Rye (Secale cereale) supernumerary (B) chromosomes associated with heat tolerance during early stages of male sporogenesis.

BACKGROUND AND AIMS: Rye supernumerary (B) chromosomes have an accumulation mechanism involving the B subtelomeric domain highly enriched in D1100- and E3900-related sequences. In this work, the effects of heat stress during the early stages of male meiosis in 0B and +B plants were studied. METHODS: In-depth cytological analyses of chromatin structure and behaviour were performed on staged rye meiocytes utilizing DAPI, fluorescence in situ hybridization and 5-methylcytosine immune labelling. Quantitative real-time PCR was used to measure heat effects on the expression of the Hsp101 gene as well as the 3·9- and 2·7-kb E3900 forms in various tissues and meiotic stages. KEY RESULTS AND CONCLUSIONS: Quantitative real-time PCR established that heat induced equal up-regulation of the Hsp101 gene in 0B and 2B plants, with a marked peak in anthers with meiocytes staged at pachytene. Heat also resulted in significant up-regulation of E3900-related transcripts, especially at pachytene and for the truncated 2·7-kb form of E3900. Cytological heat-induced anomalies in prophase I, measured as the frequency of anomalous meiocytes, were significantly greater in 0B plants. Whereas telomeric sequences were widely distributed in a manner close to normal in the majority of 2B pachytene cells, most 0B meiocytes displayed abnormally clustered telomeres after chromosome pairing had occurred. Relevantly, bioinformatic analysis revealed a significantly high-density heat responsive cis regulatory sequence on E3900, clearly supporting stress-induced response of transcription for the truncated variant. Taken together, these results are the first indication that rye B chromosomes have implications on heat tolerance and may protect meiocytes against heat stress-induced damage.

Multimegabase silencing in nucleolar dominance involves siRNA-directed DNA methylation and specific methylcytosine-binding proteins.

In genetic hybrids, the silencing of nucleolar rRNA genes inherited from one progenitor is the epigenetic phenomenon known as nucleolar dominance. An RNAi knockdown screen identified the Arabidopsis de novo cytosine methyltransferase, DRM2, and the methylcytosine binding domain proteins, MBD6 and MBD10, as activities required for nucleolar dominance. MBD10 localizes throughout the nucleus, but MBD6 preferentially associates with silenced rRNA genes and does so in a DRM2-dependent manner. DRM2 methylation is thought to be guided by siRNAs whose biogenesis requires RNA-DEPENDENT RNA POLYMERASE 2 (RDR2) and DICER-LIKE 3 (DCL3). Consistent with this hypothesis, knockdown of DCL3 or RDR2 disrupts nucleolar dominance. Collectively, these results indicate that in addition to directing the silencing of retrotransposons and noncoding repeats, siRNAs specify de novo cytosine methylation patterns that are recognized by MBD6 and MBD10 in the large-scale silencing of rRNA gene loci.

Use of Repetitive Sequences for Molecular and Cytogenetic Characterization of Avena Species from Portugal.

Genomic diversity of Portuguese accessions of Avena species--diploid A. strigosa and hexaploids A. sativa and A. sterilis--was evaluated through molecular and cytological analysis of 45S rDNA, and other repetitive sequences previously studied in cereal species--rye subtelomeric sequence (pSc200) and cereal centromeric sequence (CCS1). Additionally, retrotransposons and microsatellites targeting methodologies--IRAP (inter-retrotransposon amplified polymorphism) and REMAP (retrotransposon-microsatellite amplified polymorphism)--were performed. A very high homology was detected for ribosomal internal transcribed sequences (ITS1 and ITS2) between the species analyzed, although nucleolar organizing regions (NOR) fluorescent in situ hybridization (FISH) analysis revealed distinct number of Nor loci between diploid and hexaploid species. Moreover, morphological diversity, evidenced by FISH signals with different sizes, was observed between distinct accessions within each species. pSc200 sequences were for the first time isolated from Avena species but proven to be highly similar in all genotypes analyzed. The use of primers designed for CCS1 unraveled a sequence homologous to the Ty3/gypsy retrotransposon Cereba, that was mapped to centromeric regions of diploid and hexaploid species, being however restricted to the more related A and D haplomes. Retrotransposon-based methodologies disclosed species- and accessions-specific bands essential for the accurate discrimination of all genotypes studied. Centromeric, IRAP and REMAP profiles therefore allowed accurate assessment of inter and intraspecific variability, demonstrating the potential of these molecular markers on future oat breeding programs.

