A new path for terpenoid biosynthesis.

Triterpenes are C30 organic compounds abundantly found in all living organisms. Although previously believed to be exclusively produced from squalene or oxidosqualene, a recent report by Tao and colleagues describes a new triterpene biosynthetic route involving the cyclization of the precursor hexaprenyl diphosphate (HexPP) by unprecedented bifunctional terpene synthase (TS) enzymes.

Convergent evolution for antibiotic biosynthesis in bacteria and animals.

Convergent evolution has been described for several metabolic pathways across the kingdoms of life. However, there is hitherto no evidence for such an interkingdom process for antimicrobials. A new report suggests that marine animals have evolved the ability to biosynthesize antimicrobial polyketides, in parallel with bacteria.

Mycobiota dysbiosis: a new nexus in intestinal tumorigenesis.

While there is growing evidence that perturbation of the gut microbiota can result in a variety of pathologies including gut tumorigenesis, the influence of commensal fungi remains less clear. In this issue, Zhu et al (2021) show that mycobiota dysbiosis stimulates energy metabolism changes in subepithelial macrophages promoting colon cancer via enhancing innate lymphoid cell activity. These findings provide insights into a role of the gut flora in intestinal carcinogenesis and suggest opportunities for adjunctive antifungal or immunotherapeutic strategies to prevent colorectal cancer.

Fungal Melanin Rewires Macrophage Metabolism.

Aspergillus fumigatus is a deadly fungal pathogen in immunocompromised patients. A report by Gonçalves et al. reveals that melanin, a secondary metabolite present at the surface of infecting fungal spores, induces glycolysis in macrophages to promote inflammatory responses. This opens a window for the development of innovative host-directed antifungal therapies.

Plant to Insect Horizontal Gene Transfer: Empowering Whiteflies.

Horizontal gene transfer (HGT) is a well-documented evolutionary driving phenomenon in prokaryotes and eukaryotes, but its impact on the plant kingdom has remained elusive. A recent study provides compelling evidences, which support the idea that a plant-derived gene allows for the detoxification of plant defense metabolites in a polyphagous arthropod herbivore.

Are Histidine Kinases of Arbuscular Mycorrhizal Fungi Involved in the Response to Ethylene and Cytokinins?

Signals are exchanged at all stages of the arbuscular mycorrhizal (AM) symbiosis between fungi and their host plants. Root-exuded strigolactones are well-known early symbiotic cues, but the role of other phytohormones as interkingdom signals has seldom been investigated. Here we focus on ethylene and cytokinins, for which candidate receptors have been identified in the genome of the AM fungus Rhizophagus irregularis. Ethylene is known from the literature to affect asymbiotic development of AM fungi, and in the present study, we found that three cytokinin forms could stimulate spore germination in R. irregularis. Heterologous complementation of a Saccharomyces cerevisiae mutant strain with the candidate ethylene receptor RiHHK6 suggested that this protein can sense and transduce an ethylene signal. Accordingly, its N-terminal domain expressed in Pichia pastoris displayed saturable binding to radiolabeled ethylene. Thus, RiHHK6 displays the expected characteristics of an ethylene receptor. In contrast, the candidate cytokinin receptor RiHHK7 did not complement the S. cerevisiae mutant strain or Medicago truncatula cytokinin receptor mutants and seemed unable to bind cytokinins, suggesting that another receptor is involved in the perception of these phytohormones. Taken together, our results support the hypothesis that AM fungi respond to a range of phytohormones and that these compounds bear multiple functions in the rhizosphere beyond their known roles as internal plant developmental regulators. Our analysis of two phytohormone receptor candidates also sheds new light on the possible perception mechanisms in AM fungi. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The Evolutionary Pattern of Cocaine and Hyoscyamine Biosynthesis Provides Strategies To Produce Tropane Alkaloids.

Cocaine and hyoscyamine are two tropane alkaloids (TA) from Erythroxylaceae and Solanaceae, respectively. These famous compounds possess anticholinergic properties that can be used to treat neuromuscular disorders. While the hyoscyamine biosynthetic pathway has been fully elucidated allowing its de novo synthesis in yeast, the cocaine pathway remained only partially elucidated. Recently, the Huang research group has completed the cocaine biosynthetic route by characterizing its two missing enzymes. This allowed the whole pathway to be transferring into Nicotiana benthamiana to achieve cocaine production. Here, besides highlighting the impact of this discovery, we discuss how TA biosynthesis evolved via the recruitment of two distinct and convergent pathways in Erythroxylaceae and Solanaceae. Finally, while enriching our knowledge on TA biosynthesis, this diversification of the molecular actors involved in cocaine and hyoscyamine biosynthesis opens perspectives in metabolic engineering by exploring enzyme biochemical plasticity that can ease and shorten TA pathway reconstitution in heterologous organisms.

