Genomic imprinting in plants-revisiting existing models.

Genomic imprinting is an epigenetic phenomenon leading to parentally biased gene expression. Throughout the years, extensive efforts have been made to characterize the epigenetic marks underlying imprinting in animals and plants. As a result, DNA methylation asymmetries between parental genomes emerged as the primary factor controlling the imprinting status of many genes. Nevertheless, the data accumulated so far suggest that this process cannot solely explain the imprinting of all genes. In this review, we revisit the current models explaining imprinting regulation in plants, and discuss novel regulatory mechanisms that could function independently of parental DNA methylation asymmetries in the establishment of imprinting.

Helitrons: genomic parasites that generate developmental novelties.

Helitrons, classified as DNA transposons, employ rolling-circle intermediates for transposition. Distinguishing themselves from other DNA transposons, they leave the original template element unaltered during transposition, which has led to their characterization as 'peel-and-paste elements'. Helitrons possess the ability to capture and mobilize host genome fragments, with enormous consequences for host genomes. This review discusses the current understanding of Helitrons, exploring their origins, transposition mechanism, and the extensive repercussions of their activity on genome structure and function. We also explore the evolutionary conflicts stemming from Helitron-transposed gene fragments and elucidate their domestication for regulating responses to environmental challenges. Looking ahead, further research in this evolving field promises to bring interesting discoveries on the role of Helitrons in shaping genomic landscapes.

Auxin regulates endosperm cellularization in Arabidopsis.

The endosperm is an ephemeral tissue that nourishes the developing embryo, similar to the placenta in mammals. In most angiosperms, endosperm development starts as a syncytium, in which nuclear divisions are not followed by cytokinesis. The timing of endosperm cellularization largely varies between species, and the event triggering this transition remains unknown. Here we show that increased auxin biosynthesis in the endosperm prevents its cellularization, leading to seed arrest. Auxin-overproducing seeds phenocopy paternal-excess triploid seeds derived from hybridizations of diploid maternal plants with tetraploid fathers. Concurrently, auxin-related genes are strongly overexpressed in triploid seeds, correlating with increased auxin activity. Reducing auxin biosynthesis and signaling reestablishes endosperm cellularization in triploid seeds and restores their viability, highlighting a causal role of increased auxin in preventing endosperm cellularization. We propose that auxin determines the time of endosperm cellularization, and thereby uncovered a central role of auxin in establishing hybridization barriers in plants.

Endosperm cellularization failure induces a dehydration-stress response leading to embryo arrest.

The endosperm is a nutritive tissue supporting embryo growth in flowering plants. Most commonly, the endosperm initially develops as a coenocyte (multinucleate cell) and then cellularizes. This process of cellularization is frequently disrupted in hybrid seeds generated by crosses between different flowering plant species or plants that differ in ploidy, resulting in embryo arrest and seed lethality. The reason for embryo arrest upon cellularization failure remains unclear. In this study, we show that triploid Arabidopsis thaliana embryos surrounded by uncellularized endosperm mount an osmotic stress response that is connected to increased levels of abscisic acid (ABA) and enhanced ABA responses. Impairing ABA biosynthesis and signaling aggravated triploid seed abortion, while increasing endogenous ABA levels as well as the exogenous application of ABA-induced endosperm cellularization and suppressed embryo growth arrest. Taking these results together, we propose that endosperm cellularization is required to establish dehydration tolerance in the developing embryo, ensuring its survival during seed maturation.

Epigenetic and transcriptional consequences in the endosperm of chemically induced transposon mobilization in Arabidopsis.