Cytotoxicity of Eupatorium cannabinum L. ethanolic extract against colon cancer cells and interactions with Bisphenol A and Doxorubicin.

BACKGROUND: Eupatorium cannabinum L. has long been utilized in traditional medicine, however no information is available regarding cellular effects of full extracts. Here we assessed the effects of E. cannabinum ethanolic extract (EcEE) on the colon cancer line HT29. Potential interactions with bisphenol A (BPA) a synthetic phenolic compound to which humans are generally exposed and a commonly used chemotherapeutic agent, doxorubicin (DOX) were also evaluated. METHODS: HT29 cells were exposed to different concentrations (0.5 to 50 µg/ml) of EcEE alone or in combination with BPA or DOX. Cell viability was analyzed through resazurin assay. Gene transcription levels for NCL, FOS, p21, AURKA and bcl-xl were determined through qRT-PCR. Cytological analysis included evaluation of nuclear and mitotic anomalies after DAPI staining, immunodetection of histone H3 lysine 9 acetylation (H3K9ac) and assessment of DNA damage by TUNEL assay. RESULTS: Severe loss of HT29 cell viability was detected for 50 µg/ml EcEE immediately after 24 h exposure whereas the lower concentrations assayed (0.5, 5 and 25 µg/ml) resulted in significant viability decreases after 96 h. Exposure to 25 µg/ml EcEE for 48 h resulted in irreversible cell damage leading to a drastic decrease in cell viability after 72 h recovery in EcEE-free medium. 48 h 25 µg/ml EcEE treatment also induced alteration of colony morphology, H3K9 hyperacetylation, transcriptional up regulation of p21 and down regulation of NCL, FOS and AURKA, indicating reduced proliferation capacity. This treatment also resulted in drastic mitotic and nuclear disruption accompanied by up-regulation of bcl-xl, limited TUNEL labeling and nuclear size increase, suggestive of a non-apoptocic cell death pathway. EcEE/BPA co-exposure increased mitotic anomalies particularly for the lowest EcEE concentration, although without major effects on viability. Conversely, EcEE/DOX co-exposure decreased cell viability in relation to DOX for all EcEE concentrations, without affecting the DOX-induced cell cycle arrest. CONCLUSIONS: EcEE has cytotoxic activity on HT29 cancer cells leading to mitotic disruption and non-apoptotic cell death without severe induction of DNA damage. Interaction experiments showed that EcEE can increase BPA aneugenic effects and EcEE synergistic effects with DOX supporting a potential use as adjuvant in chemotherapeutic approaches.

Bisphenol A disrupts transcription and decreases viability in aging vascular endothelial cells.

Bisphenol A (BPA) is a widely utilized endocrine disruptor capable of mimicking endogenous hormones, employed in the manufacture of numerous consumer products, thereby interfering with physiological cellular functions. Recent research has shown that BPA alters epigenetic cellular mechanisms in mammals and may be correlated to enhanced cellular senescence. Here, the effects of BPA at 10 ng/mL and 1 µg/mL, concentrations found in human samples, were analyzed on HT29 human colon adenocarcinona cell line and Human Umbilical Vein Endothelial Cells (HUVEC). Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) transcriptional analysis of the Long Interspersed Element-1 (LINE-1) retroelement showed that BPA induces global transcription deregulation in both cell lines, although with more pronounced effects in HUVEC cells. Whereas there was an increase in global transcription in HT29 exclusively after 24 h of exposure, this chemical had prolonged effects on HUVEC. Immunoblotting revealed that this was not accompanied by alterations in the overall content of H3K9me2 and H3K4me3 epigenetic marks. Importantly, cell viability assays and transcriptional analysis indicated that prolonged BPA exposure affects aging processes in senescent HUVEC. To our knowledge this is the first report that BPA interferes with senescence in primary vascular endothelial cells, therefore, suggesting its association to the etiology of age-related human pathologies, such as atherosclerosis.

