A candidate gene cluster for the bioactive natural product gyrophoric acid in lichen-forming fungi

Natural products of lichen-forming fungi are structurally diverse and have a variety of medicinal properties. Yet they a have limited implementation in industry as for most of the natural products, the corresponding genes remain unknown. Here we implement a long-read sequencing and bioinformatic approach to identify the biosynthetic gene cluster of the bioactive natural product gyrophoric acid (GA). Using 15 high-quality genomes representing nine GA-producing species of the lichen-forming fungal genus Umbilicaria, we identify the most likely GA cluster and investigate cluster gene organization and composition across the nine species. Our results show that GA clusters are promiscuous within Umbilicaria with only three genes that are conserved across species, including the PKS gene. In addition, our results suggest that the same cluster codes for different but structurally similar NPs, i.e., GA, umbilicaric acid and hiascic acid, bringing new evidence that lichen metabolite diversity is also generated through regulatory mechanisms at the molecular level. Ours is the first study to identify the most likely GA cluster. This information is essential for opening up avenues for biotechnological approaches to producing and modifying GA, and possibly other lichen compounds. We show that bioinformatics approaches are useful in linking genes and potentially associated natural products. Genome analyses help unlocking the pharmaceutical potential of organisms such as lichens, which are biosynthetically diverse, but slow growing, and usually uncultivable due to their symbiotic nature.

Intra-specific variation in lichen secondary compounds across environmental gradients on Signy Island, maritime Antarctic

Lichens produce various carbon-based secondary compounds (CBSCs) in response to abiotic conditions and herbivory. Although lichen CBSCs have received considerable attention with regard to responses to UV-B exposure, very little is known about intra-specific variation across environmental gradients and their role in protection against herbivory in the Antarctic. Here we report on the variation in CBSCs of two widely distributed and common Antarctic lichens, Usnea antarctica and Umbilicaria antarctica, between sites with different solar exposure (NW–SE) and along natural nitrogen (N) gradients which are associated with changing lichen-invertebrate associations on Signy Island (South Orkney Islands, maritime Antarctic). Fumarprotocetraric and usnic acid concentrations in Usnea showed no relationships with solar exposure, lichen-N or associated invertebrate abundance. However, fumarprotocetraric acid concentration was 13 times higher at inland sites compared to coastal sites along the N-gradients. Gyrophoric acid concentration in Umbilicaria was 33% lower in sun-facing (northerly exposed) habitats compared to more shaded (south-facing) rocks and declined with elevation. Gyrophoric acid concentration was positively correlated with the abundance and species richness of associated microarthropods, similar to the patterns found with lichen N. This initial investigation indicates that there can be large intraspecific variation in lichen CBSC concentrations across relative short distances (< 500 m) on Signy Island and raises further questions regarding current understanding of the role of CBSCs in Antarctic lichens in relation to biotic and abiotic pressures.

Phenolic compounds from the lichen Parmotrema tinctorum

Introduction: The metabolites of lichens concentrated depsidones, depsides, and diphenyl ethers were possessed antibiotic, antifungal, antiviral, antitumor, and anticancer activities. Parmotrema tinctorum (Despr. ex Nyl.) Hale, a species of foliose lichen, is widely distributed in Lam Dong province, Vietnam. Herein, this paper describes the isolation and structure elucidation of seven compounds isolated from this lichen. Methods: Phytochemical investigations of the ethyl acetate extract of the lichen P. tinctorum led to the isolation of seven pure compounds. Their chemical structures were elucidated by extensive HR-ESI-MS and NMR spectroscopic analysis and comparison with previously published data. Results: Seven compounds, namely orcinol (1), orsellinic acid (2), methyl orsellinate (3), methyl heamatomate (4), lecanorin (5), lecanoric acid (6), and gyrophoric acid (7). These compounds were determined the α-glucosidase inhibitory activity. Conclusions: Compound 7 was determined for the first time in P. tinctorum, and this was also the first time these compounds were determined the α-glucosidase inhibitory activity.

Trapelia calyciformis sp. nov. from China

Trapelia calyciformis is described from China as a new species. The lichen is characterized by its gray-white thallus, zeorine apothecium with black disc and cracked excipulum, narrow paraphyses branched near the tip, 8-spored asci, aseptate ascospores with one vacuole, and the presence of gyrophoric acid. The specimens examined were deposited in LCUF. A key to the species of Trapelia reported in China is presented.