Genomic imprinting, an epigenetic phenomenon leading to parent-of-origin-specific gene expression, has independently evolved in the endosperm of flowering plants and the placenta of mammals-tissues crucial for nurturing embryos. While transposable elements (TEs) frequently colocalize with imprinted genes and are implicated in imprinting establishment, direct investigations of the impact of de novo TE transposition on genomic imprinting remain scarce. In this study, we explored the effects of chemically induced transposition of the Copia element ONSEN on genomic imprinting in Arabidopsis thaliana. Through the combination of chemical TE mobilization and doubled haploid induction, we generated a line with 40 new ONSEN copies. Our findings reveal a preferential targeting of maternally expressed genes (MEGs) for transposition, aligning with the colocalization of H2A.Z and H3K27me3 in MEGs-both previously identified as promoters of ONSEN insertions. Additionally, we demonstrate that chemically-induced DNA hypomethylation induces global transcriptional deregulation in the endosperm, leading to the breakdown of MEG imprinting. This study provides insights into the consequences of chemically induced TE remobilization in the endosperm, revealing that chemically-induced epigenome changes can have long-term consequences on imprinted gene expression.

Auxin: a molecular trigger of seed development.

The evolution of seeds defines a remarkable landmark in the history of land plants. A developing seed contains three genetically distinct structures: the embryo, the nourishing tissue, and the seed coat. While fertilization is necessary to initiate seed development in most plant species, apomicts have evolved mechanisms allowing seed formation independently of fertilization. Despite their socio-economical relevance, the molecular mechanisms driving seed development have only recently begun to be understood. Here we review the current knowledge on the role of the hormone auxin for the initial development of the three seed structures and as a trigger of fertilization-independent seed development.

Molecular basis and evolutionary drivers of endosperm-based hybridization barriers.

The endosperm, a transient seed tissue, plays a pivotal role in supporting embryo growth and germination. This unique feature sets flowering plants apart from gymnosperms, marking an evolutionary innovation in the world of seed-bearing plants. Nevertheless, the importance of the endosperm extends beyond its role in providing nutrients to the developing embryo by acting as a versatile protector, preventing hybridization events between distinct species and between individuals with different ploidy. This phenomenon centers on growth and differentiation of the endosperm and the speed at which both processes unfold. Emerging studies underscore the important role played by type I MADS-box transcription factors, including the paternally expressed gene PHERES1. These factors, along with downstream signaling pathways involving auxin and abscisic acid, are instrumental in regulating endosperm development and, consequently, the establishment of hybridization barriers. Moreover, mutations in various epigenetic regulators mitigate these barriers, unveiling a complex interplay of pathways involved in their formation. In this review, we discuss the molecular underpinnings of endosperm-based hybridization barriers and their evolutionary drivers.

Bypassing reproductive barriers in hybrid seeds using chemically induced epimutagenesis.

The triploid block, which prevents interploidy hybridizations in flowering plants, is characterized by a failure in endosperm development, arrest in embryogenesis, and seed collapse. Many genetic components of triploid seed lethality have been successfully identified in the model plant Arabidopsis thaliana, most notably the paternally expressed genes (PEGs), which are upregulated in tetraploid endosperm with paternal excess. Previous studies have shown that the paternal epigenome is a key determinant of the triploid block response, as the loss of DNA methylation in diploid pollen suppresses the triploid block almost completely. Here, we demonstrate that triploid seed collapse is bypassed in Arabidopsis plants treated with the DNA methyltransferase inhibitor 5-Azacytidine during seed germination and early growth. We identified strong suppressor lines showing stable transgenerational inheritance of hypomethylation in the CG context, as well as normalized expression of PEGs in triploid seeds. Importantly, differentially methylated loci segregate in the progeny of "epimutagenized" plants, which may allow epialleles involved in the triploid block response to be identified in future studies. Finally, we demonstrate that chemically induced epimutagenesis facilitates hybridization between different Capsella species, thus potentially emerging as a strategy for producing triploids and interspecific hybrids with high agronomic interest.

The miRNome function transitions from regulating developmental genes to transposable elements during pollen maturation.

