scholarly journals Transient Gene Expression is an Effective Experimental Tool for the Research into the Fine Mechanisms of Plant Gene Function: Advantages, Limitations, and Solutions

Plants ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 1187
Author(s):  
Alexander A. Tyurin ◽  
Alexandra V. Suhorukova ◽  
Ksenia V. Kabardaeva ◽  
Irina V. Goldenkova-Pavlova
Keyword(s):  
Gene Expression ◽  
Transient Expression ◽  
Gene Function ◽  
Transient Gene Expression ◽  
Physiological Role ◽  
Gene Products ◽  
Plant Gene ◽  
Gene Functions ◽  
Insight Into

A large data array on plant gene expression accumulated thanks to comparative omic studies directs the efforts of researchers to the specific or fine effects of the target gene functions and, as a consequence, elaboration of relatively simple and concurrently effective approaches allowing for the insight into the physiological role of gene products. Numerous studies have convincingly demonstrated the efficacy of transient expression strategy for characterization of the plant gene functions. The review goals are (i) to consider the advantages and limitations of different plant systems and methods of transient expression used to find out the role of gene products; (ii) to summarize the current data on the use of the transient expression approaches for the insight into fine mechanisms underlying the gene function; and (iii) to outline the accomplishments in efficient transient expression of plant genes. In general, the review discusses the main and critical steps in each of the methods of transient gene expression in plants; areas of their application; main results obtained using plant objects; their contribution to our knowledge about the fine mechanisms of the plant gene functions underlying plant growth and development; and clarification of the mechanisms regulating complex metabolic pathways.

scholarly journals Harnessing a Transient Gene Expression System in Nicotiana benthamiana to Explore Plant Agrochemical Transporters

Plants ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 524
Author(s):  
Bingqi Wu ◽  
Zhiting Chen ◽  
Xiaohui Xu ◽  
Ronghua Chen ◽  
Siwei Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transient Expression ◽  
Nicotiana Benthamiana ◽  
Heterologous Gene Expression ◽  
Expression System ◽  
Functional Characterization ◽  
Transient Gene Expression ◽  
High Mobility ◽  
Gene Expression System

Functional characterization of plant agrichemical transporters provided an opportunity to discover molecules that have a high mobility in plants and have the potential to increase the amount of pesticides reaching damage sites. Agrobacterium-mediated transient expression in tobacco is simple and fast, and its protein expression efficiency is high; this system is generally used to mediate heterologous gene expression. In this article, transient expression of tobacco nicotine uptake permease (NtNUP1) and rice polyamine uptake transporter 1 (OsPUT1) in Nicotiana benthamiana was performed to investigate whether this system is useful as a platform for studying the interactions between plant transporters and pesticides. The results showed that NtNUP1 increases nicotine uptake in N. benthamiana foliar discs and protoplasts, indicating that this transient gene expression system is feasible for studying gene function. Moreover, yeast expression of OsPUT1 apparently increases methomyl uptake. Overall, this method of constructing a transient gene expression system is useful for improving the efficiency of analyzing the functions of plant heterologous transporter-encoding genes and revealed that this system can be further used to study the functions of transporters and pesticides, especially their interactions.

scholarly journals KH-type splicing regulatory protein is a new component of chromatoid body

Reproduction ◽  
2017 ◽  
Vol 154 (6) ◽  
pp. 723-733 ◽  
Author(s):  
Huijuan Zhang ◽  
Guishuan Wang ◽  
Lin Liu ◽  
Xiaolin Liang ◽  
Yu Lin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Germ Cell ◽  
Knockout Mice ◽  
Binding Protein ◽  
Regulatory Protein ◽  
Somatic Cells ◽  
Chromatoid Body ◽  
Mechanistic Insight ◽  
Insight Into

The chromatoid body (CB) is a specific cloud-like structure in the cytoplasm of haploid spermatids. Recent findings indicate that CB is identified as a male germ cell-specific RNA storage and processing center, but its function has remained elusive for decades. In somatic cells, KH-type splicing regulatory protein (KSRP) is involved in regulating gene expression and maturation of select microRNAs (miRNAs). However, the function of KSRP in spermatogenesis remains unclear. In this study, we showed that KSRP partly localizes in CB, as a component of CB. KSRP interacts with proteins (mouse VASA homolog (MVH), polyadenylate-binding protein 1 (PABP1) and polyadenylate-binding protein 2 (PABP2)), mRNAs (Tnp2 and Odf1) and microRNAs (microRNA-182) in mouse CB. Moreover, KSRP may regulate the integrity of CB via DDX5-miRNA-182 pathway. In addition, we found abnormal expressions of CB component in testes of Ksrp-knockout mice and of patients with hypospermatogenesis. Thus, our results provide mechanistic insight into the role of KSRP in spermatogenesis.

scholarly journals Characterization of the Role of AMP-Activated Protein Kinase in the Regulation of Glucose-Activated Gene Expression Using Constitutively Active and Dominant Negative Forms of the Kinase