Cryptothecia aleurodes (Arthoniaceae), a misunderstood species

Cryptothecia aleurodes was considered to be a widespread, rare tropical lichen having white ascigerous areas and a crustose whitish-grey thallus containing norstictic acid. A revision of its type specimen from Guadeloupe and the study of recent specimens from Martinique proved that the species has been misunderstood. In this paper, Cryptothecia aleurodes is shown to have a K– and C+ red thallus containing notably gyrophoric acid as major secondary metabolite but lacking norstictic acid. A detailed description and illustrations are provided. The species is known with certainty only from the Caribbean and has probably a Neotropical distribution. Previous reports of C. aleurodes from the Seychelles and Thailand are shown to be misidentifications and reports from India are considered dubious.

Anti-Aging Effects of Gyrophoric Acid on UVA-Irradiated Normal Human Dermal Fibroblasts

This research was conducted to identify the anti-aging effects of gyrophoric acid on the skin, using normal human dermal fibroblasts. The anti-aging effects of gyrophoric acid on dermal fibroblasts were demonstrated through cell viability, verification of collagen, type I, alpha 1 (COL1A1)/COL3A1/matrix metalloproteinases 1 (MMP1) messenger ribonucleic acid (mRNA) expression levels with quantitative real-time reverse-transcription polymerase chain reaction, and protein estimation using type I collagen/MMP1-enzyme-linked immunosorbent assay. Further, the effects of gyrophoric acid on superoxide dismutases (SODs)/catalase were investigated by assessing their mRNA expression. In ultraviolet A (UVA)-treated dermal fibroblasts, gyrophoric acid was observed to increase mRNA levels of COL1A1/COL3A1/SOD2 genes and type I collagen protein levels, consistent with its anti-aging role. Furthermore, gyrophoric acid treatment decreased both MMP1 mRNA and protein expression levels. Therefore, the results of this study demonstrate that gyrophoric acid can be considered as an important natural compound with potent anti-aging effects on the skin. Based on the findings of this study, further research about the mechanism of action of gyrophoric acid should be pursued so as to develop novel anti-aging strategies not only in the field of cosmetics but also for healthcare.

PRIMARY AND SECONDARY METABOLITES LASALLIA PENSYLVANICA

Gyrophoric acid was isolated from thalluses of Lasallia pensylvanica and identification using IR, UV, MS spectrometry and differential scanning calorimetry. It was established that the content of gyrophoric acid in the lichen was 36 mg/g. It was shown that the gyrophoric acid exhibited antioxidant activity relative to the superoxide anion, NO, DPPH and ABTS radicals, inhibited the lipid peroxidation and reduced Fe (II) ions. At the same time, the antiradical activity of the gyrophoric acid relative to the superoxide anion and nitric oxide was higher by 1.3 and 1.6 times, respectively, compared with taxifolin. Neutral lipids, glycolipids, phospholipids and a fraction of water-soluble polysaccharides were isolated from the thalluses of Lasallia pensylvanica. It was shown that the main lipid fractions are neutral and phospholipids, the content of which were 84.25 and 73.25 mg/g, respectively. The yield of the fraction of water-soluble polysaccharides from the thalluses of Lasallia pensylvanica was 45.98%. It is established that the main monomers of the water-soluble fraction of the polysaccharide Lasallia pensylvanica are glucose and mannose. The study of the composition and content of low molecular weight compounds in the thalluses of Lasallia pensylvanica, performing the function of cryo- and osmoprotectants. Found compounds such as hydroxyproline, glutamine, arabitol, ribitol, mannitol, sucrose, trehalose.

