Antisense Oligonucleotide as a New Technology Application for CsLOB1 Gene Silencing Aiming at Citrus Canker Resistance

Author(s)

de Lima, Luiz Felipe Franco ; Carvalho, Isis Gabriela Barbosa ; de Souza-Neto, Reinaldo Rodrigues ; dos Santos, Larissa da Silva ; Nascimento, Cesar Augusto ; Takita, Marco Aurelio ; Tavora, Fabiano Touzdjian Pinheiro Kohlrausch ; Mehta, Angela ; de Souza, Alessandra Alves

• Document type: Journal article
• Source: PHYTOPATHOLOGY; v. 114, n. 8, p. 8-pg., 2024-08-14.

Abstract

Citrus canker disease, caused by Xanthomonas citri subsp. citri, poses a significant threat to global citrus production. The control of the disease in the field relies mainly on the use of conventional tools such as copper compounds, which are harmful to the environment and could lead to bacterial resistance. This scenario stresses the need for new and sustainable technologies to control phytopathogens, representing a key challenge in developing studies that translate basic into applied knowledge. During infection, X. citri subsp. citri secretes a transcriptional activator-like effector that enters the nucleus of plant cells, activating the expression of the canker susceptibility gene LATERAL ORGAN BOUNDARIES 1 (LOB1). In this study, we explored the use of antisense oligonucleotides (ASOs) with phosphorothioate modifications to transiently inhibit the gene expression of CsLOB1 in Citrus sinensis. We designed and validated three potential ASO sequences, which led to a significant reduction in disease symptoms compared with the control. The selected ASO3-CsLOB1 significantly decreased the expression level of CsLOB1 when delivered through two distinct delivery methods, and the reduction of the symptoms ranged from approximately 15 to 83%. Notably, plants treated with ASO3 did not exhibit an increase in symptom development over the evaluation period. This study highlights the efficacy of ASO technology, based on short oligonucleotide chemically modified sequences, as a promising tool for controlling phytopathogens without the need for genetic transformation or plant regeneration. Our results demonstrate the potential of ASOs as a biotechnological tool for the management of citrus canker disease. (AU)

• FAPESP's process: 21/09689-8 - Antisense oligonucleotides application for silencing Citrus sinensis genes pds, lob1 and calS7
• Grantee: Larissa da Silva dos Santos
• Support Opportunities: Scholarships in Brazil - Technical Training Program - Technical Training
• FAPESP's process: 17/13885-1 - Use of CRISPR/Cas9 in the editing of susceptibility genes of citrus sinensis to Xanthomonas citri subsp. Citri
• Grantee: Reinaldo Rodrigues de Souza Neto
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 20/04795-1 - Transformation of sweet orange varieties for functional study and / or gene editing for resistance to citrus bacterial diseases
• Grantee: Isis Gabriela Barbosa Carvalho
• Support Opportunities: Scholarships in Brazil - Technical Training Program - Technical Training
• FAPESP's process: 19/23217-1 - Nanoformulation development aiming at sustainable management of agricultural plant pathogens
• Grantee: Alessandra Alves de Souza
• Support Opportunities: Regular Research Grants
• FAPESP's process: 22/04112-7 - Antisense oligonucleotides application for silencing in Citrus sinensis genes LOB1 e EXPANSINA
• Grantee: Cesar Augusto Nascimento
• Support Opportunities: Scholarships in Brazil - Technical Training Program - Technical Training