scholarly journals Conserved residues in the wheat (Triticum aestivum) NAM-A1 NAC domain are required for protein binding and when mutated lead to delayed peduncle and flag leaf senescence

2019 ◽  
Author(s):  
Sophie A. Harrington ◽  
Lauren E. Overend ◽  
Nicolas Cobo ◽  
Philippa Borrill ◽  
Cristobal Uauy
Keyword(s):  
Transcription Factors ◽  
Leaf Senescence ◽  
Protein Function ◽  
Flag Leaf ◽  
Missense Mutations ◽  
In Planta ◽  
Conserved Residues ◽  
Nac Domain ◽  
Nac Transcription Factors ◽  
The Impact

AbstractBackgroundNAC transcription factors contain five highly conserved subdomains which are required for protein dimerisation and DNA binding. Few residues within these subdomains have been identified as essential for protein function, and fewer still have been shown to be of biological relevancein planta. Here we use a positive regulator of senescence in wheat,NAM-A1, to test the impact of missense mutations at specific, highly conserved residues of the NAC domain on protein function.ResultsWe identified missense mutations in five highly conserved residues of the NAC domain ofNAM-A1in a tetraploid TILLING population. TILLING lines containing these mutations, alongside synonymous and non-conserved mutation controls, were grown under glasshouse conditions and scored for senescence. Four of the five mutations showed a significant and consistent delay in peduncle senescence but had no consistent effects on flag leaf senescence. All four mutant alleles with the delayed senescence phenotype also lost the ability to interact with the homoeolog NAM-B1 in a yeast two-hybrid assay. Two of these residues were previously shown to be involved in NAC domain function in Arabidopsis, suggesting conservation of residue function between species. Three of these four alleles led to an attenuated cell death response compared to wild-typeNAM-A1when transiently over-expressed inNicotiana benthamiana. One of these mutations was further tested under field conditions, in which there was a significant and consistent delay in both peduncle and leaf senescence.ConclusionsWe combined field and glasshouse studies of a series of mutant alleles with biochemical analyses to identify four residues of the NAC domain which are required forNAM-A1function and protein interaction. We show that mutations in these residues lead to a gradient of phenotypes, raising the possibility of developing allelic series of mutations for traits of agronomic importance. We also show that mutations inNAM-A1more severely impact peduncle senescence, compared to the more commonly studied flag leaf senescence, highlighting this as an area deserving of further study. The results from this integrated approach provide strong evidence that conserved residues within the functional domains of NAC transcription factors have biological significancein planta.

scholarly journals Conserved residues in the wheat (Triticum aestivum) NAM-A1 NAC domain are required for protein binding and when mutated lead to delayed peduncle and flag leaf senescence

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Sophie A. Harrington ◽  
Lauren E. Overend ◽  
Nicolas Cobo ◽  
Philippa Borrill ◽  
Cristobal Uauy
Keyword(s):  
Transcription Factors ◽  
Leaf Senescence ◽  
Protein Function ◽  
Flag Leaf ◽  
Missense Mutations ◽  
In Planta ◽  
Conserved Residues ◽  
Nac Domain ◽  
Nac Transcription Factors ◽  
The Impact

Abstract Background NAC transcription factors contain five highly conserved subdomains which are required for protein dimerisation and DNA binding. Few residues within these subdomains have been identified as essential for protein function, and fewer still have been shown to be of biological relevance in planta. Here we use a positive regulator of senescence in wheat, NAM-A1, to test the impact of missense mutations at specific, highly conserved residues of the NAC domain on protein function. Results We identified missense mutations in five highly conserved residues of the NAC domain of NAM-A1 in a tetraploid TILLING population. TILLING lines containing these mutations, alongside synonymous and non-conserved mutation controls, were grown under glasshouse conditions and scored for senescence. Four of the five mutations showed a significant and consistent delay in peduncle senescence but had no consistent effects on flag leaf senescence. All four mutant alleles with the delayed senescence phenotype also lost the ability to interact with the homoeolog NAM-B1 in a yeast two-hybrid assay. Two of these residues were previously shown to be involved in NAC domain function in Arabidopsis, suggesting conservation of residue function between species. Three of these four alleles led to an attenuated cell death response compared to wild-type NAM-A1 when transiently over-expressed in Nicotiana benthamiana. One of these mutations was further tested under field conditions, in which there was a significant and consistent delay in both peduncle and leaf senescence. Conclusions We combined field and glasshouse studies of a series of mutant alleles with biochemical analyses to identify four residues of the NAC domain which are required for NAM-A1 function and protein interaction. We show that mutations in these residues lead to a gradient of phenotypes, raising the possibility of developing allelic series of mutations for traits of agronomic importance. We also show that mutations in NAM-A1 more severely impact peduncle senescence, compared to the more commonly studied flag leaf senescence, highlighting this as an area deserving of further study. The results from this integrated approach provide strong evidence that conserved residues within the functional domains of NAC transcription factors have biological significance in planta.