Bisphenol A at concentrations found in human serum induces aneugenic effects in endothelial cells.

Bisphenol A (BPA) is an endocrine disrupting chemical to which humans are exposed. Continuous environmental exposure to BPA leads to its detection in the majority of individuals from developed countries, with serum concentrations ranging from 0.5 to 10ng/ml in the general population and much higher when associated with occupational exposure. In this work, human umbilical vascular endothelial cells (HUVEC) and human colon adenocarcinona (HT29) cell lines were used to represent endothelial and digestive-tract tissues, which are in direct contact to BPA in vivo. Our results demonstrate that BPA has cell-type differential effects. Notably, BPA concentrations commonly found in humans induce micronucleus formation and interfere with cell-division processes in endothelial cells, resulting in mitotic abnormalities. We also found that BPA induces up-regulation of two genes encoding proteins associated with chromosome segregation, namely CDCA8 (borealin/cell division cycle A8) and SGOL2 (shugoshin-like2). Taken together, the aneugenic effects observed in endothelial cells (HUVECs) substantiate increasing concerns about BPA exposure at levels currently detected in humans.

Dynamic Regulation of Grapevine's microRNAs in Response to Mycorrhizal Symbiosis and High Temperature.

MicroRNAs (miRNAs) are non-coding small RNAs that play crucial roles in plant development and stress responses and can regulate plant interactions with beneficial soil microorganisms such as arbuscular mycorrhizal fungi (AMF). To determine if root inoculation with distinct AMF species affected miRNA expression in grapevines subjected to high temperatures, RNA-seq was conducted in leaves of grapevines inoculated with either Rhizoglomus irregulare or Funneliformis mosseae and exposed to a high-temperature treatment (HTT) of 40 °C for 4 h per day for one week. Our results showed that mycorrhizal inoculation resulted in a better plant physiological response to HTT. Amongst the 195 identified miRNAs, 83 were considered isomiRs, suggesting that isomiRs can be biologically functional in plants. The number of differentially expressed miRNAs between temperatures was higher in mycorrhizal (28) than in non-inoculated plants (17). Several miR396 family members, which target homeobox-leucine zipper proteins, were only upregulated by HTT in mycorrhizal plants. Predicted targets of HTT-induced miRNAs in mycorrhizal plants queried to STRING DB formed networks for Cox complex, and growth and stress-related transcription factors such as SQUAMOSA promoter-binding-like-proteins, homeobox-leucine zipper proteins and auxin receptors. A further cluster related to DNA polymerase was found in R. irregulare inoculated plants. The results presented herein provide new insights into miRNA regulation in mycorrhizal grapevines under heat stress and can be the basis for functional studies of plant-AMF-stress interactions.

Mediterranean diet adherence and nutritional literacy: an observational cross-sectional study of the reality of university students in a COVID-19 pandemic context.

Aim: To evaluate the adherence to the Mediterranean diet (MD) and the level of nutritional literacy (NL) among university students from different academic fields of study, within the context of the COVID-19 pandemic. Methods: A total of 1114 first-year undergraduate students at the University of Lisbon, Portugal, were included in this study. A self-administered online questionnaire was applied that included questions regarding sociodemographic information, the MD measured by the PREDIMED questionnaire (PREvención con DIeta MEDiterránea) and NL assessed using the Newest Vital Sign questionnaire. Results: The average PREDIMED score revealed a low adherence (6.79±2.14 points) to the MD. Notably, students in the Social Sciences and Humanities academic fields showed the highest level of adherence (U=21 071; p<0.05). Within the Health field, there was a greater prevalence of dietary behaviours aligned with the MD, contributing to higher overall adherence scores. Furthermore, 84.1% of the participants demonstrated adequate NL. Interestingly, students in the Exact Sciences and Engineering field demonstrated the highest levels of NL (5.07±1.19), particularly in questions involving mathematical reasoning. Conclusions: Our findings suggest that university students in Lisbon do not follow a MD and are far from the recommendations of this dietary pattern. While most participants showed adequate NL, it is essential to highlight the link between knowledge and application to daily practice. Despite positive literacy levels, there remains a deficit in translating this knowledge into correct dietary practices.

