Polymorpha Genome Research Articles

Hansenula polymorpha methanol metabolism genes enhance recombinant protein production in Komagataella phaffi

Recombinant protein production in Komagataella phaffi (K. phaffi), a widely utilized host organism, can be optimized by enhancing the metabolic flux in the central carbon metabolism pathways. The methanol utilization pathway (MUT) during methanol-based growth plays a crucial role in providing precursors and energy for cell growth and development. This study investigated the impact of boosting the methanol dissimilation pathway, a branch of MUT that plays a vital role in detoxifying formaldehyde and providing energy in the form of NADH, in K. phaffi. This was achieved by integrating two orthologous genes from Hansenula polymorpha into the K. phaffi genome: formaldehyde dehydrogenase (HpFLD) and formate dehydrogenase (HpFMDH). The HpFLD and HpFMDH genes were isolated from the Hansenula polymorpha genome and inserted under the regulation of the pAOX1 promoter in the genome of recombinant K. phaffi that already contained a single copy of model protein genes (eGFP or EGII). The expression levels of these model proteins were assessed through protein activity assays and gene expression analysis. The findings revealed that while both orthologous genes positively influenced model protein production, HpFMDH exhibited a more pronounced upregulation in expression compared to HpFLD. Co-expression of both orthologous genes demonstrated synergistic effects, resulting in approximately a twofold increase in the levels of the model proteins detected. This study provides valuable insights into enhancing the production capacity of recombinant proteins in K. phaffi.Graphical abstract

The D-mannose/L-galactose pathway plays a predominant role in ascorbate biosynthesis in the liverwort Marchantia polymorpha but is not regulated by light and oxidative stress.

Ascorbate plays an indispensable role in plants, functioning as both an antioxidant and a cellular redox buffer. It is widely acknowledged that the ascorbate biosynthesis in the photosynthetic tissues of land plants is governed by light-mediated regulation of the D-mannose/L-galactose (D-Man/L-Gal) pathway. At the core of this light-dependent regulation lies the VTC2 gene, encoding the rate-limiting enzyme GDP-L-Gal phosphorylase. The VTC2 expression is regulated by signals via the photosynthetic electron transport system. In this study, we directed our attention to the liverwort Marchantia polymorpha, representing one of the basal land plants, enabling us to conduct an in-depth analysis of its ascorbate biosynthesis. The M. polymorpha genome harbors a solitary gene for each enzyme involved in the D-Man/L-Gal pathway, including VTC2, along with three lactonase orthologs, which may be involved in the alternative ascorbate biosynthesis pathway. Through supplementation experiments with potential precursors, we observed that only L-Gal exhibited effectiveness in ascorbate biosynthesis. Furthermore, the generation of VTC2-deficient mutants through genome editing unveiled the inability of thallus regeneration in the absence of L-Gal supplementation, thereby revealing the importance of the D-Man/L-Gal pathway in ascorbate biosynthesis within M.polymorpha. Interestingly, gene expression analyses unveiled a distinct characteristic of M. polymorpha, where none of the genes associated with the D-Man/L-Gal pathway, including VTC2, showed upregulation in response to light, unlike other known land plants. This study sheds light on the exceptional nature of M.polymorpha as a land plant that has evolved distinctive mechanisms concerning ascorbate biosynthesis and its regulation.

Population Genomics of the Facultatively Sexual Liverwort Marchantia polymorpha.

The population genomics of facultatively sexual organisms are understudied compared with their abundance across the tree of life. We explore patterns of genetic diversity in two subspecies of the facultatively sexual liverwort Marchantia polymorpha using samples from across Southern Ontario, Canada. Despite the ease with which M. polymorpha should be able to propagate asexually, we find no evidence of strictly clonal descent among our samples and little to no signal of isolation by distance. Patterns of identity-by-descent tract sharing further showed evidence of recent recombination and close relatedness between geographically distant isolates, suggesting long distance gene flow and at least a modest frequency of sexual reproduction. However, the M. polymorpha genome contains overall very low levels of nucleotide diversity and signs of inefficient selection evidenced by a relatively high fraction of segregating deleterious variants. We interpret these patterns as possible evidence of the action of linked selection and a small effective population size due to past generations of asexual propagation. Overall, the M. polymorpha genome harbors signals of a complex history of both sexual and asexual reproduction.