scholarly journals Genome-wide identification and characterization of long non-coding RNAs involved in flag leaf senescence of rice

2021 ◽  
Author(s):  
Xiaoping Huang ◽  
Hongyu Zhang ◽  
Qiang Wang ◽  
Rong Guo ◽  
Lingxia Wei ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Leaf Senescence ◽  
Flag Leaf ◽  
Reduction Process ◽  
Sequencing Analysis ◽  
Biological Processes ◽  
Genome Wide ◽  
A Genome ◽  
Non Coding Rnas ◽  
Systematic Identification

Abstract Key message This study showed the systematic identification of long non-coding RNAs (lncRNAs) involving in flag leaf senescence of rice, providing the possible lncRNA-mRNA regulatory relationships and lncRNA-miRNA-mRNA ceRNA networks during leaf senescence. Abstract LncRNAs have been reported to play crucial roles in diverse biological processes. However, no systematic identification of lncRNAs associated with leaf senescence in plants has been studied. In this study, a genome-wide high throughput sequencing analysis was performed using rice flag leaves developing from normal to senescence. A total of 3953 lncRNAs and 38757 mRNAs were identified, of which 343 lncRNAs and 9412 mRNAs were differentially expressed. Through weighted gene co-expression network analysis (WGCNA), 22 continuously down-expressed lncRNAs targeting 812 co-expressed mRNAs and 48 continuously up-expressed lncRNAs targeting 1209 co-expressed mRNAs were considered to be significantly associated with flag leaf senescence. Gene Ontology results suggested that the senescence-associated lncRNAs targeted mRNAs involving in many biological processes, including transcription, hormone response, oxidation–reduction process and substance metabolism. Additionally, 43 senescence-associated lncRNAs were predicted to target 111 co-expressed transcription factors. Interestingly, 8 down-expressed lncRNAs and 29 up-expressed lncRNAs were found to separately target 12 and 20 well-studied senescence-associated genes (SAGs). Furthermore, analysis on the competing endogenous RNA (CeRNA) network revealed that 6 down-expressed lncRNAs possibly regulated 51 co-expressed mRNAs through 15 miRNAs, and 14 up-expressed lncRNAs possibly regulated 117 co-expressed mRNAs through 21 miRNAs. Importantly, by expression validation, a conserved miR164-NAC regulatory pathway was found to be possibly involved in leaf senescence, where lncRNA MSTRG.62092.1 may serve as a ceRNA binding with miR164a and miR164e to regulate three transcription factors. And two key lncRNAs MSTRG.31014.21 and MSTRG.31014.36 also could regulate the abscisic-acid biosynthetic gene BGIOSGA025169 (OsNCED4) and BGIOSGA016313 (NAC family) through osa-miR5809. The possible regulation networks of lncRNAs involving in leaf senescence were discussed, and several candidate lncRNAs were recommended for prior transgenic analysis. These findings will extend the understanding on the regulatory roles of lncRNAs in leaf senescence, and lay a foundation for functional research on candidate lncRNAs.

scholarly journals In Planta Mutagenesis Determines the Functional Regions of the Wheat Puroindoline Proteins

