Chaga Genome and Convergent Evolution of Betulinate Biosynthesis in Host (Betula pendula) and the Pathogen (Inonotus obliquus)

Inonotus obliquuse, commonly called chaga mushroom (Pakurikääpä), is known and used by many ethnic societies for its medicinal properties, to the degree that they believed to its magical and supernatural powers. Chaga mushroom mainly grows on Betula species, and easily identifiable when the sterile black conks burst through the bark of a host like Betula pendula (silver birch). The infected host tree might not even show any symptoms for several years, while the white-rot disease is progressing in the heart of the woody stem of the infected tree. However, very little known about the evolutionary history of chaga mushroom, and the molecular dynamics involves in pathogen-host interaction of chaga. In contrast, there is no lack of research over the study of secondary metabolites, extracted from chaga, and their potential for biotechnological, pharmaceutical, and industrial applications. Among the compounds are triterpenoids, such as betulin and betulinic acid, which found to be produced naturally by both chaga and its hosts Betula species. This begs a question what is the molecular mechanism which drive the production of the same compounds in host and the pathogen? To this end, we sequenced, assembled, and annotated the genome of B. pendula to predict high-quality gene models. We also studied the transcriptomic profiling of silver birch bark to understand the molecular pathways which contribute to the triterpenoid’s biosynthesis in different tissues of silver birch bark. In addition to the host genome, we sequenced, assembled, and annotated a high-quality genome of the pathogen (chaga mushroom), which provides an important foundation for our evolutionary analysis. Furthermore, we also identified and characterized the candidate CYP450 monooxygenase genes from silver birch and chaga mushroom, which then for the functional analysis, they subjected to heterologous gene expression analysis.Pakurikääpä, Inonotus obliquuse (Chaga mushroom), on tunnettu ja laajasti käytetty sieni kansanlääketieteessä etenkin Venäjällä ja Aasiassa. Jotkut uskovat sillä olevan vahvaa, jopa yliluonnollista, tehoa useiden sairauksien hoidossa. Pakurikääpä esiintyy yleisimmin koivuissa (Betula spp.) ja sen tunnistaa helpoiten puun kuoren läpi työntyvästä, rihmastopahkan aiheuttamasta tummasta epämuodostumasta, pakurista. Tartunnan saaneessa isäntäpuussa oireet saatetaan havaita vasta vuosia tartunnan jälkeen, sillä tartunnan alkuvaiheessa sienirihmasto lahottaa vain sydänpuuta. Pakurikäävän kehityshistoriasta tai taudinaiheuttajan ja isännän välisestä vuorovaikutuksesta molekyylitasolla tiedetään vielä hyvin vähän. Sen sijaan pakurikäävän toissijaisia aineenvaihduntatuotteita (sekundaarimetaboliitteja) ja niiden mahdollista käyttöä bioteknologian, lääketieteen sekä teollisuuden sovelluksissa, on tutkittu paljon. Tällaisia lupaavia yhdisteitä ovat muun muassa tartunnan saaneiden koivujen sekä pakurikäävän tuottamat triterpenoidit, kuten betuliini ja betuliinihappo. Onkin mielenkiintoinen kysymys, mikä molekyylitason mekanismi saa sekä isännän että taudinaiheuttajan tuottamaan samoja yhdisteitä? Tässä väitöskirjatyössä olemme selvittäneet rauduskoivun (Betula pendula) perimän emäsjärjestyksen sekä määrittäneet perimän geenit. Tutkimme ja analysoimme myös rauduskoivun kuoren transkriptomin ymmärtääksemme paremmin mitkä molekyylitason tapahtumat ovat mukana triterpenoidien biosynteesissä eri solukoissa. Isännän ja taudinaiheuttajan välistä evoluutiota ja vuorovaikutusta tutkiaksemme selvitimme myös pakurikäävän perimän. Lisäksi tunnistimme ja kuvailimme CYP450 mono-oksigenaasigeenit rauduskoivusta ja pakurikäävästä sekä tutkimme näiden geenien samanaikaista ilmenemistä isännässä ja taudinaiheuttajassa