MpDWF5A-encoded sterol Δ7-reductase is essential for the normal growth and development of Marchantia polymorpha.

Sterols are the essential components of the eukaryotic cell membranes. However, studies on sterol biosynthesis in bryophytes are limited. This study analyzed the sterol profiles in the bryophyte model plant Marchantia polymorpha L. The thalli contained typical phytosterols such as campesterol, sitosterol, and stigmasterol. BLASTX analysis of the M. polymorpha genome against the Arabidopsis thaliana sterol biosynthetic genes confirmed the presence of all of the enzymes responsible for sterol biosynthesis in M. polymorpha. In this study, we focused on characterizing two genes, MpDWF5A and MpDWF5B, which showed high homology with A. thaliana DWF5, encoding Δ5,Δ7-sterol Δ7-reductase. Functional analysis using a yeast expression system revealed that MpDWF5A converted 7-dehydrocholesterol to cholesterol, indicating that MpDWF5A is a Δ5,Δ7-sterol Δ7-reductase. Mpdwf5a-knockout lines (Mpdwf5a-ko) were constructed using CRISPR/Cas9 mediated genome editing. GC-MS analysis of Mpdwf5a-ko revealed that phytosterols such as campesterol, sitosterol, and stigmasterol disappeared, and instead, the corresponding Δ7-type sterols accumulated. The thalli of Mpdwf5a-ko grew smaller than those of the wild type, and excessive formation of apical meristem in the thalli was observed. In addition, the gemma cups of the Mpdwf5a-ko were incomplete, and only a limited number of gemma formations were observed. Treatment with 1 µM of castesterone or 6-deoxocastasterone, a bioactive brassinosteroid, partly restored some of these abnormal phenotypes, but far from complete recovery. These results indicate that MpDWF5A is essential for the normal growth and development of M. polymorpha and suggest that the dwarfism caused by the MpDWF5A defect is due to the deficiency of typical phytosterols and, in part, a brassinosteroid-like compound derived from phytosterols.

Genome-Wide Identification and Expression Analysis of the SWEET Gene Family in Annual Alfalfa (Medicago polymorpha).

SWEET (Sugars will eventually be exported transporter) proteins are a group of sugar transporters that are involved in sugar efflux, phloem loading, reproductive development, plant senescence, and stress responses. In this study, 23 SWEET transporter members were identified in the Medicago polymorpha genome, heterogeneously distributed on seven chromosomes. These MpSWEET genes were divided into four subfamilies, which showed similar gene structure and motif composition within the same subfamily. Seventeen MpSWEET genes encode seven transmembrane helices (TMHs), and all MpSWEET proteins possess conserved membrane domains and putative serine phosphorylation sites. Four and three pairs of MpSWEET genes were predicted to be segmentally and tandemly duplicated, respectively, which may have contributed to their evolution of M. polymorpha. The results of microarray and RNA-Seq data showed that some MpSWEET genes were specifically expressed in disparate developmental stages (including seedling stage, early flowering stage, and late flowering stage) or tissues such as flower and large pod. Based on protein network interaction and expression patterns of MpSWEET genes, six MpSWEET genes were selected for further quantitative real-time PCR validation in different stress treatments. qRT-PCR results showed that MpSWEET05, MpSWEET07, MpSWEET12, MpSWEET15, and MpSWEET21 were significantly upregulated for at least two of the three abiotic stress treatments. These findings provide new insights into the complex transcriptional regulation of MpSWEET genes, which facilitates future research to elucidate the function of MpSWEET genes in M. polymorpha and other legume crops.

Genome-Wide Identification and Expression Analysis of the Dof Transcription Factor in Annual Alfalfa Medicago polymorpha.

