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.

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.

Comparative transcriptomics of seed nourishing tissues: uncovering conserved and divergent pathways in seed plants.

The evolutionary and ecological success of spermatophytes is intrinsically linked to the seed habit, which provides a protective environment for the initial development of the new generation. This environment includes an ephemeral nourishing tissue that supports embryo growth. In gymnosperms this tissue originates from the asexual proliferation of the maternal megagametophyte, while in angiosperms it is a product of fertilization, and is called the endosperm. The emergence of these nourishing tissues is of profound evolutionary value, and they are also food staples for most of the world's population. Here, using Orthofinder to infer orthologue genes among newly generated and previously published datasets, we provide a comparative transcriptomic analysis of seed nourishing tissues from species of several angiosperm clades, including those of early diverging lineages, as well as of one gymnosperm. Our results show that, although the structure and composition of seed nourishing tissues has seen significant divergence along evolution, there are signatures that are conserved throughout the phylogeny. Conversely, we identified processes that are specific to species within the clades studied, and thus illustrate their functional divergence. With this, we aimed to provide a foundation for future studies on the evolutionary history of seed nourishing structures, as well as a resource for gene discovery in future functional studies.

BABY BOOM regulates early embryo and endosperm development.

The BABY BOOM (BBM) AINTEGUMENTA-LIKE (AIL) AP2/ERF domain transcription factor is a major regulator of plant cell totipotency, as it induces asexual embryo formation when ectopically expressed. Surprisingly, only limited information is available on the role of BBM during zygotic embryogenesis. Here we reexamined BBM expression and function in the model plant Arabidopsis thaliana (Arabidopsis) using reporter analysis and newly developed CRISPR mutants. BBM was expressed in the embryo from the zygote stage and also in the maternal (nucellus) and filial (endosperm) seed tissues. Analysis of CRISPR mutant alleles for BBM (bbm-cr) and the redundantly acting AIL gene PLETHORA2 (PLT2) (plt2-cr) uncovered individual roles for these genes in the timing of embryo progression. We also identified redundant roles for BBM and PLT2 in endosperm proliferation and cellularization and the maintenance of zygotic embryo development. Finally, we show that ectopic BBM expression in the egg cell of Arabidopsis and the dicot crops Brassica napus and Solanum lycopersicon is sufficient to bypass the fertilization requirement for embryo development. Together these results highlight roles for BBM and PLT2 in seed development and demonstrate the utility of BBM genes for engineering asexual embryo development in dicot species.

Presumed post-mortem donors: the degree of information among university students.

BACKGROUND: Organ transplantation represents the most effective and acceptable therapy for end-stage organ failure. However, its frequent practice often leads to a shortage of organs worldwide. To solve this dilemma, some countries, such as Portugal, have switched from an opt-in to an opt-out system, which has raised concerns about respect for individual autonomy. We aimed to evaluate whether young university students are aware of this opt-out system so that they can make informed, autonomous and conscious decisions, as well as to identify the factors that determine a positive attitude toward post-mortem organ donation. METHODS: An observational, cross-sectional study was developed and a questionnaire was administered to first-year students from six faculties of the University of Porto. RESULTS: Of the 841 participants, 60% were unaware that Portugal had adopted an opt-out system. Among the informed individuals, their main sources of information included social media, internet, and family. Furthermore, only 48% of all participants agreed with the current opt-out system. Female sex (p = 0.049; OR 1.393), knowledge of the law (p < 0.001; OR 4.749) and family being the primary source of information (p < 0.001; OR 2.855) were independent factors associated with a positive attitude toward post-mortem organ donation law. CONCLUSIONS: There is a significant lack of knowledge among young university students regarding the presumed post-mortem organ donation law and how it works. Female sex, having family as a primary source of information and being aware of the presumed post-mortem organ donation law are the strongest independent factors that determine a positive attitude toward the opt-out system.

Validity and reliability of a shorter version of the Geriatric Depression Scale in institutionalized older Portuguese adults.

OBJECTIVES: Depressive symptoms are common in older adults in institutional contexts; however, there is a lack of validated measures for these settings. Identifying depressive symptoms can help clinicians to manage them and to prevent or delay their complications. This study aimed to validate the Geriatric Depression Scale (GDS) in an institutionalized sample of older adults. METHOD: 493 institutionalized older people (73% women) aged 60 or over were evaluated through the GDS, the Mini International Neuropsychiatric Interview (MINI) (depression vs. no depression = 11% vs. 89%), the Geriatric Anxiety Inventory (GAI), the Positive Affect (PA) and Negative Affect (NA) Schedule, and the Satisfaction with Life Scale (SWLS). Test-retest reliability was assessed with 57 older adults. RESULTS: An 8-item version presented a Cronbach's alpha value of .87 with a single factor explaining its variance. The correlations (p < .01) attested the concurrent validity (GAI: r = .76; PA: r = -.22; AN: r = .62; SWLS: r = -.32). Test-retest reliability (6.51 months) was adequate (r = .52). ROC analysis (AUC = .82; sensitivity = 80%; specificity = 77%) and Youden index revealed a cutoff of 5/6 for the diagnosis of depression. CONCLUSION: Results support the validity and the screening capacity of a short version of GDS in institutional contexts. Short screening instruments for depressive symptoms may facilitate their identification, allowing for timely clinical interventions in institutional settings.