Genetics ◽  
2009 ◽  
Vol 183 (3) ◽  
pp. 853-860 ◽  
Author(s):  
Leila Feiz ◽  
Brian S. Beecher ◽  
John M. Martin ◽  
Michael J. Giroux
Keyword(s):  
Functional Analysis ◽  
Allelic Variation ◽  
Screening Method ◽  
Point Mutations ◽  
Missense Mutations ◽  
Grain Hardness ◽  
In Planta ◽  
Rich Region ◽  
Critical Function ◽  
The Impact

In planta analysis of protein function in a crop plant could lead to improvements in understanding protein structure/function relationships as well as selective agronomic or end product quality improvements. The requirements for successful in planta analysis are a high mutation rate, an efficient screening method, and a trait with high heritability. Two ideal targets for functional analysis are the Puroindoline a and Puroindoline b (Pina and Pinb, respectively) genes, which together compose the wheat (Triticum aestivum L.) Ha locus that controls grain texture and many wheat end-use properties. Puroindolines (PINs) together impart soft texture, and mutations in either PIN result in hard seed texture. Studies of the PINs' mode of action are limited by low allelic variation. To create new Pin alleles and identify critical function-determining regions, Pin point mutations were created in planta via EMS treatment of a soft wheat. Grain hardness of 46 unique PIN missense alleles was then measured using segregating F2:F3 populations. The impact of individual missense alleles upon PIN function, as measured by grain hardness, ranged from neutral (74%) to intermediate to function abolishing. The percentage of function-abolishing mutations among mutations occurring in both PINA and PINB was higher for PINB, indicating that PINB is more critical to overall Ha function. This is contrary to expectations in that PINB is not as well conserved as PINA. All function-abolishing mutations resulted from structure-disrupting mutations or from missense mutations occurring near the Tryptophan-rich region. This study demonstrates the feasibility of in planta functional analysis of wheat proteins and that the Tryptophan-rich region is the most important region of both PINA and PINB.

scholarly journals Genotype-Phenotype correlation in Dravet Syndrome with SCN1A mutation increase efficiency of molecular diagnosis

2012 ◽  
Vol 18 (2) ◽  
pp. 60-62
Author(s):  
MC Gonsales ◽  
P Preto ◽  
MA Montenegro ◽  
MM Guerreiro ◽  
I Lopes-Cendes
Keyword(s):  
Protein Function ◽  
Copy Number ◽  
Mutation Screening ◽  
Copy Number Variants ◽  
Febrile Seizures ◽  
Copy Number Variations ◽  
Dravet Syndrome ◽  
Missense Mutations ◽  
The Impact ◽  
Doose Syndrome

OBJECTIVES: The purpose of this study was to advance the knowledge on the clinical use of SCN1A testing for severe epilepsies within the spectrum of generalized epilepsy with febrile seizures plus by performing genetic screening in patients with Dravet and Doose syndromes and establishing genotype-phenotype correlations. METHODS: Mutation screening in SCN1A was performed in 15 patients with Dravet syndrome and 13 with Doose syndrome. Eight prediction algorithms were used to analyze the impact of the mutations in putative protein function. Furthermore, all SCN1A mutations previously published were compiled and analyzed. In addition, Multiplex Ligation-Dependent Probe Amplification (MLPA) technique was used to detect possible copy number variations within SCN1A. RESULTS: Twelve mutations were identified in patients with Dravet syndrome, while patients with Doose syndrome showed no mutations. Our results show that the most common type of mutation found is missense, and that they are mostly located in the pore region and the N- and C-terminal of the protein. No copy number variants in SCN1A were identified in our cohort. CONCLUSIONS: SCN1A testing is clinically useful for patients with Dravet syndrome, but not for those with Doose syndrome, since both syndromes do not seem to share the same genetic basis. Our results indicate that indeed missense mutations can cause severe phenotypes depending on its location and the type of amino-acid substitution. Moreover, our strategy for predicting deleterious effect of mutations using multiple computation algorithms was efficient for most of the mutations identified.