Animal and plant microRNAs (miRNAs) are essential for the spatio-temporal regulation of development. Together with this role, plant miRNAs have been proposed to target transposable elements (TEs) and stimulate the production of epigenetically active small interfering RNAs. This activity is evident in the plant male gamete containing structure, the male gametophyte or pollen grain. How the dual role of plant miRNAs, regulating both genes and TEs, is integrated during pollen development and which mRNAs are regulated by miRNAs in this cell type at a genome-wide scale are unknown. Here, we provide a detailed analysis of miRNA dynamics and activity during pollen development in Arabidopsis thaliana using small RNA and degradome parallel analysis of RNA end high-throughput sequencing. Furthermore, we uncover miRNAs loaded into the two main active Argonaute (AGO) proteins in the uninuclear and mature pollen grain, AGO1 and AGO5. Our results indicate that the developmental progression from microspore to mature pollen grain is characterized by a transition from miRNAs targeting developmental genes to miRNAs regulating TE activity.

Ectopic application of the repressive histone modification H3K9me2 establishes post-zygotic reproductive isolation in Arabidopsis thaliana.

Hybrid seed lethality as a consequence of interspecies or interploidy hybridizations is a major mechanism of reproductive isolation in plants. This mechanism is manifested in the endosperm, a dosage-sensitive tissue supporting embryo growth. Deregulated expression of imprinted genes such as ADMETOS (ADM) underpin the interploidy hybridization barrier in Arabidopsis thaliana; however, the mechanisms of their action remained unknown. In this study, we show that ADM interacts with the AT hook domain protein AHL10 and the SET domain-containing SU(VAR)3-9 homolog SUVH9 and ectopically recruits the heterochromatic mark H3K9me2 to AT-rich transposable elements (TEs), causing deregulated expression of neighboring genes. Several hybrid incompatibility genes identified in Drosophila encode for dosage-sensitive heterochromatin-interacting proteins, which has led to the suggestion that hybrid incompatibilities evolve as a consequence of interspecies divergence of selfish DNA elements and their regulation. Our data show that imbalance of dosage-sensitive chromatin regulators underpins the barrier to interploidy hybridization in Arabidopsis, suggesting that reproductive isolation as a consequence of epigenetic regulation of TEs is a conserved feature in animals and plants.

Updated Phylogeny and Protein Structure Predictions Revise the Hypothesis on the Origin of MADS-box Transcription Factors in Land Plants.

MADS-box transcription factors (TFs), among the first TFs extensively studied, exhibit a wide distribution across eukaryotes and play diverse functional roles. Varying by domain architecture, MADS-box TFs in land plants are categorized into Type I (M-type) and Type II (MIKC-type). Type I and II genes have been considered orthologous to the SRF and MEF2 genes in animals, respectively, presumably originating from a duplication before the divergence of eukaryotes. Here, we exploited the increasing availability of eukaryotic MADS-box sequences and reassessed their evolution. While supporting the ancient duplication giving rise to SRF- and MEF2-types, we found that Type I and II genes originated from the MEF2-type genes through another duplication in the most recent common ancestor (MRCA) of land plants. Protein structures predicted by AlphaFold2 and OmegaFold support our phylogenetic analyses, with plant Type I and II TFs resembling the MEF2-type structure, rather than SRFs. We hypothesize that the ancestral SRF-type TFs were lost in the MRCA of Archaeplastida (the kingdom Plantae sensu lato). The retained MEF2-type TFs acquired a Keratin-like domain and became MIKC-type before the divergence of Streptophyta. Subsequently in the MRCA of land plants, M-type TFs evolved from a duplicated MIKC-type precursor through loss of the Keratin-like domain, leading to the Type I clade. Both Type I and II TFs expanded and functionally differentiated in concert with the increasing complexity of land plant body architecture. The recruitment of these originally stress-responsive TFs into developmental programs, including those underlying reproduction, may have facilitated the adaptation to the terrestrial environment.

Hybrid seed incompatibility in Capsella is connected to chromatin condensation defects in the endosperm.