The Dof transcription factor is a plant-specific transcription gene family that plays various biological functions in plant development and stress response. However, no relevant research has been conducted on Medicago polymorpha. Here, 36 MpDof genes were identified in the M. polymorpha genome and further divided into 10 groups based on the comparative phylogenetic analysis. The essential information of MpDof genes, such as chromosomal localization, gene structure, conserved motifs, and selective pressures were systematically analyzed. All 36 MpDof genes were predicted to contain more cis-acting elements related to hormone response. MpDof24 and MpDof25 were predicted to interact with MpDof11 and MpDof26 to involve in the photoperiod blooms process. The MpDof genes showed a diverse expression pattern in different tissues. Notably, MpDof29 and MpDof31 were specifically expressed in the large pod and root, respectively, suggesting their crucial role in the pod and root development. qRT-PCR analysis indicated that the expression levels of MpDof10, MpDof25, MpDof26, and MpDof29 were obviously up-regulated under drought, salt, and cold stress. Collectively, genome-wide identification, evolutionary, and expression analysis of the Dof transcription gene family in M. polymorpha will provide new information to further understand and utilize the function of these Dof genes in Medicago plants.

ROP signaling regulates spatial pattern of cell division and specification of meristem notch

The formation of cell polarity is essential for many developmental processes such as polar cell growth and spatial patterning of cell division. A plant-specific ROP (Rho-like GTPases from Plants) subfamily of conserved Rho GTPase plays a crucial role in the regulation of cell polarity. However, the functional study of ROPs in angiosperm is challenging because of their functional redundancy. The Marchantia polymorpha genome encodes a single ROP gene, MpROP, providing an excellent genetic system to study ROP-dependent signaling pathways. Mprop knockout mutants exhibited rhizoid growth defects, and MpROP was localized at the tip of elongating rhizoids, establishing a role for MpROP in the control of polar cell growth and its functional conservation in plants. Furthermore, the Mprop knockout mutant showed defects in the formation of meristem notches associated with disorganized cell division patterns. These results reveal a critical function of MpROP in the regulation of plant development. Interestingly, these phenotypes were complemented not only by MpROP but also Arabidopsis AtROP2, supporting the conservation of ROP's function among land plants. Our results demonstrate a great potential for M. polymorpha as a powerful genetic system for functional and mechanistic elucidation of ROP signaling pathways during plant development.

An Automated High-Throughput Phenotyping System for Marchantia polymorpha.

High-throughput phenotyping (HTP) allows automation of fast and precise acquisition and analysis of digital images for the detection of key traits in real time. HTP improves characterization of the growth and development of plants in controlled environments in a nondestructive fashion. Marchantia polymorpha has emerged as a very attractive model for studying the evolution of the physiological, cellular, molecular, and developmental adaptations that enabled plants to conquer their terrestrial environments. The availability of the M. polymorpha genome in combination with a full set of functional genomic tools including genetic transformation, homologous recombination, and genome editing has allowed the inspection of its genome through forward and reverse genetics approaches. The increasing number of mutants has made it possible to perform informative genome-wide analyses to study the phenotypic consequences of gene inactivation. Here we present an HTP protocol for M. polymorpha that will aid current efforts to quantify numerous morphological parameters that can potentially reveal genotype-to-phenotype relationships and relevant connections between individual traits.

Functional Analyses of the Two Distinctive Types of Two-Pore Channels and the Slow Vacuolar Channel in Marchantia polymorpha.