Involvement of disperse repetitive sequences in wheat/rye genome adjustment.

The union of different genomes in the same nucleus frequently results in hybrid genotypes with improved genome plasticity related to both genome remodeling events and changes in gene expression. Most modern cereal crops are polyploid species. Triticale, synthesized by the cross between wheat and rye, constitutes an excellent model to study polyploidization functional implications. We intend to attain a deeper knowledge of dispersed repetitive sequence involvement in parental genome reshuffle in triticale and in wheat-rye addition lines that have the entire wheat genome plus each rye chromosome pair. Through Random Amplified Polymorphic DNA (RAPD) analysis with OPH20 10-mer primer we unraveled clear alterations corresponding to the loss of specific bands from both parental genomes. Moreover, the sequential nature of those events was revealed by the increased absence of rye-origin bands in wheat-rye addition lines in comparison with triticale. Remodeled band sequencing revealed that both repetitive and coding genome domains are affected in wheat-rye hybrid genotypes. Additionally, the amplification and sequencing of pSc20H internal segments showed that the disappearance of parental bands may result from restricted sequence alterations and unraveled the involvement of wheat/rye related repetitive sequences in genome adjustment needed for hybrid plant stabilization.

Grain Transcriptome Dynamics Induced by Heat in Commercial and Traditional Bread Wheat Genotypes.

High temperature (HT) events have negative impact on wheat grains yield and quality. Transcriptome profiles of wheat developing grains of commercial genotypes (Antequera and Bancal) and landraces (Ardito and Magueija) submitted to heatwave-like treatments during grain filling were evaluated. Landraces showed significantly more differentially expressed genes (DEGs) and presented more similar responses than commercial genotypes. DEGs were more associated with transcription and RNA and protein synthesis in Antequera and with metabolism alterations in Bancal and landraces. Landraces upregulated genes encoding proteins already described as HT responsive, like heat shock proteins and cupins. Apart from the genes encoding HSP, two other genes were upregulated in all genotypes, one encoding for Adenylate kinase, essential for the cellular homeostasis, and the other for ferritin, recently related with increased tolerance to several abiotic stress in Arabidopsis. Moreover, a NAC transcription factor involved in plant development, known to be a negative regulator of starch synthesis and grain yield, was found to be upregulated in both commercial varieties and downregulated in Magueija landrace. The detected diversity of molecular processes involved in heat response of commercial and traditional genotypes contribute to understand the importance of genetic diversity and relevant pathways to cope with these extreme events.

COVID-19 and Lockdown, as Lived and Felt by University Students.

In the last 2 years, the COVID-19 pandemic has spread all over the world, forcing the closure of universities, among other unusual measures in recent history. (1) Background: This work is based on the study HOUSE-ULisbon, a survey carried out during the second confinement (March-May 2021) in Portugal with the collaboration of all the Faculties of the University of Lisbon (UL). The present work aims to explore gender differences in how first-year college students experienced and felt COVID-19 and the second confinement. (2) Methods: A questionnaire was carried out. In total, 976 university students (19.66 years (SD = 4.033); Min = 17 and Max = 65) from the first year of the UL were included, of which 69.5% (n = 678) were female, and 30.5% were male (n = 298). SPSS v. 26 was used for quantitative data and MAXQDA 2020 for qualitative data. (3) Results: Overall, students reported various symptoms of physical and mental discomfort (especially females). Statistically significant differences were found in the problems that could arise from the pandemic, such as the prevalence of higher anxiety and worries by females, and online gaming by males. In coping strategies, differences were found in leisure and family relationships, with greater difficulty on the female side. Social interaction was perceived as difficult or very difficult by both genders. As strategies for future pandemics, they highlighted a concerted effort between the government and media in the transmission of messages to the population, facilitating information, knowledge and adoption of protective behaviors. (4) Conclusions: These results are important data for activating or maintaining resources and services for first-year university students, who in some university institutions were supported during the pandemic by psychological, material (e.g., computers, internet), and financial support measures, which are now diminished or extinct. The impacts on their lives will certainly not be extinguished post-pandemic, and health, education, and public policy measures should be prioritized for this group. These results are important data for activating resources and services for students, informing health and education professionals, and supporting public policies.