Hybridization of closely related plant species is frequently connected to endosperm arrest and seed failure, for reasons that remain to be identified. In this study, we investigated the molecular events accompanying seed failure in hybrids of the closely related species pair Capsella rubella and C. grandiflora. Mapping of QTL for the underlying cause of hybrid incompatibility in Capsella identified three QTL that were close to pericentromeric regions. We investigated whether there are specific changes in heterochromatin associated with interspecific hybridizations and found a strong reduction of chromatin condensation in the endosperm, connected with a strong loss of CHG and CHH methylation and random loss of a single chromosome. Consistent with reduced DNA methylation in the hybrid endosperm, we found a disproportionate deregulation of genes located close to pericentromeric regions, suggesting that reduced DNA methylation allows access of transcription factors to targets located in heterochromatic regions. Since the identified QTL were also associated with pericentromeric regions, we propose that relaxation of heterochromatin in response to interspecies hybridization exposes and activates loci leading to hybrid seed failure.

Endosperm Evolution by Duplicated and Neofunctionalized Type I MADS-Box Transcription Factors.

MADS-box transcription factors (TFs) are present in nearly all major eukaryotic groups. They are divided into Type I and Type II that differ in domain structure, functional roles, and rates of evolution. In flowering plants, major evolutionary innovations like flowers, ovules, and fruits have been closely connected to Type II MADS-box TFs. The role of Type I MADS-box TFs in angiosperm evolution remains to be identified. Here, we show that the formation of angiosperm-specific Type I MADS-box clades of Mγ and Mγ-interacting Mα genes (Mα*) can be tracked back to the ancestor of all angiosperms. Angiosperm-specific Mγ and Mα* genes were preferentially expressed in the endosperm, consistent with their proposed function as heterodimers in the angiosperm-specific embryo nourishing endosperm tissue. We propose that duplication and diversification of Type I MADS genes underpin the evolution of the endosperm, a developmental innovation closely connected to the origin and success of angiosperms.

Meiocyte size is a determining factor for unreduced gamete formation in Arabidopsis thaliana.

Polyploidy, the presence of more than two sets of chromosomes within a cell, is a widespread phenomenon in plants. The main route to polyploidy is considered through the production of unreduced gametes that are formed as a consequence of meiotic defects. Nevertheless, for reasons poorly understood, the frequency of unreduced gamete formation differs substantially among different plant species. The previously identified meiotic mutant jason (jas) in Arabidopsis thaliana forms about 60% diploid (2n) pollen. JAS is required to maintain an organelle band as a physical barrier between the two meiotic spindles, preventing previously separated chromosome groups from uniting into a single cell. In this study, we characterized the jas suppressor mutant telamon (tel) that restored the production of haploid pollen in the jas background. The tel mutant did not restore the organelle band, but enlarged the size of male jas tel meiocytes, suggesting that enlarged meiocytes can bypass the requirement of the organelle band. Consistently, enlarged meiocytes generated by a tetraploid jas mutant formed reduced gametes. The results reveal that meiocyte size impacts chromosome segregation in meiosis II, suggesting an alternative way to maintain the ploidy stability in meiosis during evolution.

Polymerase IV Plays a Crucial Role in Pollen Development in Capsella.

In Arabidopsis (Arabidopsis thaliana), DNA-dependent RNA polymerase IV (Pol IV) is required for the formation of transposable element (TE)-derived small RNA transcripts. These transcripts are processed by DICER-LIKE3 into 24-nucleotide small interfering RNAs (siRNAs) that guide RNA-directed DNA methylation. In the pollen grain, Pol IV is also required for the accumulation of 21/22-nucleotide epigenetically activated siRNAs, which likely silence TEs via post-transcriptional mechanisms. Despite this proposed role of Pol IV, its loss of function in Arabidopsis does not cause a discernible pollen defect. Here, we show that the knockout of NRPD1, encoding the largest subunit of Pol IV, in the Brassicaceae species Capsella (Capsella rubella), caused postmeiotic arrest of pollen development at the microspore stage. As in Arabidopsis, all TE-derived siRNAs were depleted in Capsella nrpd1 microspores. In the wild-type background, the same TEs produced 21/22-nucleotide and 24-nucleotide siRNAs; these processes required Pol IV activity. Arrest of Capsella nrpd1 microspores was accompanied by the deregulation of genes targeted by Pol IV-dependent siRNAs. TEs were much closer to genes in Capsella compared with Arabidopsis, perhaps explaining the essential role of Pol IV in pollen development in Capsella. Our discovery that Pol IV is functionally required in Capsella microspores emphasizes the relevance of investigating different plant models.