The two-pore channel (TPC) family is widely conserved in eukaryotes. Many vascular plants, including Arabidopsis and rice, possess a single TPC gene which functions as a slow vacuolar (SV) channel-voltage-dependent cation-permeable channel located in the vacuolar membrane (tonoplast). On the other hand, a liverwort Marchantia polymorpha genome encodes three TPC homologs: MpTPC1 is similar to TPCs in vascular plants (type 1 TPC), while MpTPC2 and MpTPC3 are classified into a distinctive group (type 2 TPC). Phylogenetic analysis suggested that the type 2 TPC emerged before the land colonization in plant evolution and was lost in vascular plants and hornworts. All of the three MpTPCs were shown to be localized at the tonoplast. We generated knockout mutants of tpc1, tpc2, tpc3 and tpc2 tpc3 double mutant by clustered regularly interspaced short palindromic repeats/Cas9 genome editing and performed patch-clamp analyses of isolated vacuoles. The SV channel activity was abolished in the Mptpc1 loss-of-function mutant (Mptpc1-1KO), while Mptpc2-1KO, Mptpc3-1KO and Mptpc2-2/tpc3-2KO double mutant exhibited similar activity to the wild type, indicating that MpTPC1 (type 1) is solely responsible for the SV channel activity. Activators of mammalian TPCs, phosphatidylinositol-3,5-bisphosphate and nicotinic acid adenine dinucleotide phosphate, did not affect the ion channel activity of any MpTPCs. These results indicate that the type 1 TPCs, which are well conserved in all land plant species, encode the SV channel, while the type 2 TPCs likely encode other tonoplast cation channel(s) distinct from the SV channel and animal TPCs.

The genome of the zebra mussel, Dreissena polymorpha: a resource for comparative genomics, invasion genetics, and biocontrol.

The zebra mussel, Dreissena polymorpha, continues to spread from its native range in Eurasia to Europe and North America, causing billions of dollars in damage and dramatically altering invaded aquatic ecosystems. Despite these impacts, there are few genomic resources for Dreissena or related bivalves. Although the D. polymorpha genome is highly repetitive, we have used a combination of long-read sequencing and Hi-C-based scaffolding to generate a high-quality chromosome-scale genome assembly. Through comparative analysis and transcriptomics experiments, we have gained insights into processes that likely control the invasive success of zebra mussels, including shell formation, synthesis of byssal threads, and thermal tolerance. We identified multiple intact steamer-like elements, a retrotransposon that has been linked to transmissible cancer in marine clams. We also found that D. polymorpha have an unusual 67 kb mitochondrial genome containing numerous tandem repeats, making it the largest observed in Eumetazoa. Together these findings create a rich resource for invasive species research and control efforts.

The genome of Medicago polymorpha provides insights into its edibility and nutritional value as a vegetable and forage legume

Medicago polymorpha is a nutritious and palatable forage and vegetable plant that also fixes nitrogen. Here, we reveal the chromosome-scale genome sequence of M. polymorpha using an integrated approach including Illumina, PacBio and Hi-C technologies. We combined PacBio full-length RNA-seq, metabolomic analysis, structural anatomy analysis and related physiological indexes to elucidate the important agronomic traits of M. polymorpha for forage and vegetable usage. The assembled M. polymorpha genome consisted of 457.53 Mb with a long scaffold N50 of 57.72 Mb, and 92.92% (441.83 Mb) of the assembly was assigned to seven pseudochromosomes. Comparative genomic analysis revealed that expansion and contraction of the photosynthesis and lignin biosynthetic gene families, respectively, led to enhancement of nutritious compounds and reduced lignin biosynthesis in M. polymorpha. In addition, we found that several positively selected nitrogen metabolism-related genes were responsible for crude protein biosynthesis. Notably, the metabolomic results revealed that a large number of flavonoids, vitamins, alkaloids, and terpenoids were enriched in M. polymorpha. These results imply that the decreased lignin content but relatively high nutrient content of M. polymorpha enhance its edibility and nutritional value as a forage and vegetable. Our genomic data provide a genetic basis that will accelerate functional genomic and breeding research on M. polymorpha as well as other Medicago and legume plants.

Fungal-Type Terpene Synthases in Marchantia polymorpha Are Involved in Sesquiterpene Biosynthesis in Oil Body Cells.

