of the marker gene (bar) (in the RNAi mutants of cv. Zheltozernoe 10). The research findings presented in this chapter provide strong evidence that RNA interference can be used for improvement of the nutritional properties of sorghum grain.


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