DNA binding by the plant-specific NAC transcription factors in crystal and solution: a firm link to WRKY and GCM transcription factors

2012 ◽  
Vol 444 (3) ◽  
pp. 395-404 ◽  
Author(s):  
Ditte H. Welner ◽  
Søren Lindemose ◽  
J. Günter Grossmann ◽  
Niels Erik Møllegaard ◽  
Addie N. Olsen ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Stress Responses ◽  
Crystal Form ◽  
Nutrient Distribution ◽  
Nac Domain ◽  
X Ray ◽  
Nac Transcription Factors ◽  
X Ray Crystallography ◽  
Dna Complex

NAC (NAM/ATAF/CUC) plant transcription factors regulate essential processes in development, stress responses and nutrient distribution in important crop and model plants (rice, Populus, Arabidopsis), which makes them highly relevant in the context of crop optimization and bioenergy production. The structure of the DNA-binding NAC domain of ANAC019 has previously been determined by X-ray crystallography, revealing a dimeric and predominantly β-fold structure, but the mode of binding to cognate DNA has remained elusive. In the present study, information from low resolution X-ray structures and small angle X-ray scattering on complexes with oligonucleotides, mutagenesis and (DNase I and uranyl photo-) footprinting, is combined to form a structural view of DNA-binding, and for the first time provide experimental evidence for the speculated relationship between plant-specific NAC proteins, WRKY transcription factors and the mammalian GCM (Glial cell missing) transcription factors, which all use a β-strand motif for DNA-binding. The structure shows that the NAC domain inserts the edge of its core β-sheet into the major groove, while leaving the DNA largely undistorted. The structure of the NAC–DNA complex and a new crystal form of the unbound NAC also indicate limited flexibility of the NAC dimer arrangement, which could be important in recognizing suboptimal binding sites.

scholarly journals SNAC‐As, stress‐responsive NAC transcription factors, mediate ABA‐inducible leaf senescence

2015 ◽  
Vol 84 (6) ◽  
pp. 1114-1123 ◽  
Author(s):  
Hironori Takasaki ◽  
Kyonoshin Maruyama ◽  
Fuminori Takahashi ◽  
Miki Fujita ◽  
Takuya Yoshida ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Leaf Senescence ◽  
Nac Transcription Factors ◽  
As Stress

scholarly journals Genome-wide and comparative phylogenetic analysis of senescence-associated NAC transcription factors in sunflower (Helianthus annuus)

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Sofia A. Bengoa Luoni ◽  
Alberto Cenci ◽  
Sebastian Moschen ◽  
Salvador Nicosia ◽  
Laura M. Radonic ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Transcription Factors ◽  
Leaf Senescence ◽  
Large Scale ◽  
Gene Families ◽  
Orthologous Group ◽  
Specific Gene ◽  
Significant Differential Expression ◽  
Nac Transcription Factors ◽  
Functional Relationships

Abstract Background Leaf senescence delay impacts positively in grain yield by maintaining the photosynthetic area during the reproductive stage and during grain filling. Therefore a comprehensive understanding of the gene families associated with leaf senescence is essential. NAC transcription factors (TF) form a large plant-specific gene family involved in regulating development, senescence, and responses to biotic and abiotic stresses. The main goal of this work was to identify sunflower NAC TF (HaNAC) and their association with senescence, studying their orthologous to understand possible functional relationships between genes of different species. Results To clarify the orthologous relationships, we used an in-depth comparative study of four divergent taxa, in dicots and monocots, with completely sequenced genomes (Arabidopsis thaliana, Vitis vinifera, Musa acuminata and Oryza sativa). These orthologous groups provide a curated resource for large scale protein sequence annotation of NAC TF. From the 151 HaNAC genes detected in the latest version of the sunflower genome, 50 genes were associated with senescence traits. These genes showed significant differential expression in two contrasting lines according to an RNAseq assay. An assessment of overexpressing the Arabidopsis line for HaNAC001 (a gene of the same orthologous group of Arabidopsis thaliana ORE1) revealed that this line displayed a significantly higher number of senescent leaves and a pronounced change in development rate. Conclusions This finding suggests HaNAC001 as an interesting candidate to explore the molecular regulation of senescence in sunflower.

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