Storage of monokaryotic strains of Podospora anserina

Maintenance of Podospora anserina strains for experimental purposes is very time consuming (see Esser 1969 Neurospora Newsl. 15:27-31) and methods have been published that address this issue by freezing ascospores at -80 oC (Begel and Belcour 1991 Fungal Genet. Newsl. 38:67). Although the latter approach does reduce the amount of time required for yearly sexual crosses and ascospore isolation, it does not resolve the problem of the time required to rapidly generate monokaryotic hyphae, that are needed as a source for inoculum for many types of experiments

Two new species of Ophiostomatales (Sordariomycetes) associated with the bark beetle Dryocoetes alni from Poland

Bark beetles belonging to the genus Dryocoetes (Coleoptera, Curculionidae, Scolytinae) are known vectors of fungi, such as the pathogenic species Grosmannia dryocoetidis involved in alpine fir (Abies lasiocarpa) mortality. Associations between hardwood-infesting Dryocoetes species and fungi in Europe have received very little research attention. Ectosymbiotic fungi residing in Ceratocystiopsis and Leptographium (Ophiostomatales, Sordariomycetes, Ascomycota) were commonly detected in previous surveys of the Dryocoetes alni-associated mycobiome in Poland. The aim of this study was to accurately identify these isolates and to provide descriptions of the new species. The identification was conducted based on morphology and DNA sequence data for six loci (ITS1-5.8S, ITS2-28S, ACT, CAL, TUB2, and TEF1-α). This revealed two new species, described here as Ceratocystiopsis synnemata sp. nov. and Leptographium alneum sp. nov. The host trees for the new species included Alnus incana and Populus tremula. Ceratocystiopsis synnemata can be distinguished from its closely related species, C. pallidobrunnea, based on conidia morphology and conidiophores that aggregate in loosely arranged synnemata. Leptographium alneum is closely related to Grosmannia crassivaginata and differs from this species in having a larger ascomatal neck, and the presence of larger club-shaped cells.202

Uncovering the temporal dynamics and regulatory networks of thermal stress response in a hyperthermophile using transcriptomics and proteomics

Facing rapid fluctuations in their natural environment, extremophiles, like the hyperthermophilic archaeon Pyrococcus furiosus, exhibit remarkable adaptability to extreme conditions. However, our understanding of their dynamic cellular responses remains limited. This study integrates RNA-sequencing and mass spectrometry data, thereby elucidating transcriptomic and proteomic responses to heat and cold shock stress in P. furiosus. Our results reveal rapid and dynamic changes in gene and protein expression following these stress responses. Heat shock triggers extensive transcriptome reprogramming, orchestrated by the transcriptional regulator Phr, targeting a broader gene repertoire than previously demonstrated. For heat shock signature genes, RNA levels swiftly return to baseline upon recovery, while protein levels remain persistently upregulated, reflecting a rapid but sustained response. Intriguingly, cold shock at 4°C elicits distinct short- and long-term responses at both RNA and protein levels. Cluster analysis identified gene sets with either congruent or contrasting trends in RNA and protein changes, representing well-separated arCOG groups tailored to their individual cellular responses. Particularly, upregulation of ribosomal proteins and significant enrichment of 5′-leadered sequences in cold-shock responsive genes suggest that translation regulation is important during cold shock adaption. Further investigating transcriptomic features, we reveal that thermal stress genes are equipped with basal sequence elements, such as strong promoter and poly(U)-terminators, facilitating a regulated response of the respective transcription units. Our study provides a comprehensive overview of the cellular response to temperature stress, advancing our understanding of stress response mechanisms in hyperthermophilic archaea and providing valuable insights into the molecular adaptations that facilitate life in extreme environments

The mitochondrial genome of Ophiostoma himal-ulmi and comparison with other Dutch elm disease causing fungi.

The mitochondrial genome of Ophiostoma himal-ulmi, a species endemic to the Western Himalayas and a member of the Dutch elm disease-causing fungi, has been sequenced and characterized. The mitochondrial genome was compared with other available genomes for members of the Ophiostomatales, including other agents of Dutch elm disease (Ophiostoma ulmi, Ophiostoma novo-ulmi subspecies novo-ulmi and Ophiostoma novo-ulmi subspecies americana) and it was noted that gene synteny is highly conserved and variability among members of the Dutch-elm disease-causing fungi is primarily due to the number of intron insertions. Among the Dutch elm disease-causing fungi examined, O. himal-ulmi has the largest mitochondrial genomes ranging from 94 934 bp to 111 712 bp due to the expansion of the number of introns.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Organellar Introns in Fungi, Algae, and Plants

