Second messenger c-di-GMP modulates exopolysaccharide Pea-dependent phenotypes via regulating eppA expression in Pseudomonas putida

2022 ◽  
Author(s):  
Yujie Xiao ◽  
Qingyuan Liang ◽  
Meina He ◽  
Nianqi Wu ◽  
Liang Nie ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Structure Prediction ◽  
Direct Interaction ◽  
Second Messenger ◽  
Genetically Engineered ◽  
Colony Morphology ◽  
Base Pairs ◽  
Pellicle Formation ◽  
Unknown Protein ◽  
Relationship Of

Exopolysaccharides (EPSs) Pea is essential for wrinkly colony morphology, pellicle formation, and robust biofilm production in Pseudomonas putida . The second messenger cyclic diguanylate monophosphate (c-di-GMP) induces wrinkly colony morphology in P. putida through unknown mechanism(s). Herein, we found that c-di-GMP modulated wrinkly colony morphology via regulating expression of eppA ( PP_5586 ), a small individually transcribed gene with 177 base pairs, and this gene was adjacent to the upstream of pea cluster. Phenotype observation revealed that eppA was essential for Pea-dependent phenotypes. The deletion of eppA led to smooth colony morphology and impaired biofilm, which was analogous to the phenotypes with the loss of the entire pea operon. EppA expression was positively regulated by c-di-GMP via the transcriptional effector FleQ, and eppA was essential for the c-di-GMP-induced wrinkly colony morphology. Structure prediction results implied that EppA had two transmembrane regions, and Western blot revealed that EppA was located on cell membrane. Transcriptomic analysis indicated that EppA had no significant effect on transcriptomic profile of P. putida . Bacterial two-hybrid (BTH) assay suggested that there was no direct interaction between EppA and the proteins in pea cluster and adjacent operons. Overall, these findings reveal that EppA is essential for Pea-dependent phenotypes, and that c-di-GMP modulates Pea-dependent phenotypes via regulating eppA expression in P. putida . IMPORTANCE Microbe-secreted EPSs are high molecular weight polysaccharides that have the potential to be used as industrially important biomaterials. The EPS Pea in P. putida is essential for wrinkly colony morphology and pellicle formation. Here, we identified a function-unknown protein EppA, which was also essential for Pea-dependent wrinkly colony morphology and pellicle formation, and EppA was probably involved in Pea secretion. Meanwhile, our results indicated that the second messenger c-di-GMP positively regulated the expression of EppA, resulting in Pea-dependent wrinkly colony morphology. Our results reveal the relationship of c-di-GMP, EppA, and Pea-dependent phenotypes, and provide possible pathway to construct genetically engineered strain for high Pea production.

scholarly journals Cellular L-arginine pools modulate c-di-GMP turnover and biofilm formation in Pseudomonas putida

2020 ◽  
Author(s):  
Laura Barrientos-Moreno ◽  
María Antonia Molina-Henares ◽  
María Isabel Ramos-González ◽  
Manuel Espinosa-Urgel
Keyword(s):  
Pseudomonas Putida ◽  
Biofilm Formation ◽  
Response Regulator ◽  
Second Messenger ◽  
Colony Morphology ◽  
Diguanylate Cyclase ◽  
Arginine Biosynthesis ◽  
Amino Acid Transport System ◽  
Metabolic Signal ◽  
Biofilm Development

<p>The intracellular second messenger cyclic diguanylate (c-di-GMP) is broadly conserved in bacteria, where it influences processes such as virulence, stress resistance and biofilm development. In the plant-beneficial bacterium <em>Pseudomonas putida</em> KT2440, the response regulator with diguanylate cyclase activity CfcR is the main contributor to c-di-GMP levels in the stationary phase of growth. When overexpressed, CfcR increases c-di-GMP levels and gives rise to a pleiotropic phenotype that includes enhanced biofilm formation and crinkly colony morphology. Our group has previously reported that insertion mutants in <em>argG</em> and <em>argH</em>, the genes that encode the last two enzymes in the arginine biosynthesis pathway, do not display the crinkly colony morphology phenotype and show decreased c-di-GMP levels even in the presence of <em>cfcR</em> in multicopy (Ramos-González, M.I. <em>et al.</em> 2016. Front. Microbiol. 7, 1093). Here we present results indicating that L-arginine acts both as an environmental and as a metabolic signal that influences the lifestyles of <em>P. putida</em> through the modulation of c-di-GMP levels and changes in the expression of structural elements of biofilms. Exogenous L-arginine partially restores c-di-GMP levels in arginine biosynthesis mutants, a response that is transduced through CfcR and possibly (an)other diguanylate cyclase(s). At least three periplasmic binding proteins, each forming part of an amino acid transport system, contribute in different ways to the response to external L-arginine. We propose that the turnover of the second messenger c-di-GMP is modulated by the state of global arginine pools in the cell resulting both from anabolism and from uptake.</p>

scholarly journals Variasi Genetik Plutella xylostella L. (Lepidoptera: Plutellidae) berdasarkan Gen Cytochrome C Oxidase I