INT-Hi-C reveals distinct chromatin architecture in endosperm and leaf tissues of Arabidopsis.

Higher-order chromatin structure undergoes striking changes in response to various developmental and environmental signals, causing distinct cell types to adopt specific chromatin organization. High throughput chromatin conformation capture (Hi-C) allows studying higher-order chromatin structure; however, this technique requires substantial amounts of starting material, which has limited the establishment of cell type-specific higher-order chromatin structure in plants. To overcome this limitation, we established a protocol that is applicable to a limited amount of nuclei by combining the INTACT (isolation of nuclei tagged in specific cell types) method and Hi-C (INT-Hi-C). Using this INT-Hi-C protocol, we generated Hi-C data from INTACT purified endosperm and leaf nuclei. Our INT-Hi-C data from leaf accurately reiterated chromatin interaction patterns derived from conventional leaf Hi-C data. We found that the higher-order chromatin organization of mixed leaf tissues and endosperm differs and that DNA methylation and repressive histone marks positively correlate with the chromatin compaction level. We furthermore found that self-looped interacting genes have increased expression in leaves and endosperm and that interacting intergenic regions negatively impact on gene expression in the endosperm. Last, we identified several imprinted genes involved in long-range and trans interactions exclusively in endosperm. Our study provides evidence that the endosperm adopts a distinct higher-order chromatin structure that differs from other cell types in plants and that chromatin interactions influence transcriptional activity.

Polycomb Repressive Complex 2 and KRYPTONITE regulate pathogen-induced programmed cell death in Arabidopsis.

The Polycomb Repressive Complex 2 (PRC2) is well-known for its role in controlling developmental transitions by suppressing the premature expression of key developmental regulators. Previous work revealed that PRC2 also controls the onset of senescence, a form of developmental programmed cell death (PCD) in plants. Whether the induction of PCD in response to stress is similarly suppressed by the PRC2 remained largely unknown. In this study, we explored whether PCD triggered in response to immunity- and disease-promoting pathogen effectors is associated with changes in the distribution of the PRC2-mediated histone H3 lysine 27 trimethylation (H3K27me3) modification in Arabidopsis thaliana. We furthermore tested the distribution of the heterochromatic histone mark H3K9me2, which is established, to a large extent, by the H3K9 methyltransferase KRYPTONITE, and occupies chromatin regions generally not targeted by PRC2. We report that effector-induced PCD caused major changes in the distribution of both repressive epigenetic modifications and that both modifications have a regulatory role and impact on the onset of PCD during pathogen infection. Our work highlights that the transition to pathogen-induced PCD is epigenetically controlled, revealing striking similarities to developmental PCD.

Brachycephaly in French bulldogs and pugs is associated with narrow ear canals.

BACKGROUND: Brachycephalic dog breeds have multiple skull malformations which may lead to anatomical changes in the external auditory canal. It is our frequent observation that in the otoscopic examination of the external ear in these breeds we are unable to visualise the tympanic membrane as a consequence of extreme narrowing of the proximal ear canal. Additionally brachycephalic dogs reportedly are predisposed to otitis externa (OE) and otitis media. OBJECTIVES: To characterizse the transition of the cartilaginous ear canal to the bony meatus acusticus externus using computed tomography (CT) and to investigate a possible association with OE in brachycephalic dogs. MATERIALS AND METHODS: Seventy-five client-owned dogs [pugs (n = 20), French bulldogs (n = 55)] were included and assessed for OE using an owner questionnaire and otoscopic and cytological examinations. In dorsal plane CT scans, the diameter of the porus acusticus externus was measured using novel methodology. The results were compared with a normocephalic control group without preexisting otological disorders. RESULTS: Brachycephalic dogs had a significantly smaller porus acusticus externus diameter (2.6 mm) than normocephalic dogs (5.0 mm). Of the brachycephalic dogs, 32% had OE yet this was not statistically significantly related to the diameter of the porus acusticus externus. Middle ear effusion (44%) and narrowing of the external ear canal (82.6%) were significantly more frequent in brachycephalic dogs. Only five of 150 eardrums could be visualised otoscopically. CONCLUSIONS AND CLINICAL RELEVANCE: Malformation of the porus acusticus externus causes severe stenosis of the proximal ear canal in brachycephalic dogs. A connection between stenosis of the external auditory canal and OE could not be confirmed.

