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250508s2025 xx |||||o 00| ||eng c |
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|a 10.1016/j.plantsci.2025.112449
|2 doi
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|a pubmed25n1343.xml
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|a (DE-627)NLM384921582
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|a (NLM)40015341
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|a (PII)S0168-9452(25)00067-6
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|e rakwb
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|a eng
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|a Alok, Anshu
|e verfasserin
|4 aut
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|a CRISPR/dCas9-KRAB mediated transcriptional suppression of NtbHLH47 enhances tolerance to iron stress and modulates iron content in tobacco
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|c 2025
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|a Text
|b txt
|2 rdacontent
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|a ƒaComputermedien
|b c
|2 rdamedia
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|a ƒa Online-Ressource
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|2 rdacarrier
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|a Date Completed 13.03.2025
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|a Date Revised 13.03.2025
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|a published: Print-Electronic
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|a Citation Status MEDLINE
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|a Copyright © 2025 Elsevier B.V. All rights reserved.
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|a Iron homeostasis is a multifaceted regulatory process that needs to be studied to elucidate iron distribution, uptake, and storage in plants. NtbHLH47, a homologue to AtbHLH47, is a negative regulator of iron. The current study deploys CRISPR interference-dCas9-KRAB (Krüppel-associated box) in the transcriptional suppression of NtbHLH47 and its effect on iron uptake by plants. The pHSN6I01 harbouring dCas9-KRAB and gRNA targeting NtbHHLH47 was constructed. Four gRNAs were designed, G1, G2, G3, and G4, located at + 19, + 111, + 232, and + 335 bp upstream from the ATG start codon in the promoter region of NtbHLH47. The NtbHLH47 was repressed in the developed transgenic lines of tobacco and the qRT-PCR analysis showed that target sites G1 and G2 suppressed NtbHLH47 effectively. The transgenic pHSN6I01 +G1 plants were tolerant to the elevated levels of iron, copper, zinc, and magnesium. The root Ferric chelate reductase activity of pHSN6I01 +G1 lines was reduced against wild type. The Perl staining showed high iron content in the roots of the pHSN6I01 +G1 plants. ICP-MS analysis showed increased Fe content in the roots of pHSN6I01 +G1 line suggesting that NtbHLH47 modulates it. The expression of NtbHLH38, NtbHLH100, NtbHLH101, and NtFIT was found to be upregulated in the pHSN6I01 +G1 line. This is the first report of using CRISPRi based on dCas9-KRAB in tobacco and its application in the functional validation of a gene. Using this, NtbHLH47 was transcriptionally suppressed and the generated lines expressed increased levels of iron in the roots of N. tabacum and gave insight in the iron homeostasis
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|a Journal Article
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|a CRISPRi
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|a DCas9
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|a Iron homeostasis
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|a KRAB
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|a Transcription factor
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|a bHLH47
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|a Iron
|2 NLM
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| 650 |
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|a E1UOL152H7
|2 NLM
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| 650 |
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|a Plant Proteins
|2 NLM
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| 700 |
1 |
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|a Chauhan, Hanny
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Rout, Biswaranjan
|e verfasserin
|4 aut
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| 700 |
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|a Pandey, Ashutosh
|e verfasserin
|4 aut
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| 700 |
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|a Singh, Kashmir
|e verfasserin
|4 aut
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| 773 |
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|i Enthalten in
|t Plant science : an international journal of experimental plant biology
|d 1985
|g 354(2025) vom: 15. Mai, Seite 112449
|w (DE-627)NLM098174193
|x 1873-2259
|7 nnas
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| 773 |
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|g volume:354
|g year:2025
|g day:15
|g month:05
|g pages:112449
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|u http://dx.doi.org/10.1016/j.plantsci.2025.112449
|3 Volltext
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