Jurnal MIPA ◽  
2019 ◽  
Vol 8 (1) ◽  
pp. 1
Author(s):  
Claudius F. Kairupan ◽  
Jantje Pelealu ◽  
Juliet M.E. Mamahit
Keyword(s):  
Cytochrome C ◽  
Cytochrome C Oxidase ◽  
Plutella Xylostella ◽  
Coi Gene ◽  
Cytochrome C Oxidase I ◽  
Base Pairs ◽  
Insecticide Application ◽  
Nucleotide Base ◽  
North Sulawesi ◽  
Relationship Of

Daerah Modoinding dan Tomohon di Sulawesi Utara, dikenal sebagai  daerah penghasil sayuran kubis di Indonesia. Sayuran kubis memiliki hama utama yaitu Plutella xylostella. Penyebab serangga ini dapat bertahan hingga saat ini karena adanya sifat resistensi akibat pemberian insektisida yang berlebihan. Penelitian ini dilakukan untuk menganalisis variasi pada gen cytochrome C oxidase IPlutella xylostella yang diperoleh dari dua lokasi yang berbeda, yaitu Modoinding dan Tomohon. Analisis sekuens menunjukkan adanya perbedaan pasang basa nukleotida dari sampel yang berbeda lokasi. Selain itu, variasi juga ditunjukkan pada sampel yang diperoleh dari basis data GenBank dengan adanya perbedaan 1-14 pasang basa nukleotida dengan spesimen pada penelitian ini. Hubungan kekerabatan gen COI P. xylostella keseluruhan sampel tergolong dalam variasi intraspesies dengan nilai jarak genetik berkisar antara 0-0,022 (0-2,20%).Modoinding and Tomohon areas in North Sulawesi, are known as regions in Indonesia that produce a cabbage. The main pest of cabbage, Plutella xylostella. This insect can survive due to its resistance resulted from prolonged insecticide application. This study aims to analyze genetic variation of COI genes in P. xylostella from Modoinding and Tomohon areas. Sequence analysis showed there were differences in nucleotide base pairs between these locations. In addition, variations were also shown in samples obtained from the GenBank database with differences in 1-14 nucleotide base pairs with specimens in this study. The genetic relationship of P. xylostella COI gene in all samples was classified as intraspecific variation with genetic distance values ranging from 0-0,022 (0-2,20%).D aerah Modoinding dan Tomohon di Sulawesi Utara, dikenal sebagaidaerah penghasil sayuran kubis di Indonesia. Sayuran kubis memilikihama utama yaitu Plutella xylostella. Penyebab serangga ini dapatbertahan hingga saat ini karena adanya sifat resistensi akibat pemberianinsektisida yang berlebihan. Penelitian ini dilakukan untuk menganalisisvariasi pada gen cytochrome C oxidase I Plutella xylostella yang diperolehdari dua lokasi yang berbeda, yaitu Modoinding dan Tomohon. Analisissekuens menunjukkan adanya perbedaan pasang basa nukleotida darisampel yang berbeda lokasi. Selain itu, variasi juga ditunjukkan padasampel yang diperoleh dari basis data GenBank dengan adanyaperbedaan 1-14 pasang basa nukleotida dengan spesimen padapenelitian ini. Hubungan kekerabatan gen COI P. xylostella keseluruhansampel tergolong dalam variasi intraspesies dengan nilai jarak genetikberkisar antara 0-0,022 (0-2,20%).

Computer Simulations of Nonlinear Optical Chromophore Containing Polypeptides

1993 ◽  
Vol 330 ◽  
Author(s):  
Ruth Pachter ◽  
Steven B. Fairchild ◽  
James A. Lupo ◽  
Brian S. Sennett ◽  
Robert L. Crane ◽  
...  
Keyword(s):  
Molecular Dynamics ◽  
Structure Prediction ◽  
Nonlinear Optical ◽  
Integrated Approach ◽  
Genetically Engineered ◽  
Spatial Proximity ◽  
Tertiary Structures ◽  
Eglin C ◽  
Nonlinear Optical Chromophores ◽  
Dynamics Simulations

