Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i>
Functional analysis of promoter sequences is important to understand the regulation of gene expression. This study aimed to investigate the promoter region of the <i>Lupinus luteus</i> cytoplasmic cyclophilin gene (<i>LlCyP</i>; AF178458). After bioinformatic analysis, four p...
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MDPI AG
2021-05-01
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| Series: | Agriculture |
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| author | Katarzyna Nuc Przemysław Olejnik Mirella Samardakiewicz Przemysław Nuc |
| author_facet | Katarzyna Nuc Przemysław Olejnik Mirella Samardakiewicz Przemysław Nuc |
| author_sort | Katarzyna Nuc |
| collection | DOAJ |
| description | Functional analysis of promoter sequences is important to understand the regulation of gene expression. This study aimed to investigate the promoter region of the <i>Lupinus luteus</i> cytoplasmic cyclophilin gene (<i>LlCyP</i>; AF178458). After bioinformatic analysis, four promoter deletion fragments were fused to the β-glucuronidase reporter gene. We used <i>Lotus japonicus</i> as a model plant. After <i>Agrobacterium rhizogenes</i> transformation of <i>L. japonicus</i>, only the longest promoter region (−1055 bp to ATG) supported the β-glucuronidase expression in root nodule parenchyma. Putative <i>cis</i>-elements located between −1055 and −846 bp were subjected to site-directed mutagenesis. Mutations incorporated in the TGATT and AGATT motifs (cytokinin response) abolished <i>GUS</i> expression in nodules, but the mutated AAAGAT motif (OSE, organ-specific element) still activated the <i>GUS</i> expression in root nodules, mainly in cells surrounding the vascular bundle. Promoter deletion and mutation experiments suggest that <i>cis</i>-elements responsible for gene expression in the nodule are located in the region spanning from −1055 to −846 bp. We constructed a deletion fragment, in which the DNA sequence located between −822 and −198 bp was removed (pCYPMG). The promoter region arranged in the pCYPMG supports the expression in the parenchyma of <i>L. japonicus</i> nodules, but it is lower than the whole promoter region. The obtained results may be useful for transgene expression in determinate and indeterminate root nodules. |
| first_indexed | 2024-03-10T11:31:38Z |
| format | Article |
| id | doaj.art-5d384d82c4e54e6d9ca286da2760ec3b |
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| issn | 2077-0472 |
| language | English |
| last_indexed | 2024-03-10T11:31:38Z |
| publishDate | 2021-05-01 |
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| record_format | Article |
| series | Agriculture |
| spelling | doaj.art-5d384d82c4e54e6d9ca286da2760ec3b2023-11-21T19:12:06ZengMDPI AGAgriculture2077-04722021-05-0111543510.3390/agriculture11050435Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i>Katarzyna Nuc0Przemysław Olejnik1Mirella Samardakiewicz2Przemysław Nuc3Department of Biochemistry and Biotechnology, Poznań University of Life Sciences, Dojazd 11, 60-632 Poznań, PolandDepartment of Biochemistry and Biotechnology, Poznań University of Life Sciences, Dojazd 11, 60-632 Poznań, PolandDepartment of Animal Physiology and Developmental Biology, Institute of Experimental Biology, Faculty of Biology, Adam Mickiewicz University in Poznań, Umultowska 89, 61-614 Poznań, PolandInstitute of Molecular Biology and Biotechnology, Adam Mickiewicz University, Umultowska 89, 61-614 Poznań, PolandFunctional analysis of promoter sequences is important to understand the regulation of gene expression. This study aimed to investigate the promoter region of the <i>Lupinus luteus</i> cytoplasmic cyclophilin gene (<i>LlCyP</i>; AF178458). After bioinformatic analysis, four promoter deletion fragments were fused to the β-glucuronidase reporter gene. We used <i>Lotus japonicus</i> as a model plant. After <i>Agrobacterium rhizogenes</i> transformation of <i>L. japonicus</i>, only the longest promoter region (−1055 bp to ATG) supported the β-glucuronidase expression in root nodule parenchyma. Putative <i>cis</i>-elements located between −1055 and −846 bp were subjected to site-directed mutagenesis. Mutations incorporated in the TGATT and AGATT motifs (cytokinin response) abolished <i>GUS</i> expression in nodules, but the mutated AAAGAT motif (OSE, organ-specific element) still activated the <i>GUS</i> expression in root nodules, mainly in cells surrounding the vascular bundle. Promoter deletion and mutation experiments suggest that <i>cis</i>-elements responsible for gene expression in the nodule are located in the region spanning from −1055 to −846 bp. We constructed a deletion fragment, in which the DNA sequence located between −822 and −198 bp was removed (pCYPMG). The promoter region arranged in the pCYPMG supports the expression in the parenchyma of <i>L. japonicus</i> nodules, but it is lower than the whole promoter region. The obtained results may be useful for transgene expression in determinate and indeterminate root nodules.https://www.mdpi.com/2077-0472/11/5/435<i>Lupinus luteus</i><i>Lotus japonicus</i>cyclophilinβ-glucuronidase (<i>GUS</i>)plant transformation |
| spellingShingle | Katarzyna Nuc Przemysław Olejnik Mirella Samardakiewicz Przemysław Nuc Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> Agriculture <i>Lupinus luteus</i> <i>Lotus japonicus</i> cyclophilin β-glucuronidase (<i>GUS</i>) plant transformation |
| title | Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> |
| title_full | Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> |
| title_fullStr | Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> |
| title_full_unstemmed | Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> |
| title_short | Functional Analysis of the <i>Lupinus luteus</i> Cyclophilin Gene Promoter Region in <i>Lotus japonicus</i> |
| title_sort | functional analysis of the i lupinus luteus i cyclophilin gene promoter region in i lotus japonicus i |
| topic | <i>Lupinus luteus</i> <i>Lotus japonicus</i> cyclophilin β-glucuronidase (<i>GUS</i>) plant transformation |
| url | https://www.mdpi.com/2077-0472/11/5/435 |
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