2000 ◽  
Vol 20 (18) ◽  
pp. 6704-6711 ◽  
Author(s):  
Angela Woods ◽  
Dalila Azzout-Marniche ◽  
Marc Foretz ◽  
Silvie C. Stein ◽  
Patricia Lemarchand ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Kinase ◽  
Fatty Acid Synthase ◽  
Physiological Role ◽  
Rat Hepatocytes ◽  
Dominant Negative ◽  
Ampk Activity ◽  
Amp Activated Protein Kinase ◽  
Constitutively Active

ABSTRACT In the liver, glucose induces the expression of a number of genes involved in glucose and lipid metabolism, e.g., those encoding L-type pyruvate kinase and fatty acid synthase. Recent evidence has indicated a role for the AMP-activated protein kinase (AMPK) in the inhibition of glucose-activated gene expression in hepatocytes. It remains unclear, however, whether AMPK is involved in the glucose induction of these genes. In order to study further the role of AMPK in regulating gene expression, we have generated two mutant forms of AMPK. One of these (α1312) acts as a constitutively active kinase, while the other (α1DN) acts as a dominant negative inhibitor of endogenous AMPK. We have used adenovirus-mediated gene transfer to express these mutants in primary rat hepatocytes in culture in order to determine their effect on AMPK activity and the transcription of glucose-activated genes. Expression of α1312 increased AMPK activity in hepatocytes and blocked completely the induction of a number of glucose-activated genes in response to 25 mM glucose. This effect is similar to that observed following activation of AMPK by 5-amino-imidazolecarboxamide riboside. Expression of α1DN markedly inhibited both basal and stimulated activity of endogenous AMPK but had no effect on the transcription of glucose-activated genes. Our results suggest that AMPK is involved in the inhibition of glucose-activated gene expression but not in the induction pathway. This study demonstrates that the two mutants we have described will provide valuable tools for studying the wider physiological role of AMPK.

Insight into the physiological role of a compartmentalised ferritin like protein in Rhodospirillum rubrum

2014 ◽  
Vol 31 ◽  
pp. S43-S44
Author(s):  
Jon Marles-Wright ◽  
Didi He ◽  
Atanas Georgiev ◽  
David Clarke
Keyword(s):  
Rhodospirillum Rubrum ◽  
Physiological Role ◽  
Insight Into

The calcitonin receptor regulates osteocyte lacunae acidity during lactation in mice

2021 ◽  
Vol 249 (1) ◽  
pp. 31-41
Author(s):  
Rachel A Davey ◽  
Michele V Clarke ◽  
Suzanne B Golub ◽  
Patricia K Russell ◽  
Jeffrey D Zajac
Keyword(s):  
Gene Expression ◽  
Direct Action ◽  
Physiological Role ◽  
Calcitonin Receptor ◽  
Ph Indicator ◽  
Osteocyte Lacunae ◽  
Osteocytic Osteolysis ◽  
Indicator Dye

The physiological role of calcitonin, and its receptor, the CTR (or Calcr), has long been debated. We previously provided the first evidence for a physiological role of the CTR to limit maternal bone loss during lactation in mice by a direct action on osteocytes to inhibit osteocytic osteolysis. We now extend these findings to show that CTR gene expression is upregulated two- to three-fold in whole bone of control mice at the end of pregnancy (E18) and lactation (P21) compared to virgin controls. This was associated with an increase in osteoclast activity evidenced by increases in osteoclast surface/bone surface and Dcstamp gene expression. To investigate the mechanism by which the CTR inhibits osteocytic osteolysis, in vivo acidification of the osteocyte lacunae during lactation (P14 days) was assessed using a pH indicator dye. A lower pH was observed in the osteocyte lacunae of lactating Global-CTRKOs compared to controls and was associated with an increase in the gene expression of ATPase H+ transporting V0 subunit D2 (Atp6v0d2) in whole bone of Global-CTRKOs at the end of lacation (P21). To determine whether the CTR is required for the replacement of mineral within the lacunae post-lactation, lacunar area was determined 3 weeks post-weaning. Comparison of the largest 20% of lacunae by area did not differ between Global-CTRKOs and controls post-lactation. These results provide evidence for CTR activation to inhibit osteocytic osteolysis during lactation being mediated by regulating the acidity of the lacunae microenvironment, whilst the CTR is dispensable for replacement of bone mineral within lacunae by osteocytes post-lactation.

scholarly journals Biolistic-mediated transient gene expression in shoot apical meristems of the prickly-pear (Opuntia ficus-indica)

1999 ◽  
Vol 42 (3) ◽  
pp. 299-302 ◽  
Author(s):  
Romulo Marino Llamoca-Zárate ◽  
Luiz Ferreira Aguiar Ponte ◽  
Joerg Landsmann ◽  
Francisco de Assis Paiva Campos
Keyword(s):  
Gene Expression ◽  
Transient Expression ◽  
Transient Gene Expression ◽  
Dna Delivery ◽  
Prickly Pear ◽  
Target Tissue ◽  
Gus Gene ◽  
Transformation System ◽  
Opuntia Ficus Indica ◽  
Apical Meristems

