OSMaC strategies to discover new natural products

• Autor(a) principal: Ferreira, Dora de Fátima Medeiros
• Data de Publicação: 2023
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.14/43044
• topic: Natural products; Cyanobacteria; OSMaC strategies; Siderophores; Produtos naturais; Cianobactérias; Estratégias OSMaC; Sideróforos; Domínio/Área Científica::Ciências Naturais::Ciências Biológicas

Cyanobacteria are photosynthetic microorganisms known to produce a wide range of secondary metabolites, from toxins to other compounds with recognized biotechnological and pharmaceutical applications. Recent advances in Genomics and Bioinformatics have shown that cyanobacterial genomes encode for more biosynthetic gene clusters (BGCs) than the identified compounds so far, possibly because some are cryptic or silent under standard laboratory conditions. Given that secondary metabolites can be produced as a response to environmental stress conditions, employing the OSMaC (One Strain, Many Compounds) strategy might help unlocking the full biosynthetic potential of these microorganisms. This approach underlies that, in different culture conditions, a single strain can produce different molecules. CIIMAR harbors an important cyanobacterial culture collection (LEGE-CC) that has been investigated for the discovery of new bioactive compounds. Previous bioactivity-guided studies discovered that strains Lusitaniella coriacea LEGE 01767 and Leptothoe sp. LEGE 181152 produce different groups of compounds whose structure indicates a possible role as metal chelating agents. Additionally, the genome of Leptothoe sp. LEGE 181152 contains a BGC whose product has not been found. The main goal of this work was to use OSMaC on two strains from the LEGE-CC Culture Collection (CIIMAR) to (i) study the production of lusichelins (1-5) on Lusitaniella coriacea LEGE 01767 (ii) modulate the secondary metabolism Leptothoe sp. LEGE 181152 to induce expression of cryptic/silent BGCs in. To achieve this, a workflow was followed involving genome mining, cultivation under different culture conditions and metabolomic analyses by mass spectrometry. Lusitaniella coriacea LEGE 01767 was cultivated under standard (Z8-TM), iron-limited and iron depleted conditions, as well as with 0, 5, 200 and 450 nM copper sulfate, to evaluate their effect on production of lusichelins 1-5. Given the initial hypothesis that lusichelins are involved in iron chelation, an increase of their presence in iron-limited and iron-depleted conditions was expected. However, it did not occur, and peak production was detected on standard conditions. This points to iron chelation not being their main biological purpose. The conditions with 200 and 450 nM of copper sulfate seemed to cause some toxicity to the cyanobacterium. In these, a potential new compound with the same exact mass of lusichelin 5 was identified. Leptothoe sp. LEGE 181152 was cultivated under different light conditions: white, blue, and red light in 16/8h light/dark cycle and 24h of continuous light. In these conditions, major modulation of the metabolome was verified, with blue and red light having a more pronounced effect than the white light. Moreover, continuous light stimulated more changes in the metabolome than the 16/8h light/dark cycle. A new group of halogenated compounds, predominant in blue light conditions, was discovered. The extracts from these different experimental conditions did not reveal anticancer or antibacterial activity. However, extracts from the red light conditions presented anti-settlement activity in mussel larvae (Mytilus galloprovincialis). Ultimately, the manipulation of the culture conditions in these two strains caused alterations in their biosynthetic capabilities, which reflected in their bioactive potential and allowed the identification of potential new compounds.