Chronic Conditions and School Participation of First-Year University Students-HOUSE ULisbon Study.

Students with chronic conditions (CC) tend to experience several barriers in terms of their school participation and performance. Therefore, the present study aims to explore the factors related to the time of diagnosis of CC (recent/non-recent), the barriers to participation and academic success (health condition, people's attitude towards CC and school physical environment), the physical and mental health (physical/psychological symptoms and concerns) and school-related variables (relationship with teachers and peers), regarding the school participation of first-year students with CC. This work is part of the HOUSE-Colégio F3 Project, University of Lisbon, which includes 1143 first-year university students from 17 Faculties and Institutes of the University of Lisbon. In this specific study, only the subsample of 207 students with CC was considered, 72.4% of which were female, aged between 18 and 54 years (M = 20.00; SD = 4.83). The results showed that students with a recent diagnosis of CC and students with school participation affected by the CC were those who presented more negative indicators regarding barriers to school participation, physical and mental health, and school-related variables. A greater impact of CC in terms of school participation was associated with having a recent diagnosis, with people's attitude towards CC and with the health condition as barriers, with more psychological symptoms and worse relationships with teachers and peers. This is a relevant message for the organization of health services for students with CC at the beginning of their university studies, especially since they are often displaced from home and managing their health conditions alone (in many cases, for the first time).

Size matters in Triticeae polyploids: larger genomes have higher remodeling.

Polyploidization is one of the major driving forces in plant evolution and is extremely relevant to speciation and diversity creation. Polyploidization leads to a myriad of genetic and epigenetic alterations that ultimately generate plants and species with increased genome plasticity. Polyploids are the result of the fusion of two or more genomes into the same nucleus and can be classified as allopolyploids (different genomes) or autopolyploids (same genome). Triticeae synthetic allopolyploid species are excellent models to study polyploids evolution, particularly the wheat-rye hybrid triticale, which includes various ploidy levels and genome combinations. In this review, we reanalyze data concerning genomic analysis of octoploid and hexaploid triticale and different synthetic wheat hybrids, in comparison with other polyploid species. This analysis reveals high levels of genomic restructuring events in triticale and wheat hybrids, namely major parental band disappearance and the appearance of novel bands. Furthermore, the data shows that restructuring depends on parental genomes, ploidy level, and sequence type (repetitive, low copy, and (or) coding); is markedly different after wide hybridization or genome doubling; and affects preferentially the larger parental genome. The shared role of genetic and epigenetic modifications in parental genome size homogenization, diploidization establishment, and stabilization of polyploid species is discussed.

Rye Bs disclose ancestral sequences in cereal genomes with a potential role in gametophyte chromatid segregation.

Two sequence families, E3900 and D1100, are amplified on the subtelomeric domain of the long arm of rye B chromosomes, the region that controls its drive mechanism. In this work, polymerase chain reaction (PCR) with a number of primers spanning E3900 shows that the organization and nucleotide sequence of E3900-related portions are present and highly conserved on rye A chromosomes as well as in other cereals. Quantitative Real-Time PCR estimates two E3900 sequences to be represented in 100-150 copies on Bs and at least as single copies on As. A novel E3900-related sequence, with a deletion that results in a frameshift and subsequently an open reading frame with putative DNA binding motifs, is identified. Expression analysis of E3900 indicates identical transcription levels in leaves from plants with and without Bs, showing that the expression of these sequences must be silenced on Bs and tightly regulated on As in leaves. In contrast, E3900 transcription is upregulated during meiosis exclusively in plants with Bs, maintaining a high level of transcription in the gametophyte. Interestingly, Bs not only influence their own chromatid segregation but also that of the regular chromosome complement of both rye and wheat. There is a drastic increase in frequency of disrupted metaphase and anaphase cells in the first mitosis of pollen grains carrying Bs, which appears to be due to anomalous adherences between sister chromatids. Taken together, this work provides insight into how E3900 sequences are potentially associated with important evolutionary mechanisms involved in basic cellular processes.

Genome merger: from sequence rearrangements in triticale to their elimination in wheat-rye addition lines.