We have demonstrated the transient expression of the GUS gene in cells of the meristematic apical dome of Opuntia ficus-indica. DNA delivery into the cells was achieved using a biolistic PDS-1000He instrument from Bio-Rad Laboratories. The transforming DNA was coated in tungsten particles with diameter of 1.3 m m and the distance between the flying disk and the target tissue was 7.5cm and the shooting pressure was adjusted to 1200 psi. This is the first demonstration that the biolistic transformation system can be used to express a transgene in a member of the Cactaceae.

scholarly journals Atlas of gene expression in the mouse kidney: new features of glomerular parietal cells

2011 ◽  
Vol 43 (3) ◽  
pp. 161-173 ◽  
Author(s):  
Lydie Cheval ◽  
Fabien Pierrat ◽  
Carole Dossat ◽  
Mathieu Genete ◽  
Martine Imbert-Teboul ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mouse Kidney ◽  
Thick Ascending Limb ◽  
Proliferating Cells ◽  
Quantitative Expression ◽  
Nephron Segments ◽  
Nephron Segment ◽  
Mrna Markers ◽  
Insight Into

To gain molecular insight into kidney function, we performed a high-resolution quantitative analysis of gene expression in glomeruli and nine different nephron segments dissected from mouse kidney using Serial Analysis of Gene Expression (SAGE). We also developed dedicated bioinformatics tools and databases to annotate mRNA tags as transcripts. Over 800,000 mRNA SAGE tags were sequenced corresponding to >20,000 different mRNA tags present at least twice in at least one library. Hierarchical clustering analysis of tags demonstrated similarities between the three anatomical subsegments of the proximal tubule, between the cortical and medullary segments of the thick ascending limb of Henle's loop, and between the three segments constituting the aldosterone-sensitive distal nephron segments, whereas the glomerulus and distal convoluted tubule clusterized independently. We also identified highly specific mRNA markers of each subgroup of nephron segments and of most nephron segments. Tag annotation also identified numbers of putative antisense mRNAs. This database constitutes a reference resource in which the quantitative expression of a given gene can be compared with that of other genes in the same nephron segment, or between different segments of the nephron. To illustrate possible applications of this database, we performed a deeper analysis of the glomerulus transcriptome that unexpectedly revealed expression of several ion and water carriers; within the glomerulus, they were found to be preferentially expressed in the parietal sheet. It also revealed the major role of the zinc finger transcription factor Wt1 in the specificity of gene expression in the glomerulus. Finally, functional annotation of glomerulus-specific transcripts suggested a high proliferation activity of glomerular cells. Immunolabeling for PCNA confirmed a high percentage of proliferating cells in the glomerulus parietal sheet.

Role of non-specific DNA in reducing coding DNA requirement for transient gene expression with CHO and HEK-293E cells

2012 ◽  
Vol 109 (9) ◽  
pp. 2271-2278 ◽  
Author(s):  
Yashas Rajendra ◽  
Divor Kiseljak ◽  
Sagar Manoli ◽  
Lucia Baldi ◽  
David L. Hacker ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transient Gene Expression

scholarly journals A New Insight into the Physiological Role of Bile Salt Hydrolase among Intestinal Bacteria from the Genus Bifidobacterium

PLoS ONE ◽  
2014 ◽  
Vol 9 (12) ◽  
pp. e114379 ◽  
Author(s):  
Piotr Jarocki ◽  
Marcin Podleśny ◽  
Paweł Glibowski ◽  
Zdzisław Targoński
Keyword(s):  
Bile Salt ◽  
Physiological Role ◽  
Intestinal Bacteria ◽  
Bile Salt Hydrolase ◽  
Insight Into

scholarly journals Autoregulation Is Essential for Precise Temporal and Steady-State Regulation by the Bordetella BvgAS Phosphorelay

2006 ◽  
Vol 189 (5) ◽  
pp. 1974-1982 ◽  
Author(s):  
Corinne L. Williams ◽  
Peggy A. Cotter
Keyword(s):  
Gene Expression ◽  
Steady State ◽  
Response Regulator ◽  
Expression Patterns ◽  
State Regulation ◽  
Temporal Expression ◽  
Multiple Gene ◽  
Insight Into

ABSTRACT The Bordetella BvgAS virulence control system is prototypical of phosphorelays that use a polydomain sensor and a response regulator to control gene expression in response to environmental cues. BvgAS controls the expression of at least three distinct phenotypic phases (Bvg−, Bvgi, and Bvg+) by differentially regulating the expression of at least four classes of genes. Among the loci regulated by BvgAS is bvgAS itself. We investigated the role of autoregulation in the ability of BvgAS to control multiple gene expression patterns in a temporal and steady-state manner by constructing Bordetella bronchiseptica strains in which the bvgAS promoter was replaced with constitutively active promoters. Our results show that positive autoregulation of bvgAS transcription is required for the temporal expression of multiple phenotypic phases that occurs in response to a shift from Bvg−-phase conditions to Bvg+-phase conditions. Autoregulation was also shown to contribute to steady-state regulation; it influences the sensitivity of the system in response to subtle differences in signal intensity. In addition, considered in relation to BvgA and BvgS activities demonstrated in vitro, our results provide insight into how BvgA and BvgS function mechanistically.

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