The role of beta-carotene derivatives in speciation and communication in zygomycete fungi

References
Schultze K, Schimek C, Wöstemeyer J, Burmester A (2005) Sexuality and parasitism share common regulatory pathways in the fungus Parasitella parasitica. Gene 348, 33-44.
Parasitella parasitica, a facultative mycoparasite of zygomycetous fungi, forms cytoplasmic fusions with its hosts during infection. Thus, the organism is an efficient donor of genetic material in parasexual host-parasite interactions. Recognition between parasite and host is mediated by trisporoids, which are also responsible for sexual communication. The TDH gene for one of the key enzymes of trisporic acid biosynthesis, 4-dihydromethyl-trisporate dehydrogenase, was cloned and its transcription analysed. TDH was cloned on a 6175 bp insert and was found to map in a complex cluster of genes that suggest post-transcriptional antisense regulation. Histochemical TDH analysis in developing parasitic or sexual structures shows high enzymatic activity in Parasitella. TDH is linked to a gene for a putative acyl-CoA thioesterase (ACT). Two ORFs were identified in the 5'-region of the TDH gene, a third one, coding for 176 amino acids overlaps the ACT gene in antisense direction completely. Expression levels of ACT and ORF1 depend on parasitic and sexual interactions.

Schachtschabel D, Schimek C, Wöstemeyer J, Boland W (2005) Biological activity of trisporoids and trisporoid analogues in Mucor mucedo (-). Phytochemistry 66, 1358-1365.
In the course of their sexual interactions, zygomycete fungi communicate via an elaborate series of carotene-derived compounds, namely trisporic acid and its biosynthetic progenitors. A novel building-block strategy allowed the systematic generation of structurally modified trisporoids along with putative early biosynthetic precursors for physiological tests. The impact of discrete structural elements was documented by the ability of individual compounds to induce sexually committed hyphae in Mucor mucedo. The activity screening contributed to establish general structure-functional relationships for trisporoid action. Most crucial for activity were the dimension of the longer side chain, the polarity of functional groups at C(4) and C(13), and the number of conjugated double bonds in the side chain. The presence of an oxygen substituent at the cyclohexene ring is not essential for function. The overall biological activity apparently results from the combination of the various structural elements.

Schimek C, Petzold A, Schultze K, Wetzel J, Wolschendorf F, Burmester A, Wöstemeyer J (2005) 4-Dihydromethyltrisporate dehydrogenase, an enzmye of the sex hormone pathway in Mucor mucedo, is constitutively transcribed but its activity is differently regulated in (+) and (-) mating types. Fungal Genet Biol 42, 804-812.
4-dihydromethyltrisporate dehydrogenase (TDH) converts the (+) mating type sex pheromone 4-dihydromethyltrisporate into methyltrisporate. In Mucor mucedo, this conversion is required only in the (-) mating type. Gene expression of the TDH encoding TSP1 gene was analyzed qualitatively using reverse transcribed-PCR. TSP1 is constitutively transcribed in the (+) and in the (-) mating type, irrespective of the mating situation. By immunodetection, the translation product is also formed constitutively. Contrarily to gene expression, TDH enzyme activity depends on the sexual status of the mycelium. Activity is restricted to the sexually stimulated (-) mating type. Non-stimulated (-)-, as well as stimulated and non-stimulated (+)-mycelia exhibit no activity and do not influence activity in stimulated (-) mycelia. Time course analysis shows strongly increased enzyme activity at 80 minutes after stimulation. Low activity exists from the onset of stimulation, indicating that additional regulation mechanisms are involved in TDH function.

Schachtschabel D, Boland W (2007) Efficient generation of a trisporoid library by combination of synthesis and biotransformation. J Org Chem 72, 1366-1372.
Trisporic acids and their biosynthetic precursors represent a family of powerful fungal pheromones and morphogenetic factors. A highly flexible synthetic protocol is described that (i) provides rapid access to nonfunctionalized early trisporoids from b-ionone, (ii) includes a regiospecific oxidative functionalization of b-ionone leading to 1-acetoxy-ß-ionone giving access to functionalized trisporoids, and (iii) utilizes a biotransformation of early synthetic trisporoids by growing cells of Blakeslea trispora to prepare late trisporoids including trisporic acids. The same protocol also provides deuterium-labeled trisporoids such as trisporin B [²H₃]-19. Administration of [²H₃]-19 to growing cells of the (-)-mating type of B. trispora resulted in the formation of the labeled trisporols [²H₃]-20 and [²H₃]-21. Growing cultures containing both mating types can be used to prepare trisporic acids from early precursors.

Schimek C, Kleppe K, Saleem A-R, Voigt K, Burmester A, Wöstemeyer J (2003) Sexual reactions in Mortierellales are mediated by the trisporic acid system. Mycol Res 107, 736-747.
Several species of the genus Mortierella (Mortierellales, Zygomycota) were examined for substances regulating their sexual reactions. Compounds isolated from both mated and single growing Mortierella strains were purified by thin layer chromatography. Some of these compounds showed UV absorbance-characteristics similar to those of trisporoids, a group of compounds involved in sexual regulation in Mucorales. A compound with a 4-dihydromethyltrisporate-like absorbance spectrum was detected. To test for the interspecific sexual responses typically induced by trisporoids, the compounds extracted from Mortierella spp. were tested against the Mucorales Mucor mucedo and Phycomyces blakesleeanus and were found to induce sexual reactions in both tester strains. A gene encoding 4-dihydromethyltrisporate dehydrogenase was identified in several Mortierella species and the activity of the gene product was shown using a histochemical assay. We propose that the regulation of sexual processes by trisporoids is common to both Mucorales and Mortierellales and may be more widespread within the Zygomycota.

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