Contexte - Les races de chiens brachycéphales présentent de multiples malformations crâniennes qui peuvent entraîner des modifications anatomiques du conduit auditif externe. Nous observons fréquemment que lors de l'examen otoscopique de l'oreille externe chez ces races, nous sommes incapables de visualiser la membrane tympanique en raison d'un rétrécissement extrême du conduit auditif proximal. De plus, les chiens brachycéphales seraient prédisposés à l'otite externe (OE) et à l'otite moyenne. Objectifs - Caractériser la transition du conduit auditif cartilagineux au méat osseux externe par tomodensitométrie (TDM) et étudier une éventuelle association avec l'OE chez les chiens brachycéphales. Matériels et méthodes - Soixante-quinze chiens de propriétaires [carlins (n = 20), bouledogues français (n = 55)] ont été inclus et évalués pour l'OE à l'aide d'un questionnaire du propriétaire et d'examens otoscopiques et cytologiques. Dans les tomodensitogrammes du plan dorsal, le diamètre du porus acusticus externus a été mesuré à l'aide d'une nouvelle méthodologie. Les résultats ont été comparés à un groupe témoin normocéphale sans trouble otologique préexistant. Résultats - Les chiens brachycéphales avaient un diamètre de porus acusticus externus significativement plus petit (2,6 mm) que les chiens normocéphales (5,0 mm). Parmi les chiens brachycéphales, 32 % avaient une OE, mais cela n'était pas statistiquement lié de manière significative au diamètre du porus acusticus externus. L'épanchement de l'oreille moyenne (44 %) et le rétrécissement du conduit auditif externe (82,6 %) étaient significativement plus fréquents chez les chiens brachycéphales. Seuls cinq des 150 tympans ont pu être visualisés par otoscopie. Conclusions et pertinence clinique - La malformation du porus acusticus externus provoque une sténose sévère du conduit auditif proximal chez le chien brachycéphale. Un lien entre la sténose du conduit auditif externe et l'OE n'a pas pu être confirmé.

Contexto - Raças de cães braquicefálicos apresentam múltiplas malformações cranianas que podem levar a alterações anatômicas no conduto auditivo externo. Frequentemente, observamos que na avaliação otoscópica da orelha externa destas raças não conseguimos visualizar a membrana timpânica como uma consequência do estreitamento extremo do conduto auditivo proximal. Além disso, cães braquicefálicos são sabidamente predispostos à otite externa (OE) e otite média. Objetivos - Caracterizar a transição do conduto auditivo cartilaginoso para o meato acústico ósseo externo utilizando tomografia computadorizada (TC) e investigar uma possível associação com OE em cães braquicefálicos. Materiais e métodos - Setenta cães de clientes [pugs (n=20), buldogues franceses (n = 55)] foram inclusos e avaliados para OE utilizando um questionário para os proprietários, citologia e otoscopia. Nas TCs em plano dorsal, o diâmetro do poro acústico externo foi mensurado utilizando uma metodologia nova. Os resultados foram comparados com um grupo de cães normocefálicos controle sem alterações otológicas pré-existentes. Resultados - Os cães braquicefálicos apresentaram o diâmetro do poro acústico externo (2,6mm) significativamente menor que os cães normocefálicos (5,0mm). Dos cães braquicefálicos, 32% apresentavam OE, mas não houve relação significativa com o diâmetro do poro acústico externo. Presença de efusão na orelha média (44%) e estreitamento do conduto auditivo externo (82,6%) foram significativamente mais frequentes em cães braquicefálicos. Apenas cinco de 150 tímpanos puderam ser visualizados por otoscopia. Conclusões e relevância clínica - Malformações do poro acústico externo causa estenose grave do conduto auditivo proximal em cães braquicefálicos. Não foi possível comprovar a conexão entre a estenose do conduto auditivo externo e OE.