Functional insights into human macrophage response against Scedosporium apiospermum and Scedosporium dehoogii.

Fungal infections caused by Scedosporium species are rising among immunocompromised and immunocompetent patients. Within the immunocompetent group, patients with cystic fibrosis (pwCF) are at high risk of developing a chronic airway colonization by these molds. While S. apiospermum is one of the major species encountered in the lungs of pwCF, S. dehoogii has rarely been reported. The innate immune response is believed to be critical for host defense against fungal infections. However, its role has only recently been elucidated and the immune mechanisms against Scedosporium species are currently unknown. In this context, we undertook a comparative investigation of macrophage-mediated immune responses toward S. apiospermum and S. dehoogii conidia. Our data showed that S. apiospermum and S. dehoogii conidia strongly stimulated the expression of a set of pro-inflammatory cytokines and chemokines such as IL-1ß, IL-8, IL-6 and TNFα. We demonstrated that S. dehoogii was more potent in stimulating the early release of pro-inflammatory cytokines and chemokines while S. apiospermum induced a late inflammatory response at a higher level. Flow cytometry analysis showed that M1-like macrophages were able to internalize both S. apiospermum and S. dehoogii conidia, with a similar intracellular killing rate for both species. In conclusion, these results suggest that M1-like macrophages can rapidly initiate a strong immune response against both S. apiospermum and S. dehoogii. This response is characterized by a similar killing of internalized conidia, but a different time course of cytokine production.

Split-marker-mediated genome editing improves homologous recombination frequency in the CTG clade yeast Candida intermedia.

Genome-editing toolboxes are essential for the exploration and exploitation of nonconventional yeast species as cell factories, as they facilitate both genome studies and metabolic engineering. The nonconventional yeast Candida intermedia is a biotechnologically interesting species due to its capacity to convert a wide range of carbon sources, including xylose and lactose found in forestry and dairy industry waste and side-streams, into added-value products. However, possibilities of genetic manipulation have so far been limited due to lack of molecular tools for this species. We describe here the development of a genome editing method for C. intermedia, based on electroporation and gene deletion cassettes containing the Candida albicans NAT1 dominant selection marker flanked by 1000 base pair sequences homologous to the target loci. Linear deletion cassettes targeting the ADE2 gene originally resulted in <1% targeting efficiencies, suggesting that C. intermedia mainly uses nonhomologous end joining for integration of foreign DNA fragments. By developing a split-marker based deletion technique for C. intermedia, we successfully improved the homologous recombination rates, achieving targeting efficiencies up to 70%. For marker-less deletions, we also employed the split-marker cassette in combination with a recombinase system, which enabled the construction of double deletion mutants via marker recycling. Overall, the split-marker technique proved to be a quick and reliable method for generating gene deletions in C. intermedia, which opens the possibility to uncover and enhance its cell factory potential.

Oxidative stress response pathways in fungi.

Fungal response to any stress is intricate, specific, and multilayered, though it employs only a few evolutionarily conserved regulators. This comes with the assumption that one regulator operates more than one stress-specific response. Although the assumption holds true, the current understanding of molecular mechanisms that drive response specificity and adequacy remains rudimentary. Deciphering the response of fungi to oxidative stress may help fill those knowledge gaps since it is one of the most encountered stress types in any kind of fungal niche. Data have been accumulating on the roles of the HOG pathway and Yap1- and Skn7-related pathways in mounting distinct and robust responses in fungi upon exposure to oxidative stress. Herein, we review recent and most relevant studies reporting the contribution of each of these pathways in response to oxidative stress in pathogenic and opportunistic fungi after giving a paralleled overview in two divergent models, the budding and fission yeasts. With the concept of stress-specific response and the importance of reactive oxygen species in fungal development, we first present a preface on the expanding domain of redox biology and oxidative stress.

Tonoplast and Peroxisome Targeting of γ-tocopherol N-methyltransferase Homologs Involved in the Synthesis of Monoterpene Indole Alkaloids.