Discovery of Novel Multi-target Inhibitor of angiotensin type 1 receptor and neprilysin inhibitors from Traditional Chinese Medicine

Angiotensin II type-1 receptor–neprilysin inhibitor (ARNi) is consisted of Angiotensin II type-1 receptor (AT1) antagonist and neprilysin (NEP) inhibitor, which could simultaneously increase the vasodilators of the natriuretic peptides and antagonize vasoconstrictors of Ang II. ARNi has been proved a superior effect and lower risks of death on chronic heart failure (CHF) and hypertension. In this paper, ARNi from Traditional Chinese Medicines (TCM) was discovered based on target combination of AT1 and NEP by virtual screening, biological assay and molecular dynamics (MD) simulations. Two customized strategies of combinatorial virtual screening were implemented to discover AT1 antagonist and NEP inhibitor based on pharmacophore modeling and docking computation respectively. Gyrophoric acid (PubChem CID: 135728) from Parmelia saxatilis was selected as AT1 antagonist and assayed with IC50 of 29.76 μM by calcium influx assay. And 3,5,3′-triiodothyronine (PubChem CID: 861) from Bos taurus domesticus was screened as NEP inhibitor and has a dose dependent inhibitory activity by biochemistry fluorescence assay. Combined with MD simulations, these compounds can generate interaction with the target, key interactive residues of ARG167, TRP84, and VAL108 in AT1, and HIS711 in NEP were also identified respectively. This study designs the combinatorial strategy to discover novel frames of ARNi from TCM, and gyrophoric acid and 3,5,3′-triiodothyronine could provide the clues and revelations of drug design and therapeutic method of CHF and hypertension for TCM clinical applications.

Studies on Micarea in Australasia II. A synopsis of the genus in Tasmania, with the description of ten new species

Thirty-five species of Micarea are recorded for Tasmania. Ten are described as new to science: M. ceracea Coppins & Kantvilas (also known from Victoria and New South Wales), characterized by a thallus containing perlatolic and didymic acids, pallid apothecia and 3(–4)-septate ascospores, 10–21 × 3·5–6 µm; M. cinereopallida Coppins & Kantvilas (also known from Chile), with a granular to coralloid, goniocyst-like thallus containing superlatolic acid, pallid to piebald apothecia and (0–)1-septate ascospores, 8–15 × 2·5–5 µm; M. micromelaena Kantvilas & Coppins, similar to the widespread M. melaena but with markedly smaller, 0–1-septate ascospores, 8–12·5 × 2·5–4 µm; M. oreina Kantvilas & Coppins, characterized by a thallus of globose areoles containing gyrophoric acid, black, subglobose apothecia, and 1-septate ascospores, 11–16·5 × 4·5–6·5 µm; M. pallida Coppins & Kantvilas, similar to M. ceracea but distinguished by the presence of porphyrilic acid and relatively small, 3-septate ascospores, 9·5–15 × 2·5–4 µm; M. prasinastra Coppins & Kantvilas (also known from New Zealand), a member of the M. prasina group with a finely granular-sorediose thallus containing gyrophoric acid, unpigmented apothecia and (0–)1-septate ascospores, 7–11·5 × 1·8–3·5 µm; M. rubiginosa Coppins & Kantvilas (also known from Chile), likewise allied to M. prasina but with apothecia containing Rubella-orange pigment and ascospores 0–1-septate, 9·5–17 × 3·5–5·5 µm; M. sandyana Kantvilas, related to M. ternaria (Nyl.) Vĕzda but differing by smaller ascospores, 7–13·5 × 3·5–6 µm; M. saxicola Coppins & Kantvilas, characterized by a relatively thick, grey-brown, areolate thallus, convex, black apothecia and 0(–1)-septate ascospores, 7–18 × 4·5–7 µm; and M. tubaeformis Coppins & Kantvilas, related to M. flagellispora and with filiform ascospores, 45–100 × 1–2 µm, but differing by containing 2′-O-methylperlatolic acid and having funnel-shaped pycnidia. Ten species of Micarea are reported for Tasmania for the first time: M. almbornii Coppins, M. argopsinosa P. M. McCarthy & Elix, M. byssacea (Th. Fr.) Czarnota et al., M. contexta Hedl., M. farinosa Coppins & Aptroot, M. humilis P. M. McCarthy & Elix, M. incrassata Hedl., M. myriocarpa V. Wirth & Vězda ex Coppins, M. nowakii Czarnota & Coppins and M. pseudocoppinsii Brand et al. Also recorded for the first time for Victoria are M. alabastrites (Nyl.) Coppins and M. cinerea (Schaer.) Hedl. A key to Micarea-like lichens in Tasmania, which includes Micarea itself as well as Brianaria, Psilolechia and Leimonis, is presented. Leimonis erratica (Körb.) R. C. Harris & Lendemer and Brianaria tuberculata (Sommerf.) S. Ekman & M. Svensson are recorded for Tasmania for the first time.