The liverwort Marchantia polymorpha possesses oil bodies in idioblastic oil body cells scattered in its thallus. Oil bodies are subcellular organelles in which specific sesquiterpenes and bisbibenzyls are accumulated. Therefore, a specialized system for the biosynthesis and accumulation of these defense compounds specifically in oil bodies has been implied. A recent study on M. polymorpha genome sequencing revealed 10 genes that shared high similarities with fungal-type terpene synthases (TPSs). Eight of these fungal-type TPS-like genes in M. polymorpha (MpFTPSL1-6, -9 and -10) are located within a 376-kb stretch on chromosome 6 and share similarities of over 94% at the nucleotide level. Therefore, these genes have likely originated from recent gene duplication events. The expression of a subset of MpFTPSLs was induced under non-axenic growth on vermiculite, which increased the amounts of sesquiterpenes and number of oil bodies. The tdTomato fluorescent protein-based in-fusion reporter assay with MpFTPSL2 promoter revealed fluorescent signals specifically in oil body cells of the thallus, indicating that MpFTPSL2 functions in oil body cells. Recombinant MpFTPSL2 expression in Escherichia coli led to sesquiterpene synthesis from farnesyl pyrophosphate. Moreover, suppression of a subset of MpFTPSLs through RNA interference reduced sesquiterpene accumulation in thalli grown on vermiculite. Taken together, these results suggest that at least a subset of MpFTPSLs is involved in sesquiterpene synthesis in oil body cells.

Diversity of Pectin Rhamnogalacturonan I Rhamnosyltransferases in Glycosyltransferase Family 106.

Rhamnogalacturonan I (RG-I) comprises approximately one quarter of the pectin molecules in land plants, and the backbone of RG-I consists of a repeating sequence of [2)-α-L-Rha(1-4)-α-D-GalUA(1-] disaccharide. Four Arabidopsis thaliana genes encoding RG-I rhamnosyltransferases (AtRRT1 to AtRRT4), which synthesize the disaccharide repeats, have been identified in the glycosyltransferase family (GT106). However, the functional role of RG-I in plant cell walls and the evolutional history of RRTs remains to be clarified. Here, we characterized the sole ortholog of AtRRT1–AtRRT4 in liverwort, Marchantia polymorpha, namely, MpRRT1. MpRRT1 had RRT activity and genetically complemented the AtRRT1-deficient mutant phenotype in A. thaliana. However, the MpRRT1-deficient M. polymorpha mutants showed no prominent morphological changes and only an approximate 20% reduction in rhamnose content in the cell wall fraction compared to that in wild-type plants, suggesting the existence of other RRT gene(s) in the M. polymorpha genome. As expected, we detected RRT activities in other GT106 family proteins such as those encoded by MpRRT3 in M. polymorpha and FRB1/AtRRT8 in A. thaliana, the deficient mutant of which affects cell adhesion. Our results show that RRT genes are more redundant and diverse in GT106 than previously thought.

Epiphyllous bryophytes in Arboretum Mlyňany (Slovakia)

Epiphyllous bryophytes in Arboretum Mlyňany (Slovakia)

Insights into Land Plant Evolution Garnered from the Marchantia polymorpha Genome

The evolution of land flora transformed the terrestrial environment. Land plants evolved from an ancestral charophycean alga from which they inherited developmental, biochemical, and cell biological attributes. Additional biochemical and physiological adaptations to land, and a life cycle with an alternation between multicellular haploid and diploid generations that facilitated efficient dispersal of desiccation tolerant spores, evolved in the ancestral land plant. We analyzed the genome of the liverwort Marchantia polymorpha, a member of a basal land plant lineage. Relative to charophycean algae, land plant genomes are characterized by genes encoding novel biochemical pathways, new phytohormone signaling pathways (notably auxin), expanded repertoires of signaling pathways, and increased diversity in some transcription factor families. Compared with other sequenced land plants, M.polymorpha exhibits low genetic redundancy in most regulatory pathways, with this portion of its genome resembling that predicted for the ancestral land plant. PAPERCLIP.