ABSTRACTIn our continuing efforts towards the design of nonlinear (NLO) optical chromophore containing polypeptides we present an integrated computational approach, in which the design of biomolecular materials with defined secondary and tertiary structures is investigated by means of novel predictive tools, while the effects of the nonlinear optical chromophores are studied with molecular dynamics simulations. A neural network that was trained to predict the spatial proximity of Cα atoms that are less than a given threshold apart, is applied. The double-iterated Kalman filter (DIKF) technique is then employed with a constraints set that includes these pairwise atomic distances, and the distances and angles that define the structure as it is known from the protein's sequence. The results for test cases, particularly Crambin and genetically engineered Eglin-C, demonstrate that this integrated approach is useful for structure prediction at an intermediate resolution. Defined structural motifs of NLO chromophore containing polypeptides are investigated by using molecular dynamics techniques, particularly for the design of coil coiled amphiphatic biopolymers.

scholarly journals GGeneration of an efficient artificial promoter of bovine skeletal muscle alpha-actin gene (ACTA1) through addition of cis-acting element

2015 ◽  
Vol 20 (1) ◽  
Author(s):  
Qian Hu ◽  
Huili Tong ◽  
Dandan Zhao ◽  
Yunkao Cao ◽  
Weiwei Zhang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Core Promoter ◽  
Genetically Engineered ◽  
Actin Gene ◽  
Base Pairs ◽  
Cis Acting ◽  
The Core ◽  
Binding Factor ◽  
Bovine Skeletal Muscle ◽  
Alpha Actin

AbstractThe promoter of skeletal muscle α-actin gene (ACTA1) is highly muscle specific. The core of the bovine ACTA1 promoter extends from +29 to −233, about 262 base pairs (bp), which is sufficient to activate transcription in bovine muscle satellite cells. In this study, analysis by PCR site-specific mutagenesis showed that the cis-acting element SRE (serum response element binding factor) was processed as a transcriptional activator. In order to enhance the bovine ACTA1 promoter’s activity, we used a strategy to modify it. We cloned a fragment containing three SREs from the promoter of ACTA1, and then one or two clones were linked upstream of the core promoter (262 bp) of ACTA1. One and two clones increased the activity of the ACTA1 promoter 3-fold and 10-fold, respectively, and maintained muscle tissue specificity. The modified promoter with two clones could increase the level of ACTA1 mRNA and protein 4-fold and 1.1-fold, respectively. Immunofluorescence results showed that green fluorescence of ACTA1 increased. Additionally, the number of total muscle microfilaments increased. These genetically engineered promoters might be useful for regulating gene expression in muscle cells and improving muscle mass in livestock.

Drosophila Ortholog of Succinyl-CoA Synthetase β Subunit: A Novel Modulator of Drosophila KCNQ Channels

2008 ◽  
Vol 99 (5) ◽  
pp. 2736-2740 ◽  
Author(s):  
Lei Gao ◽  
Hong Fei ◽  
Nathan C. Connors ◽  
Jiaming Zhang ◽  
Irwin B. Levitan
Keyword(s):  
Potassium Channels ◽  
Cho Cells ◽  
Chinese Hamster ◽  
Direct Interaction ◽  
Β Subunit ◽  
Glutathione S Transferase ◽  
Kcnq Channels ◽  
Cell Surface Biotinylation ◽  
Relationship Of ◽  
Succinyl Coa Synthetase

Voltage-gated KCNQ potassium channels are responsible for slowly activating potassium currents in heart, brain, and other tissues. Functional defects of KCNQ channels are linked with many diseases, including epilepsy and cardiac arrhythmias. Therefore KCNQ potassium channels have been widely studied, especially in the CNS. We have identified Drosophila CG11963, which encodes a protein orthologous to the β subunit of mammalian succinyl-CoA synthetase (SCS, also known as succinate thiokinase), as a novel modulator of Drosophila KCNQ channels. Direct interaction of CG11963 and dKCNQ was demonstrated by yeast two-hybrid screen and coimmunoprecipitation. Cell surface biotinylation experiments further confirmed that CG11963 resides on the plasma membrane of tsA-201 cells. Coexpression of CG11963 with dKCNQ shifts the conductance–voltage ( G– V) relationship of dKCNQ channels to more positive membrane potentials in Chinese hamster ovary (CHO) cells. Moreover, directly dialyzing glutathione S-transferase fusion CG11963 protein into CHO cells also shifts the dKCNQ G– V curve rightward. The effect of CG11963 persists in the presence of 1 mM adenosine triphosphate (ATP), a substrate of SCS. Taken together, our data define CG11963 as a new dKCNQ-binding protein capable of modulating the properties of the channel. Our evidence suggests that this modulation is mediated by direct interaction of CG11963 with the channel and is not dependent on ATP.