Many plant species from the Apocynaceae, Loganiaceae and Rubiaceae families evolved a specialized metabolism leading to the synthesis of a broad palette of monoterpene indole alkaloids (MIAs). These compounds are believed to constitute a cornerstone of the plant chemical arsenal but above all several MIAs display pharmacological properties that have been exploited for decades by humans to treat various diseases. It is established that MIAs are produced in planta due to complex biosynthetic pathways engaging a multitude of specialized enzymes but also a complex tissue and subcellular organization. In this context, N-methyltransferases (NMTs) represent an important family of enzymes indispensable for MIA biosynthesis but their characterization has always remained challenging. In particular, little is known about the subcellular localization of NMTs in MIA-producing plants. Here, we performed an extensive analysis on the subcellular localization of NMTs from four distinct medicinal plants but also experimentally validated that two putative NMTs from Catharanthus roseus exhibit NMT activity. Apart from providing unprecedented data regarding the targeting of these enzymes in planta, our results point out an additional layer of complexity to the subcellular organization of the MIA biosynthetic pathway by introducing tonoplast and peroxisome as new actors of the final steps of MIA biosynthesis.

Highly Conserved gsc1 Gene of Pneumocystis jirovecii in Patients with or without Prior Exposure to Echinocandins.

Echinocandins are noncompetitive inhibitors of the GSC1 subunit of the enzymatic complex involved in synthesis of 1,3-beta-d-glucan, a cell wall component of most fungi, including Pneumocystis spp. Echinocandins are widely used for treating systemic candidiasis and rarely used for treating Pneumocystis pneumonia. Consequently, data on P. jirovecii gsc1 gene diversity are still scarce compared to that for the homologous fks1 gene of Candida spp. In this study, we analyzed P. jirovecii gsc1 gene diversity and the putative selection pressure of echinocandins on P. jirovecii. gsc1 gene sequences of P. jirovecii specimens from two patient groups were compared. One group of 27 patients had prior exposure to echinocandins, whereas the second group of 24 patients did not, at the time of P. jirovecii infection diagnoses. Two portions of the P. jirovecii gsc1 gene, HS1 and HS2, homologous to hot spots described in Candida spp., were sequenced. Three single-nucleotide polymorphisms (SNPs) at positions 2204, 2243, and 2303 close to the HS1 region and another SNP at position 4540 more distant from the HS2 region were identified. These SNPs represent synonymous mutations. Three gsc1 HS1 alleles, A, B, and C, and two gsc1 HS2 alleles, a and b, and four haplotypes, Ca, Cb, Aa, and Ba, were defined, without significant difference in haplotype distribution in both patient groups (P = 0.57). Considering the identical diversity of P. jirovecii gsc1 gene and the detection of synonymous mutations in both patient groups, no selection pressure of echinocandins among P. jirovecii microorganisms can be pointed out so far.

Towards a Better Understanding of Toxin Biosynthesis in Seaweeds.

Harmful algal blooms (HABs) represent both ecological and public health hazards in the marine environment. Indeed, some algae can produce metabolites that have negative effects on marine ecosystems and mammals. Kainoid derivatives such as kainic acid (KA) and domoic acid (DA) are considered some of the most toxic metabolites of marine origin biosynthesized by a limited number of micro- and macroalgae. While recent works have provided the first insights into the biosynthetic route of KA in red algae and DA in diatoms, the DA biosynthetic pathway has remained uncharacterized for red algae. In a recent work, the research groups of Chekan and Moore have not only elucidated the biosynthetic pathway of DA in the red alga Chondria armata but also shed light on its complex evolution among marine species. We discuss here the importance of pursuing active research in this area to gain insights into secondary biosynthetic pathways in marine organisms for diagnostic and metabolic engineering perspectives.

The high osmolarity glycerol (HOG) pathway in fungi†.

While fungi are widely occupying nature, many species are responsible for devastating mycosis in humans. Such niche diversity explains how quick fungal adaptation is necessary to endow the capacity of withstanding fluctuating environments and to cope with host-imposed conditions. Among all the molecular mechanisms evolved by fungi, the most studied one is the activation of the phosphorelay signalling pathways, of which the high osmolarity glycerol (HOG) pathway constitutes one of the key molecular apparatus underpinning fungal adaptation and virulence. In this review, we summarize the seminal knowledge of the HOG pathway with its more recent developments. We specifically described the HOG-mediated stress adaptation, with a particular focus on osmotic and oxidative stress, and point out some lags in our understanding of its involvement in the virulence of pathogenic species including, the medically important fungi Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus, compared to the model yeast Saccharomyces cerevisiae. Finally, we also highlighted some possible applications of the HOG pathway modifications to improve the fungal-based production of natural products in the industry.