Mitochondrial Nascent Chain Quality Control Determines Organelle Form and Function

Proteotoxicity has long been considered a key factor in mitochondrial dysfunction and human disease. The origin of the endogenous offending toxic substrates and the regulatory pathways to deal with these insults, however, have remained unclear. Mitochondria maintain a compartmentalized gene expression system that in animals is only responsible for synthesis of 1% of the organelle proteome. Because of the relatively small contribution of the mitochondrial genome to the overall proteome, the synthesis and quality control of these nascent chains to maintain organelle proteostasis has long been overlooked. However, recent research has uncovered mechanisms by which defects to the quality control of mitochondrial gene expression are linked to a novel cellular stress response that impinges upon organelle form and function and cell fitness. In this review, we discuss the mechanisms for a key event in the response: activation of the metalloprotease OMA1. This severs the membrane tether of the dynamin-related GTPase OPA1, which is a critical determinant for mitochondrial morphology and function. We also highlight the evolutionary conservation from bacteria of these quality-control mechanisms to maintain membrane integrity, gene expression, and cell fitness.Peer reviewe

Ozone responses in Arabidopsis : beyond stomatal conductance

Tropospheric ozone (O-3) is a major air pollutant that decreases yield of important crops worldwide. Despite long-lasting research of its negative effects on plants, there are many gaps in our knowledge on how plants respond to O-3. In this study, we used natural variation in the model plant Arabidopsis (Arabidopsis thaliana) to characterize molecular and physiological mechanisms underlying O-3 sensitivity. A key parameter in models for O-3 damage is stomatal uptake. Here we show that the extent of O-3 damage in the sensitive Arabidopsis accession Shahdara (Sha) does not correspond with O-3 uptake, pointing toward stomata-independent mechanisms for the development of O-3 damage. We compared tolerant (Col-0) versus sensitive accessions (Sha, Cvi-0) in assays related to photosynthesis, cell death, antioxidants, and transcriptional regulation. Acute O-3 exposure increased cell death, development of lesions in the leaves, and decreased photosynthesis in sensitive accessions. In both Sha and Cvi-0, O-3-induced lesions were associated with decreased maximal chlorophyll fluorescence and low quantum yield of electron transfer from Photosystem II to plastoquinone. However, O-3-induced repression of photosynthesis in these two O-3-sensitive accessions developed in different ways. We demonstrate that O-3 sensitivity in Arabidopsis is influenced by genetic diversity given that Sha and Cvi-0 developed accession-specific transcriptional responses to O-3. Our findings advance the understanding of plant responses to O-3 and set a framework for future studies to characterize molecular and physiological mechanisms allowing plants to respond to high O-3 levels in the atmosphere as a result of high air pollution and climate change.Peer reviewe

Arabidopsis MLO2 is a negative regulator of sensitivity to extracellular reactive oxygen species

The atmospheric pollutant ozone (O-3) is a strong oxidant that causes extracellular reactive oxygen species (ROS) formation, has significant ecological relevance, and is used here as a non-invasive ROS inducer to study plant signalling. Previous genetic screens identified several mutants exhibiting enhanced O-3 sensitivity, but few with enhanced tolerance. We found that loss-of-function mutants in Arabidopsis MLO2, a gene implicated in susceptibility to powdery mildew disease, exhibit enhanced dose-dependent tolerance to O-3 and extracellular ROS, but a normal response to intracellular ROS. This phenotype is increased in a mlo2 mlo6 mlo12 triple mutant, reminiscent of the genetic redundancy of MLO genes in powdery mildew resistance. Stomatal assays revealed that enhanced O-3 tolerance in mlo2 mutants is not caused by altered stomatal conductance. We explored modulation of the mlo2-associated O-3 tolerance, powdery mildew resistance, and early senescence phenotypes by genetic epistasis analysis, involving mutants with known effects on ROS sensitivity or antifungal defence. Mining of publicly accessible microarray data suggests that these MLO proteins regulate accumulation of abiotic stress response transcripts, and transcript accumulation of MLO2 itself is O-3 responsive. In summary, our data reveal MLO2 as a novel negative regulator in plant ROS responses, which links biotic and abiotic stress response pathways.Peer reviewe