Introns are ubiquitous in eukaryotic genomes and have long been considered as ‘junk RNA’ but the huge energy expenditure in their transcription, removal, and degradation indicate that they may have functional significance and can offer evolutionary advantages. In fungi, plants and algae introns make a significant contribution to the size of the organellar genomes. Organellar introns are classified as catalytic self-splicing introns that can be categorized as either Group I or Group II introns. There are some biases, with Group I introns being more frequently encountered in fungal mitochondrial genomes, whereas among plants Group II introns dominate within the mitochondrial and chloroplast genomes. Organellar introns can encode a variety of proteins, such as maturases, homing endonucleases, reverse transcriptases, and, in some cases, ribosomal proteins, along with other novel open reading frames. Although organellar introns are viewed to be ribozymes, they do interact with various intron- or nuclear genome-encoded protein factors that assist in the intron RNA to fold into competent splicing structures, or facilitate the turn-over of intron RNAs to prevent reverse splicing. Organellar introns are also known to be involved in non-canonical splicing, such as backsplicing and trans-splicing which can result in novel splicing products or, in some instances, compensate for the fragmentation of genes by recombination events. In organellar genomes, Group I and II introns may exist in nested intronic arrangements, such as introns within introns, referred to as twintrons, where splicing of the external intron may be dependent on splicing of the internal intron. These nested or complex introns, with two or three-component intron modules, are being explored as platforms for alternative splicing and their possible function as molecular switches for modulating gene expression which could be potentially applied towards heterologous gene expression. This review explores recent findings on organellar Group I and II introns, focusing on splicing and mobility mechanisms aided by associated intron/nuclear encoded proteins and their potential roles in organellar gene expression and cross talk between nuclear and organellar genomes. Potential application for these types of elements in biotechnology are also discussed

Using Group II Introns for Attenuating the In Vitro and In Vivo Expression of a Homing Endonuclease.

In Chaetomium thermophilum (DSM 1495) within the mitochondrial DNA (mtDNA) small ribosomal subunit (rns) gene a group IIA1 intron interrupts an open reading frame (ORF) encoded within a group I intron (mS1247). This arrangement offers the opportunity to examine if the nested group II intron could be utilized as a regulatory element for the expression of the homing endonuclease (HEase). Constructs were generated where the codon-optimized ORF was interrupted with either the native group IIA1 intron or a group IIB type intron. This study showed that the expression of the HEase (in vivo) in Escherichia coli can be regulated by manipulating the splicing efficiency of the HEase ORF-embedded group II introns. Exogenous magnesium chloride (MgCl2) stimulated the expression of a functional HEase but the addition of cobalt chloride (CoCl2) to growth media antagonized the expression of HEase activity. Ultimately the ability to attenuate HEase activity might be useful in precision genome engineering, minimizing off target activities, or where pathways have to be altered during a specific growth phase

Evolutionary dynamics of the mS952 intron: a novel mitochondrial group II intron encoding a LAGLIDADG homing endonuclease gene.

Examination of the mitochondrial small subunit ribosomal RNA (rns) gene of five species of the fungal genus Leptographium revealed that the gene has been invaded at least once at position 952 by a group II intron encoding a LAGLIDADG homing endonuclease gene. Phylogenetic analyses of the intron and homing endonuclease sequences indicated that each element in Leptographium species forms a single clade and is closely related to the group II intron/homing endonuclease gene composite element previously reported at position 952 of the mitochondrial rns gene of Cordyceps species and of Cryphonectria parasitica. The results of an intron survey of the mt rns gene of Leptographium species superimposed onto the phylogenetic analysis of the host organisms suggest that the composite element was transmitted vertically in Leptographium lundbergii. However, its stochastic distribution among strains of L. wingfieldii, L. terebrantis, and L. truncatum suggests that it has been horizontally transmitted by lateral gene transfer among these species, although the random presence of the intron may reflect multiple random loss events. A model is proposed describing the initial invasion of the group II intron in the rns gene of L. lundbergii by a LAGLIDADG homing endonuclease gene and subsequent evolution of this gene to recognize a novel DNA target site, which may now promote the mobility of the intron and homing endonuclease gene as a composite element.Journal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Programmable Genome Editing Tools and their Regulation for Efficient Genome Engineering

Targeted genome editing has become a powerful genetic tool for studying gene function or for modifying genomes by correcting defective genes or introducing genes. A variety of reagents have been developed in recent years that can generate targeted double-stranded DNA cuts which can be repaired by the error-prone, non-homologous end joining repair system or via the homologous recombination-based double-strand break repair pathway provided a suitable template is available. These genome editing reagents require components for recognizing a specific DNA target site and for DNA-cleavage that generates the double-stranded break. In order to reduce potential toxic effects of genome editing reagents, it might be desirable to control the in vitro or in vivo activity of these reagents by incorporating regulatory switches that can reduce off-target activities and/or allow for these reagents to be turned on or off. This review will outline the various genome editing tools that are currently available and describe the strategies that have so far been employed for regulating these editing reagents. In addition, this review will examine potential regulatory switches/strategies that can be employed in the future in order to provide temporal control for these reagents. Keywords: Meganuclease, TALEN, Zinc finger nuclease, CRISPR/Cas9, Regulatory switch, Hammerhead ribozym