THE p-WAVE THRESHOLD EFFECT AND QUASIRESONANT SCATTERING

2005 ◽  
Vol 20 (03) ◽  
pp. 187-201 ◽  
Author(s):  
CORNEL HATEGAN ◽  
GERHARD GRAW ◽  
HORIA COMISEL
Keyword(s):  
Reaction Mechanism ◽  
Single Particle ◽  
Strength Function ◽  
Direct Interaction ◽  
Threshold Effect ◽  
Scattering Matrix ◽  
P Wave ◽  
Threshold Models ◽  
Relationship Of ◽  
The Relationship

The p-wave threshold effect is described in terms of the reduced scattering matrix. The relationship of this approach to previous theoretical threshold models is established. We prove that this phenomenon is related to the reaction-mechanism of quasiresonant scattering: a single particle neutron threshold state and direct interaction coupling to open observed channels. Spectroscopic aspects of the threshold effect, both with respect to the magnitude and microstructure, are discussed in terms of the neutron strength function.

scholarly journals Kajian Variasi Sekuens Interspesies dan Filogeni Kelelawar Pteropus sp. Menggunakan Gen COI

Jurnal MIPA ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 71
Author(s):  
Era Monalisa ◽  
Feky Recky Mantiri ◽  
Hanry Jefri Lengkong
Keyword(s):  
Genetic Distance ◽  
Phylogenetic Relationship ◽  
Coi Gene ◽  
Gene Sequences ◽  
Base Pairs ◽  
Probable Explanation ◽  
Nucleotide Base ◽  
Tree Construction ◽  
Most Probable Explanation ◽  
Relationship Of

Penelitian ini bertujuan untuk menganalisis variasi interspesies kelelawar Pteropus sp. dan menjelaskan hubungan filogeni Pteropus sp. dengan spesies Pteropus lain yang terdata di GenBank berdasarkan Gen COI. Analisis sekuens menggunakan Geneious v5.6.4 dan menunjukkan adanya variasi interspesies sekuens gen COI pada ketiga sampel Pteropus sp. yang ditunjukkan oleh adanya perbedaan 5 pasang basa nukleotida pada urutan sekuens sampel nomor 157, 160, 421, 427 dan 652 dengan jarak genetik 0,006. Filogeni Ke-3 sampel kelelawar Pteropus sp. dengan spesies Pteropus lain dilakukan menggunakan MEGAX. Hasil filogeni menunjukkan bahwa sampel yang diteliti merupakan kelelawar dari genus Pteropus tetapi belum dapat dipastikan spesiesnya, karena ketika pohon filogeni dikonstruksikan membentuk satu klaster sendiri. Penjelasan dari proses tersebut adalah penyortiran garis keturunan yang tidak lengkap dan terjadinya hibridisasi, serta diduga bahwa primer yang digunakan kurang mampu dalam membedakan variasi intrespesies terhadap kelelawar genus PteropusThis study aimed to analyze the interspecificvariations of bats from Pteropus sp. and describethe phylogenetic relationship of Pteropus sp. with other Pteropus species recorded inGenBank based on the COI gene. Sequenceanalysis by Geneious v5.6.4 showed interspecificvariations of COI gene sequences in all threesamples of Pteropus sp. which was indicated byvariations in 5 nucleotide base pairs in thesequences number 157, 160, 421, 427 and 652with 0.006 of genetic distance value. Phylogeneticof the 3 bat samples of Pteropus sp. with otherPteropus species was carried out by MEGAX.Phylogenetic analyses showed that the samplesstudied are bats of the genus Pteropus, but theexact species cannot be determined because thesamples were grouped in the same cluster duringphylogenetic tree construction. The most probable explanation for this observation is hybridization between two different Pteropus spesies and also it is assumed that the primersused are not capable to distinguish interspecificvariations of the bats from the Pteropus genusPenelitian ini bertujuan untuk menganalisis variasi interspesies kelelawar Pteropus sp. dan menjelaskan hubungan filogeni Pteropus sp. dengan spesies Pteropus lain yang terdata di GenBank berdasarkan Gen COI. Analisis sekuens menggunakan Geneious v5.6.4 dan menunjukkan adanya variasi interspesies sekuens gen COI pada ketiga sampel Pteropus sp. yang ditunjukkan oleh adanya perbedaan 5 pasang basa nukleotida pada urutan sekuens sampel nomor 157, 160, 421, 427 dan 652 dengan jarak genetik 0,006. Filogeni Ke-3 sampel kelelawar Pteropus sp. dengan spesies Pteropus lain dilakukan menggunakan MEGAX. Hasil filogeni menunjukkan bahwa sampel yang diteliti merupakan kelelawar dari genus Pteropus tetapi belum dapat dipastikan spesiesnya, karena ketika pohon filogeni dikonstruksikan membentuk satu klaster sendiri. Penjelasan dari proses tersebut adalah penyortiran garis keturunan yang tidak lengkap dan terjadinya hibridisasi, serta diduga bahwa primer yang digunakan kurang mampu dalam membedakan variasi intrespesies terhadap kelelawar genus Pteropus

Sign in / Sign up

Export Citation Format

Share Document