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However, the specific function of the BR receptor BRL3 under osmotic stress remains largely unexplored outside\n                    <jats:styled-content style=\"fixed-case\">\n                      <jats:italic>Arabidopsis thaliana<\/jats:italic>\n                    <\/jats:styled-content>\n                    . In this study, we used a CRISPR\/dCas9\u2010based transcriptional activation (CRISPRa) system to upregulate the\n                    <jats:styled-content style=\"fixed-case\">\n                      <jats:italic>Nicotiana tabacum<\/jats:italic>\n                    <\/jats:styled-content>\n                    BRASSINOSTEROID INSENSITIVE\u2010LIKE 3 receptor (\n                    <jats:italic>NtBRL3<\/jats:italic>\n                    ) and assessed its impact on osmotic stress tolerance. Synthetic activation vectors were constructed using Loop Assembly, featuring dCas9\u20136TAL\u2013VP128 modules driven by either a constitutive (CaMV35S) or ABA\u2010inducible (\n                    <jats:italic>SlAREB<\/jats:italic>\n                    ) promoter, paired with dual sgRNAs targeting the\n                    <jats:italic>NtBRL3<\/jats:italic>\n                    promoter. Transient\n                    <jats:italic>Agrobacterium<\/jats:italic>\n                    \u2010mediated transformation followed by PEG treatment was used to impose osmotic stress. RT\u2010qPCR confirmed a 3\u2010 to 4\u2010fold activation of\n                    <jats:italic>NtBRL3<\/jats:italic>\n                    transcripts in CRISPRa\u2010infiltrated leaves. The stress\u2010inducible\n                    <jats:italic>SlAREB<\/jats:italic>\n                    promoter produced the strongest improvements, yielding nearly four\u2010fold higher leaf biomass and a five\u2010fold increase in root biomass relative to PEG\u2010stressed controls. Both constructs reduced malondialdehyde (MDA) accumulation, indicating diminished oxidative damage, and modulated osmoprotectant balance, including reduced root proline and increased total soluble solids, particularly under SlAREB\u2010driven activation. Histological segmentation revealed promoter\u2010dependent anatomical remodeling, with\n                    <jats:italic>NtBRL3<\/jats:italic>\n                    \u2010activated plants exhibiting a higher frequency of enlarged leaf cells and expanded tissue domains, consistent with brassinosteroid\u2010mediated structural plasticity. Collectively, these findings demonstrate that CRISPR\/dCas9\u2010mediated transcriptional activation of\n                    <jats:italic>NtBRL3<\/jats:italic>\n                    enhances osmotic stress resilience in tobacco through coordinated biomass recovery, oxidative stress mitigation, osmolyte homeostasis, and tissue remodeling. This transient, non\u2010integrative CRISPRa approach provides a robust synthetic biology framework for dissecting BR signaling and engineering stress\u2010tolerant crops.\n                  <\/jats:p>","DOI":"10.1111\/ppl.70816","type":"journal-article","created":{"date-parts":[[2026,3,10]],"date-time":"2026-03-10T10:02:36Z","timestamp":1773136956000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["<scp>CRISPR<\/scp>\n                    \/\n                    <scp>dCas9<\/scp>\n                    \u2010Mediated\n                    <scp>BRL3<\/scp>\n                    Activation Enhances Growth and Metabolic Resilience Under Osmotic Stress in\n                    <scp>\n                      <i>Nicotiana tabacum<\/i>\n                    <\/scp>"],"prefix":"10.1111","volume":"178","author":[{"given":"Francisca","family":"Parada","sequence":"first","affiliation":[{"name":"Laboratorio de Fisiolog\u00eda del Estr\u00e9s Centro de Estudios Avanzados en Fruticultura (CEAF)  Rengo Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Pamela","family":"Cabedo\u2010D\u00edaz","sequence":"additional","affiliation":[{"name":"Facultad de Ingenier\u00eda y Ciencias Universidad Adolfo Ib\u00e1\u00f1ez  Pe\u00f1alol\u00e9n Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ariel","family":"Cerda","sequence":"additional","affiliation":[{"name":"Centro de Biotecnolog\u00eda Vegetal Universidad Andr\u00e9s Bello  Santiago Chile"},{"name":"ANID\u2010Millennium Nucleus in Data Science for Plant Resilience (PhytoLearning)  Santiago Chile"},{"name":"ANID\u2010Millennium Science Initiative Program \u2010 Millennium Institute for Integrative Biology (iBio)  Santiago Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Claudio","family":"Osorio\u2010Navarro","sequence":"additional","affiliation":[{"name":"Depto. De Sanidad Vegetal, Facultad de Ciencias Agron\u00f3micas Universidad de Chile  Santiago Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jorge A.","family":"Toledo","sequence":"additional","affiliation":[{"name":"Health Sciences Department Universidad de Ays\u00e9n  Coyhaique Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Luis","family":"Villalobos\u2010Gonz\u00e1lez","sequence":"additional","affiliation":[{"name":"Laboratorio de Fisiolog\u00eda del Estr\u00e9s Centro de Estudios Avanzados en Fruticultura (CEAF)  Rengo Chile"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Michael","family":"Handford","sequence":"additional","affiliation":[{"name":"Centro de Biolog\u00eda Molecular Vegetal, Depto. 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