Nonstop mRNAs generate a ground state of mitochondrial gene expression noise

Funding Information: This work was supported by the Academy of Finland (307431 and 314706 to B.J.B.), the Sigrid Juselius Foundation Senior Investigator Award to B.J.B., and United Mitochondrial Disease Foundation (PI-16-0598 to B.J.B.) and donations from the Hereditary Neuropathy Foundation, Lindsey Flynt, and Medtronic to B.J.B.; the Orion Research Foundation and the Finnish Cultural Foundation to K.Y.N.; the Academy of Finland (321961 to U.R.); the Sigrid Juselius Foundation, the Academy of Finland (331556), and the Jane and Aatos Erkko Foundation to C.D.D.; Action Medical Research (GN2494 to W.G.N.) and the Manchester NIHR Biomedical Research Centre (IS-BRC-1215-20007 to W.G.N.); the Wellcome Centre for Mitochondrial Research (203105/Z/16/Z to R.W.T.), the Mitochondrial Disease Patient Cohort (UK) (G0800674 to R.W.T.), the Medical Research Council International Centre for Genomic Medicine in Neuromuscular Disease (MR/S005021/1 to R.W.T.), the Lily Foundation, the UK NIHR Biomedical Research Centre for Ageing and Age-related disease award to the Newcastle upon Tyne Foundation Hospitals NHS Trust, the Pathological Society, and the UK NHS Highly Specialised Service for Rare Mitochondrial Disorders of Adults and Children to R.W.T.; Medical Research Council (MR/W019027/1 to W.G.N. and R.W.T.); the Academy of Finland (338836 and 314672 to V.O.P.); and the Sigrid Juselius Foundation and the Jane and Aatos Erkko Foundation. Publisher Copyright: Copyright © 2022 The Authors, some rights reserved;A stop codon within the mRNA facilitates coordinated termination of protein synthesis, releasing the nascent polypeptide from the ribosome. This essential step in gene expression is impeded with transcripts lacking a stop codon, generating nonstop ribosome complexes. Here, we use deep sequencing to investigate sources of nonstop mRNAs generated from the human mitochondrial genome. We identify diverse types of nonstop mRNAs on mitochondrial ribosomes that are resistant to translation termination by canonical release factors. Failure to resolve these aberrations by the mitochondrial release factor in rescue (MTRFR) imparts a negative regulatory effect on protein synthesis that is associated with human disease. Our findings reveal a source of underlying noise in mitochondrial gene expression and the importance of responsive ribosome quality control mechanisms for cell fitness and human health.Peer reviewe

The photoreceptor UVR8 mediates the perception of both UV-B and UV-A wavelengths up to 350 nm of sunlight with responsivity moderated by cryptochromes

ABSTRACT The photoreceptors UV RESISTANCE LOCUS 8 (UVR8) and CRYPTOCHROMES 1 and 2 (CRYs) play major roles in the perception of UV-B (280?315?nm) and UV-A/blue radiation (315?500?nm), respectively. However, it is poorly understood how they function in sunlight. The roles of UVR8 and CRYs were assessed in a factorial experiment with Arabidopsis thaliana wild-type and photoreceptor mutants exposed to sunlight for 6?h or 12?h under five types of filters with cut-offs in UV and blue-light regions. Transcriptome-wide responses triggered by UV-B and UV-A wavelengths shorter than 350?nm (UV-Asw) required UVR8 whereas those induced by blue and UV-A wavelengths longer than 350?nm (UV-Alw) required CRYs. UVR8 modulated gene expression in response to blue light while lack of CRYs drastically enhanced gene expression in response to UV-B and UV-Asw. These results agree with our estimates of photons absorbed by these photoreceptors in sunlight and with in vitro monomerization of UVR8 by wavelengths up to 335?nm. Motif enrichment analysis predicted complex signaling downstream of UVR8 and CRYs. Our results highlight that it is important to use UV waveband definitions specific to plants' photomorphogenesis as is routinely done in the visible region. This article is protected by copyright. All rights reserved.Peer reviewe

