Pharmacological and transcriptomic analyses of a B. cinerea flavohemoglobin deficient mutant demonstrate that nitric oxide affects germination, DNA replication and cell cycle

Francisco Anta 1 Daniela Santander 1,4 Wilson Acosta 1 Pedro San Segundo 3 Rodrigo Santamaría 2 Ernesto P. Benito 1 Jose M. Diaz-Minguez 1
1CIALE (Instituto Hispano-Luso de Investigaciones Agrarias), Departamento de Microbiología y Genética, Universidad de Salamanca, Villamayor, Spain
2Departamento de Informática y Automática, Facultad de Ciencias, Universidad de Salamanca, Salamanca, Spain
3Instituto de Biología Funcional y Genómica, CSIC-Universidad de Salamanca, Salamanca, Spain
4Facultad de Ciencias Agropecuarias y Ambientales, Universidad Técnica del Norte, Ibarra, Ecuador

Nitric oxide (NO) is a highly reactive molecule with fundamental roles in the biology of all living systems. The participation of NO in developmental processes has been described in several fungal species. However, the nature of the mechanisms and factors being affected by NO in fungi is poorly characterized. Botrytis cinerea is a plant pathogenic fungus which has attracted much attention given its wide host range and its necrotrophic life style. The production of NO by the fungus has been demonstrated as well as its detoxification by means of a flavohemoglobin enzyme encoded by gene Bcfhg1. It has been suggested that the physiological functions of the flavohemoglobin could be related to its involvement in the modulation of endogenous NO levels produced by the fungus during specific developmental stages.

Pharmacological studies in which germinating spores and mature mycelium were exposed either to NO donors or to NO scavengers indicate that NO affects germination. From the data obtained, in combination with the results derived from the analysis of the response of ΔBcfhg1 germinating spores to NO, it can be concluded that NO exerts and inmediate and transitoy effect on germination efficiency, on germ tube elongation and on nuclear division rate. Global expression analysis of ΔBcfhg1 in these conditions detected major changes in the expression pattern with about one third of the genes predicted in the B. cinerea genome responding to exposition to NO. Functional enrichment analysis allowed to identify links between exposition to NO, growth arrest and down-regulation of “DNA replication”, “nucleolus” and “cell cycle” genes.