Transcriptional activator Cat8 is involved in regulation of xylose alcoholic fermentation in the thermotolerant yeast Ogataea (Hansenula) polymorpha

BackgroundEfficient xylose alcoholic fermentation is one of the key to a successful lignocellulosic ethanol production. However, regulation of this process in the native xylose-fermenting yeasts is poorly understood. In this work, we paid attention to the transcriptional factor Cat8 and its possible role in xylose alcoholic fermentation in Ogataea (Hansenula) polymorpha. In Saccharomyces cerevisiae, organism, which does not metabolize xylose, gene CAT8 encodes a Zn-cluster transcriptional activator necessary for expression of genes involved in gluconeogenesis, respiration, glyoxylic cycle and ethanol utilization. Xylose is a carbon source that could be fermented to ethanol and simultaneously could be used in gluconeogenesis for hexose synthesis. This potentially suggests involvement of CAT8 in xylose metabolism.ResultsHere, the role of CAT8 homolog in the natural xylose-fermenting thermotolerant yeast O. polymorpha was characterized. The CAT8 ortholog was identified in O. polymorpha genome and deleted both in the wild-type strain and in advanced ethanol producer from xylose. Constructed cat8Δ strain isolated from wild strain showed diminished growth on glycerol, ethanol and xylose as well as diminished respiration on the last substrate. At the same time, cat8Δ mutant isolated from the best available O. polymorpha ethanol producer showed only visible defect in growth on ethanol. CAT8 deletant was characterized by activated transcription of genes XYL3, DAS1 and RPE1 and slight increase in the activity of several enzymes involved in xylose metabolism and alcoholic fermentation. Ethanol production from xylose in cat8Δ mutants in the background of wild-type strain and the best available ethanol producer from xylose increased for 50 and 30%, respectively. The maximal titer of ethanol during xylose fermentation was 12.5 g ethanol/L at 45 °C. Deletion of CAT8 did not change ethanol production from glucose. Gene CAT8 was also overexpressed under control of the strong constitutive promoter GAP of glyceraldehyde-3-phosphate dehydrogenase. Corresponding strains showed drop in ethanol production in xylose medium whereas glucose alcoholic fermentation remained unchanged. Available data suggest on specific role of Cat8 in xylose alcoholic fermentation.ConclusionsThe CAT8 gene is one of the first identified genes specifically involved in regulation of xylose alcoholic fermentation in the natural xylose-fermenting yeast O. polymorpha.

Genome sequencing of Metrosideros polymorpha (Myrtaceae), a dominant species in various habitats in the Hawaiian Islands with remarkable phenotypic variations.

Whole genome sequences, which can be provided even for non-model organisms owing to high-throughput sequencers, are valuable in enhancing the understanding of adaptive evolution. Metrosideros polymorpha, a tree species endemic to the Hawaiian Islands, occupies a wide range of ecological habitats and shows remarkable polymorphism in phenotypes among/within populations. The biological functions of genetic variations observed within this species could provide significant insights into the adaptive radiation found in a single species. Here de novo assembled genome sequences of M. polymorpha are presented to reveal basic genomic parameters about this species and to develop our knowledge of ecological divergences. The assembly yielded 304-Mbp genome sequences, half of which were covered by 19 scaffolds with>5 Mbp, and contained 30K protein-coding genes. Demographic history inferred from the genome-wide heterozygosity indicated that this species experienced a dramatic rise and fall in the effective population size, possibly owing to past geographic or climatic changes in the Hawaiian Islands. This M. polymorpha genome assembly represents a high-quality genome resource useful for future functional analyses of both intra- and interspecies genetic variations or comparative genomics.

SNARE Molecules in Marchantia polymorpha: Unique and Conserved Features of the Membrane Fusion Machinery.