Genetic and epigenetic modifications resulting from different genomes adjusting to a common nuclear environment have been observed in polyploids. Sequence restructuring within genomes involving retrotransposon/microsatellite-rich regions has been reported in triticale. The present study uses inter-retrotransposon amplified polymorphisms (IRAP) and retrotransposon microsatellite amplified polymorphisms (REMAP) to assess genome rearrangements in wheat-rye addition lines obtained by the controlled backcrossing of octoploid triticale to hexaploid wheat followed by self-fertilization. The comparative analysis of IRAP and REMAP banding profiles, involving a complete set of wheat-rye addition lines, and their parental species revealed in those lines the presence of wheat-origin bands absent in triticale, and the absence of rye-origin and triticale-specific bands. The presence in triticale x wheat backcrosses (BC) of rye-origin bands that were absent in the addition lines demonstrated that genomic rearrangement events were not a direct consequence of backcrossing, but resulted from further genome structural rearrangements in the BC plant progeny. PCR experiments using primers designed from different rye-origin sequences showed that the absence of a rye-origin band in wheat-rye addition lines results from sequence elimination rather than restrict changes on primer annealing sites, as noted in triticale. The level of genome restructuring events evaluated in all seven wheat-rye addition lines, compared to triticale, indicated that the unbalanced genome merger situation observed in the addition lines induced a new round of genome rearrangement, suggesting that the lesser the amount of rye chromatin introgressed into wheat the larger the outcome of genome reshuffling.

Chromosome and DNA methylation dynamics during meiosis in the autotetraploid Arabidopsis arenosa.

Variation in chromosome number due to polyploidy can seriously compromise meiotic stability. In autopolyploids, the presence of more than two homologous chromosomes may result in complex pairing patterns and subsequent anomalous chromosome segregation. In this context, chromocenter, centromeric, telomeric and ribosomal DNA locus topology and DNA methylation patterns were investigated in the natural autotetraploid, Arabidopsis arenosa. The data show that homologous chromosome recognition and association initiates at telomeric domains in premeiotic interphase, followed by quadrivalent pairing of ribosomal 45S RNA gene loci (known as NORs) at leptotene. On the other hand, centromeric regions at early leptotene show pairwise associations rather than associations in fours. These pairwise associations are maintained throughout prophase I, and therefore likely to be related to the diploid-like behavior of A. arenosa chromosomes at metaphase I, where only bivalents are observed. In anthers, both cells at somatic interphase as well as at premeiotic interphase show 5-methylcytosine (5-mC) dispersed throughout the nucleus, contrasting with a preferential co-localization with chromocenters observed in vegetative nuclei. These results show for the first time that nuclear distribution patterns of 5-mC are simultaneously reshuffled in meiocytes and anther somatic cells. During prophase I, 5-mC is detected in extended chromatin fibers and chromocenters but interestingly is excluded from the NORs what correlates with the pairing pattern.

Assessment of Four Portuguese Wheat Landrace Diversity to Cope With Global Warming.

Wheat is a dietary staple consumed worldwide strongly responsible for proteins and carbohydrate population intake. However, wheat production and quality will scarcely fulfill forward demands, which are compounded by high-temperature (HT) events as heatwaves, increasingly common in Portugal. Thus, landraces assume crucial importance as potential reservoirs of useful traits for wheat breeding and may be pre-adapted to extreme environmental conditions. This work evaluates four Portuguese landrace yield and grain composition through attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, particularly protein content, and their responses to HT treatment mimicking a heatwave. Landraces showed distinct yield traits, especially plant height and first spike grain number, and a similar pattern in FTIR spectra, although revealing differences in grain components' proportions. Comparison between spectra band intensity indicates that Ardito has the highest protein-related peaks, contrary to Magueija, which appears to be the landrace with higher lipid content. In plants submitted to 1 week of HT treatment 10 days after anthesis, the first spike grain size and weight were markedly reduced in all landraces. Additionally, it was observed that a general increase in grain protein content in the four landraces, being the increment observed in Ardito and Grécia, is statistically significant. The comparative assessment of control and HT average FTIR spectra denoted also the occurrence of alterations in grain polysaccharide composition. An integrated assessment of the evaluations performed revealed that Ardito and Magueija landraces presented diverse yield-related characteristics and distinct responses to cope with HT. In fact, the former landrace revealed considerable grain yield diminution along with an increase in grain protein proportion after HT, while the latter showed a significant increase in spikes and grain number, with grain quality detriment. These results reinforce the relevance of scrutinizing old genotype diversity seeking for useful characteristics, particularly considering HT impact on grain production and quality.