Alternative splicing creates a pseudo-strictosidine ß-d-glucosidase modulating alkaloid synthesis in Catharanthus roseus.

Deglycosylation is a key step in the activation of specialized metabolites involved in plant defense mechanisms. This reaction is notably catalyzed by ß-glucosidases of the glycosyl hydrolase 1 (GH1) family such as strictosidine ß-d-glucosidase (SGD) from Catharanthus roseus. SGD catalyzes the deglycosylation of strictosidine, forming a highly reactive aglycone involved in the synthesis of cytotoxic monoterpene indole alkaloids (MIAs) and in the crosslinking of aggressor proteins. By exploring C. roseus transcriptomic resources, we identified an alternative splicing event of the SGD gene leading to the formation of a shorter isoform of this enzyme (shSGD) that lacks the last 71-residues and whose transcript ratio with SGD ranges from 1.7% up to 42.8%, depending on organs and conditions. Whereas it completely lacks ß-glucosidase activity, shSGD interacts with SGD and causes the disruption of SGD multimers. Such disorganization drastically inhibits SGD activity and impacts downstream MIA synthesis. In addition, shSGD disrupts the metabolic channeling of downstream biosynthetic steps by hampering the recruitment of tetrahydroalstonine synthase in cell nuclei. shSGD thus corresponds to a pseudo-enzyme acting as a regulator of MIA biosynthesis. These data shed light on a peculiar control mechanism of ß-glucosidase multimerization, an organization common to many defensive GH1 members.

Smoking-aggravated oral candidiasis: Nrf2 pathway dampens NLRP3 inflammasome.

While cigarette smoke compounds are known to have immunosuppressive effects on the oral mucosa, the relationship between in vivo immune dysfunction caused by smoking and the development of oral Candida infections remains largely unexplored. In a recent issue of The Journal of Cellular and Molecular Medicine, Ye and colleagues provide evidence that smoking increases oral mucosa susceptibility to Candida albicans infection via the activation of the Nrf2 pathway, which in turn negatively regulates the NLRP3 inflammasome. This opens new perspective in considering Nrf2 as a relevant target for smoking-induced C. albicans-related oral diseases.

Metabolic engineering for plant natural products biosynthesis: new procedures, concrete achievements and remaining limits.

Microorganisms and plants represent major sources of natural compounds with a plethora of bioactive properties. Among these, plant natural products (PNPs) remain indispensable to human health. With few exceptions, PNP-based pharmaceuticals come from plant specialized metabolisms and display a structure far too complex for a profitable production by total chemical synthesis. Accordingly, their industrial processes of supply are still mostly based on the extraction of final products or precursors directly from plant materials. This implies that particular contexts (e.g. pandemics, climate changes) and natural resource overexploitation are main drivers for the high production cost and recurrent supply shortages. Recently, biotechnological manufacturing alternatives gave rise to a multitude of benchmark studies implementing the production of important PNPs in various heterologous hosts. Here, we spotlight unprecedented advancements in the field of metabolic engineering dedicated to the heterologous production of a prominent series of PNPs that were achieved during the year 2020. We also discuss how the knowledge accumulated in recent years could pave the way for a broader manufacturing palette of natural products from a wide range of natural resources.

Engineered Microbes for Producing Anticholinergics.

The tropane alkaloids (TAs) hyoscyamine and scopolamine function as acetylcholine receptor antagonists and are used clinically as parasympatholytics to treat neuromuscular disorders in humans. While TAs are synthesized in a small subset of plant families, these specialized metabolites are only accumulated in limited quantities in plant organs. The complex chemical structures of these compounds make their industrial production by chemical synthesis very challenging, Therefore, the supply of these TAs still relies on intensive farming of Duboisia shrubs in tropical countries. Many adverse factors such as climate fluctuations and pandemics can thus influence annual world production. Based on the landmark microbial production of the antimalarial semi-synthetic artemisinin, the Smolke group recently developed a yeast cell factory capable of de novo synthesizing hyoscyamine and scopolamine, thus paving the way for an alternative production of these compounds.