Detecting early signs of heat and drought stress in Phoenix dactylifera (date palm)

Plants adapt to the environment by either long-term genome evolution or by acclimatization processes where the cellular processes and metabolism of the plant are adjusted within the existing potential in the genome. Here we studied the adaptation strategies in date palm, Phoenix dactylifera, under mild heat, drought and combined heat and drought by transcriptomic and metabolomic profiling. In transcriptomics data, combined heat and drought resembled heat response, whereas in metabolomics data it was more similar to drought. In both conditions, soluble carbohydrates, such as fucose, and glucose derivatives, were increased, suggesting a switch to carbohydrate metabolism and cell wall biogenesis. This result is consistent with the evidence from transcriptomics and cis-motif analysis. In addition, transcriptomics data showed transcriptional activation of genes related to reactive oxygen species in all three conditions (drought, heat, and combined heat and drought), suggesting increased activity of enzymatic antioxidant systems in cytosol, chloroplast and peroxisome. Finally, the genes that were differentially expressed in heat and combined heat and drought stresses were significantly enriched for circadian and diurnal rhythm motifs, suggesting new stress avoidance strategies.Peer reviewe

Tissue-specific study across the stem reveals the chemistry and transcriptome dynamics of birch bark.

Tree bark is a highly specialized array of tissues that plays important roles in plant protection and development. Bark tissues develop from two lateral meristems; the phellogen (cork cambium) produces the outermost stem-environment barrier called the periderm, while the vascular cambium contributes with phloem tissues. Although bark is diverse in terms of tissues, functions and species, it remains understudied at higher resolution. We dissected the stem of silver birch (Betula pendula) into eight major tissue types, and characterized these by a combined transcriptomics and metabolomics approach. We further analyzed the varying bark types within the Betulaceae family. The two meristems had a distinct contribution to the stem transcriptomic landscape. Furthermore, inter- and intraspecies analyses illustrated the unique molecular profile of the phellem. We identified multiple tissue-specific metabolic pathways, such as the mevalonate/betulin biosynthesis pathway, that displayed differential evolution within the Betulaceae. A detailed analysis of suberin and betulin biosynthesis pathways identified a set of underlying regulators and highlighted the important role of local, small-scale gene duplication events in the evolution of metabolic pathways. This work reveals the transcriptome and metabolic diversity among bark tissues and provides insights to its development and evolution, as well as its biotechnological applications.peerReviewe

Genome sequencing and population genomic analyses provide insights into the adaptive landscape of silver birch

Silver birch (Betula pendula) is a pioneer boreal tree that can be induced to flower within 1 year. Its rapid life cycle, small (440-Mb) genome, and advanced germplasm resources make birch an attractive model for forest biotechnology. We assembled and chromosomally anchored the nuclear genome of an inbred B. pendula individual. Gene duplicates from the paleohexaploid event were enriched for transcriptional regulation, whereas tandem duplicates were overrepresented by environmental responses. Population resequencing of 80 individuals showed effective population size crashes at major points of climatic upheaval. Selective sweeps were enriched among polyploid duplicates encoding key developmental and physiological triggering functions, suggesting that local adaptation has tuned the timing of and cross-talk between fundamental plant processes. Variation around the tightly-linked light response genes PHYC and FRS10 correlated with latitude and longitude and temperature, and with precipitation for PHYC. Similar associations characterized the growth-promoting cytokinin response regulator ARR1, and the wood development genes KAK and MED5A.Peer reviewe

Author Correction: Genome sequencing and population genomic analyses provide insights into the adaptive landscape of silver birch.

In the version of this article initially published, there was a mistake in the calculation of the nucleotide mutation rate per site per generation: 1 × 10−9 mutations per site per generation was used, whereas 9.5 × 10−9 was correct. This error affects the interpretation of population-size changes over time and their possible correspondence with known geological events, as shown in the original Fig. 4 and supporting discussion in the text, as well as details in the Supplementary Note. Neither the data themselves nor any other results are affected. Figure 4 has been revised accordingly. Images of the original and corrected figure panels are shown in the correction notice