Introducción- las razas de perros braquicéfalos tienen múltiples malformaciones craneales que pueden provocar cambios anatómicos en el conducto auditivo externo. Con frecuencia observamos que en el examen otoscópico del oído externo en estas razas no podemos visualizar la membrana timpánica como consecuencia del estrechamiento extremo del conducto auditivo proximal. Además, según la literatura al respecto, los perros braquicéfalos están predispuestos a padecer otitis externa (OE) y otitis media. Objetivos - Caracterizar la transición del canal auditivo cartilaginoso al meato acusticus externus óseo mediante tomografía computerizada (CT) e investigar una posible asociación con OE en perros braquicefálicos. Materiales y métodos- setenta y cinco perros de propietarios particulares [carlinos (n = 20), bulldogs franceses (n = 55)] fueron incluidos y evaluados por posible OE mediante un cuestionario para propietarios y exámenes otoscópicos y citológicos. En tomografías computerizadas del plano dorsal, el diámetro del porus acusticus externus se midió utilizando una metodología novedosa. Los resultados se compararon con un grupo control normocefálico sin trastornos otológicos preexistentes. Resultados- los perros braquicefálicos tenían un diámetro del porus acusticus externus significativamente más pequeño (2,6 mm) que los perros normocefálicos (5,0 mm). De los perros braquicefálicos, el 32 % tenía OE, pero esto no se relacionó estadísticamente de manera significativa con el diámetro del porus acusticus externus. La efusión del oído medio (44 %) y el estrechamiento del conducto auditivo externo (82,6 %) fueron significativamente más frecuentes en perros braquicefálicos. Solo cinco de 150 tímpanos pudieron visualizarse por otoscopia. Conclusiones y relevancia clínica - La malformación del porus acusticus externus causa estenosis severa del canal auditivo proximal en perros braquicefálicos. No se pudo confirmar una conexión entre la estenosis del conducto auditivo externo y la OE.

MRI diagnosis of spontaneous intraventricular tension-pneumocephalus in a 10-month-old male Saarloos Wolfdog.

A 10-month-old male Saarloos Wolfdog was presented with a history of multiple neurologic deficits that had acutely progressed. Neurologic examination findings localized signs to the cerebrum and brainstem. Magnetic resonance imaging revealed markedly enlarged and gas-filled lateral ventricles with a mass effect leading to cerebellar herniation. A right-sided defect of the cribriform plate with a dysplastic ethmoturbinate was identified as the inlet of air and origin of the intraventricular tension pneumocephalus. Surgical findings were consistent with a ruptured, congenital, nasal meningocele.

Role of H1 and DNA methylation in selective regulation of transposable elements during heat stress.

Heat-stressed Arabidopsis plants release heterochromatin-associated transposable element (TE) silencing, yet it is not accompanied by major reductions of epigenetic repressive modifications. In this study, we explored the functional role of histone H1 in repressing heterochromatic TEs in response to heat stress. We generated and analyzed RNA and bisulfite-sequencing data of wild-type and h1 mutant seedlings before and after heat stress. Loss of H1 caused activation of pericentromeric Gypsy elements upon heat treatment, despite these elements remaining highly methylated. By contrast, nonpericentromeric Copia elements became activated concomitantly with loss of DNA methylation. The same Copia elements became activated in heat-treated chromomethylase 2 (cmt2) mutants, indicating that H1 represses Copia elements through maintaining DNA methylation under heat. We discovered that H1 is required for TE repression in response to heat stress, but its functional role differs depending on TE location. Strikingly, H1-deficient plants treated with the DNA methyltransferase inhibitor zebularine were highly tolerant to heat stress, suggesting that both H1 and DNA methylation redundantly suppress the plant response to heat stress.