Carbon catabolite repression in filamentous fungi is regulated by phosphorylation of the transcription factor CreA

Filamentous
fungi of the genus Aspergillus, are
of particular interest for biotechnological applications due to their natural
capacity to secrete carbohydrate-active enzymes (CAZy) that target plant
biomass. The presence of easily metabolizable sugars such as glucose, whose
concentrations increase during plant biomass hydrolysis, results in the
repression of CAZy-encoding genes, in a process known as carbon catabolite
repression (CCR), which is undesired for the purpose of large-scale enzyme
production. To date, the C2H2 transcription factor CreA
has been described as the major CC-repressor in Aspergillus spp, although little is known about the role of
post-translational modifications in this process. In this work, phosphorylation
sites were identified by mass spectrometry on Aspergillus nidulans CreA, and subsequently, the previously
identified but uncharacterized site S262, the characterized site S319, and the
newly identified sites S268 and T308, were chosen to be mutated to
non-phosphorylatable residues before their effect on CCR was investigated. Sites S262, S268 and T308 are important for
CreA protein accumulation and cellular localization, DNA binding and repression
of enzyme activities. In agreement with a previous study, site S319 was not
important for several here tested phenotypes, but is key for CreA degradation
and induction of enzyme activities. All sites were shown to be important for
glycogen and trehalose metabolism. This study highlights the importance of CreA
phosphorylation sites for the regulation of CCR. These sites are interesting
targets for biotechnological strain engineering without the need to delete
essential genes, which could result in undesired side effects.