Response of Mycorrhizal 'Touriga Nacional' Variety Grapevines to High Temperatures Measured by Calorespirometry and Near-Infrared Spectroscopy.

Heat stress negatively affects several physiological and biochemical processes in grapevine plants. In this work, two new methods, calorespirometry, which has been used to determine temperature adaptation in plants, and near-infrared (NIR) spectroscopy, which has been used to determine several grapevine-related traits and to discriminate among varieties, were tested to evaluate grapevine response to high temperatures. 'Touriga Nacional' variety grapevines, inoculated or not with Rhizoglomus irregulare or Funneliformis mosseae, were used in this study. Calorespirometric parameters and NIR spectra, as well as other parameters commonly used to assess heat injury in plants, were measured before and after high temperature exposure. Growth rate and substrate carbon conversion efficiency, calculated from calorespirometric measurements, and stomatal conductance, were the most sensitive parameters for discriminating among high temperature responses of control and inoculated grapevines. The results revealed that, although this vine variety can adapt its physiology to temperatures up to 40 °C, inoculation with R. irregulare could additionally help to sustain its growth, especially after heat shocks. Therefore, the combination of calorespirometry together with gas exchange measurements is a promising strategy for screening grapevine heat tolerance under controlled conditions and has high potential to be implemented in initial phases of plant breeding programs.

Epigenetic marks in the mature pollen of Quercus suber L. (Fagaceae).

We have analysed the distribution of epigenetic marks for histone modifications at lysine residues H3 and H4, and DNA methylation, in the nuclei of mature pollen cells of the Angiosperm tree Quercus suber; a monoecious wind pollinated species with a protandrous system, and a long post-pollination period. The ultrasonic treatment developed for the isolation of pollen nuclei proved to be a fast and reliable method, preventing the interference of cell wall autofluorescence in the in situ immunolabelling assays. In contrast with previous studies on herbaceous species with short progamic phases, our results are consistent with a high level of silent (5-mC and H3K9me2) epigenetic marks on chromatin of the generative nucleus, and the prevalence of active marks (H3K9me3 and H4Kac) in the vegetative nucleus. The findings are discussed in terms of the pollination/fertilization timing strategy adopted by this plant species.

A century of B chromosomes in plants: so what?

BACKGROUND: Supernumerary B chromosomes (Bs) are a major source of intraspecific variation in nuclear DNA amounts in numerous species of plants. They favour large genomes, and create polymorphisms for DNA variation in natural populations. By studying Bs we can gain useful knowledge about the organization, function and evolution of genomes. There are also significant biological questions concerning the origin and structural organization of Bs, and the way in which these selfish elements can establish themselves by exploiting the replicative machinery of their host genome nucleus. SCOPE: It is a sine qua non that Bs originate from the A chromosomes, in a variety of ways. We can study their modes of drive and ask how it is that chromosomes which apparently lack genes can have control over their own drive process which leads to their survival in natural populations. Molecular cytogenetic studies are opening up new avenues of investigation. Population equilibria for B frequencies are determined by a balance between accumulation and harmful effects. Bs are also subject to meiotic loss due to polysomy and to elimination at meiosis as univalents. These balancing forces can be seen in the context of host/parasite interaction, based on a dissection of the genetic elements in both As and Bs (in maize) which interact to bring about a stable equilibrium, at least for a snapshot in time. CONCLUSIONS: Aside from their intrinsic enigmatic properties, B chromosomes make useful experimental tools to study genome organization. Thus far they have not been exploited for their applications, other than through the use of A-B translocations used for gene mapping in maize; but there are opportunities to use them to modulate the frequency and distribution of recombination, to diploidize allopolyploids, to study centromeres and to be developed as plant artificial chromosomes; given that they can be structurally modified and their inheritance stabilized.