The membrane trafficking pathway has been diversified in a specific way for each eukaryotic lineage, probably to fulfill specific functions in the organisms. In green plants, comparative genomics has supported the possibility that terrestrialization and/or multicellularization could be associated with the elaboration and diversification of membrane trafficking pathways, which have been accomplished by an expansion of the numbers of genes required for machinery components of membrane trafficking, including soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins. However, information regarding membrane trafficking pathways in basal land plant lineages remains limited. In the present study, we conducted extensive analyses of SNARE molecules, which mediate membrane fusion between target membranes and transport vesicles or donor organelles, in the liverwort, Marchantia polymorpha. The M. polymorpha genome contained at least 34 genes for 36 SNARE proteins, comprising fundamental sets of SNARE proteins that are shared among land plant lineages with low degrees of redundancy. We examined the subcellular distribution of a major portion of these SNARE proteins by expressing Citrine-tagged SNARE proteins in M. polymorpha, and the results showed that some of the SNARE proteins were targeted to different compartments from their orthologous products in Arabidopsis thaliana. For example, MpSYP12B was localized to the surface of the oil body, which is a unique organelle in liverworts. Furthermore, we identified three VAMP72 members with distinctive structural characteristics, whose N-terminal extensions contain consensus sequences for N-myristoylation. These results suggest that M. polymorpha has acquired unique membrane trafficking pathways associated with newly acquired machinery components during evolution.

Functional analysis of allene oxide cyclase, MpAOC, in the liverwort Marchantia polymorpha

12-Oxo-phytodienoic acid (OPDA) is an intermediate in jasmonic acid (JA) biosynthesis. OPDA exerts JA-dependent and JA-independent biological effects; therefore, it is considered a signaling molecule in flowering plants. OPDA is induced by bacterial infection and wounding and inhibits growth in the moss Physcomitrella patens. The functions of OPDA and allene oxide cyclase (AOC) in the liverwort Marchantia polymorpha were explored, which represents the most basal lineage of extant land plants. The analysis of OPDA showed that it is present in M. polymorpha and is increased by wounding. OPDA has been suggested to be involved in the response to environmental stresses. Moreover, OPDA showed growth inhibitory activity in M. polymorpha. Nonetheless JA in M. polymorpha was not found in this study. AOC synthesizes OPDA from an unstable allene oxide. A database search of the M. polymorpha genome identified only a putative gene encoding allene oxide cyclase (MpAOC). Recombinant MpAOC showed AOC activity similar to that in flowering plants. MpAOC was localized to chloroplasts, as in flowering plants. Expression of MpAOC was induced by wounding and OPDA treatment, and positive feedback regulation of OPDA was demonstrated in M. polymorpha. Overexpression of MpAOC increased the endogenous OPDA level and suppressed growth in M. polymorpha. These results indicate the role of OPDA as a signaling molecule regulating growth and the response to wounding in the liverwort M. polymorpha.

Inversion of the chromosomal region between two mating type loci switches the mating type in Hansenula polymorpha.

Yeast mating type is determined by the genotype at the mating type locus (MAT). In homothallic (self-fertile) Saccharomycotina such as Saccharomyces cerevisiae and Kluveromyces lactis, high-efficiency switching between a and α mating types enables mating. Two silent mating type cassettes, in addition to an active MAT locus, are essential components of the mating type switching mechanism. In this study, we investigated the structure and functions of mating type genes in H. polymorpha (also designated as Ogataea polymorpha). The H. polymorpha genome was found to harbor two MAT loci, MAT1 and MAT2, that are ∼18 kb apart on the same chromosome. MAT1-encoded α1 specifies α cell identity, whereas none of the mating type genes were required for a identity and mating. MAT1-encoded α2 and MAT2-encoded a1 were, however, essential for meiosis. When present in the location next to SLA2 and SUI1 genes, MAT1 or MAT2 was transcriptionally active, while the other was repressed. An inversion of the MAT intervening region was induced by nutrient limitation, resulting in the swapping of the chromosomal locations of two MAT loci, and hence switching of mating type identity. Inversion-deficient mutants exhibited severe defects only in mating with each other, suggesting that this inversion is the mechanism of mating type switching and homothallism. This chromosomal inversion-based mechanism represents a novel form of mating type switching that requires only two MAT loci.