BRR2a Affects Flowering Time via FLC Splicing.

Several pathways control time to flowering in Arabidopsis thaliana through transcriptional and posttranscriptional gene regulation. In recent years, mRNA processing has gained interest as a critical regulator of flowering time control in plants. However, the molecular mechanisms linking RNA splicing to flowering time are not well understood. In a screen for Arabidopsis early flowering mutants we identified an allele of BRR2a. BRR2 proteins are components of the spliceosome and highly conserved in eukaryotes. Arabidopsis BRR2a is ubiquitously expressed in all analyzed tissues and involved in the processing of flowering time gene transcripts, most notably FLC. A missense mutation of threonine 895 in BRR2a caused defects in FLC splicing and greatly reduced FLC transcript levels. Reduced FLC expression increased transcription of FT and SOC1 leading to early flowering in both short and long days. Genome-wide experiments established that only a small set of introns was not correctly spliced in the brr2a mutant. Compared to control introns, retained introns were often shorter and GC-poor, had low H3K4me1 and CG methylation levels, and were often derived from genes with a high-H3K27me3-low-H3K36me3 signature. We propose that BRR2a is specifically needed for efficient splicing of a subset of introns characterized by a combination of factors including intron size, sequence and chromatin, and that FLC is most sensitive to splicing defects.

Rice phytochrome-interacting factor protein OsPIF14 represses OsDREB1B gene expression through an extended N-box and interacts preferentially with the active form of phytochrome B.

DREB1/CBF genes, known as major regulators of plant stress responses, are rapidly and transiently induced by low temperatures. Using a yeast one-hybrid screening, we identified a putative Phytochrome-Interacting bHLH Factor (OsPIF14), as binding to the OsDREB1B promoter. bHLH proteins are able to bind to hexameric E-box (CANNTG) or N-box (CACG(A/C)G) motifs, depending on transcriptional activity. We have shown that OsPIF14 binds to the OsDREB1B promoter through two N-boxes and that the flanking regions of the hexameric core are essential for protein-DNA interaction and stability. We also showed that OsPIF14 down-regulates OsDREB1B gene expression in rice protoplasts, corroborating the OsPIF14 repressor activity observed in the transactivation assays using Arabidopsis protoplasts. In addition, we showed that OsPIF14 is indeed a phytochrome interacting factor, which preferentially binds to the active form (Pfr) of rice phytochrome B. This raises the possibility that OsPIF14 activity might be modulated by light. However, we did not observe any regulation of the OsDREB1B gene expression by light under control conditions. Moreover, OsPIF14 gene expression was shown to be modulated by different treatments, such as drought, salt, cold and ABA. Interestingly, OsPIF14 showed also a specific cold-induced alternative splicing. All together, these results suggest the possibility that OsPIF14 is involved in cross-talk between light and stress signaling through interaction with the OsDREB1B promoter. Although in the absence of stress, OsDREB1B gene expression was not regulated by light, given previous reports, it remains possible that OsPIF14 has a role in light modulation of stress responses.

Helminth fauna of chiropterans in Amazonia: biological interactions between parasite and host.

Amazonia, the largest Brazilian biome, is one of the most diverse biomes around the world. Considering the Brazilian chiropteran species, 120 out of known 167 species are registered in Pará state, with 10 endemic species. Despite the high diversity of bats in Amazonia, studies on their parasites, especially on helminths, are scarce. Therefore, the present study aims to study the helminth fauna of different bats from the Pará state, Amazon biome, determine the descriptors of infection, and evaluate the host-parasite interactions, as well as evaluate differences in ecological indexes in accordance with the feeding guilds. The study was developed on 67 bats of 21 species captured in several areas of the Pará state. The animals were identified, divided into feeding guilds, and necropsied. The parasites obtained were identified and quantified. A total of 182 parasites were found in 20.89 % of the studied bats, representing nine species, as follows: Anenterotrema eduardocaballeroi, Anenterotrema liliputianum, Ochoterenatrema caballeroi, Tricholeiperia sp., Parahistiostrongylus octacanthus, Litomosoides guiterasi, Litomosoides brasiliensis, Capillariinae gen. sp., and Hymenolepididae gen. sp. Also, the results indicated that there was no impact of parasitism on host body condition and no relationship between sex and parasite intensity. In relation to the feeding guilds, the omnivores showed higher prevalence and mean intensity. Animals from regions closer to the equator tend to have greater richness in parasite species, but the present study revealed low diversity and richness in species. In conclusion, the ecological pattern observed for other animal groups, in which higher parasitic diversity are registered in lower latitudes, is not applicable to chiropterans from the study area.

Signalling events regulating seed coat development.

The evolution of seeds was a major reason for the rise of angiosperms to ecological dominance. Seeds of angiosperms are composed of three main structures: the embryo, which will give rise to the next generation; the endosperm, a nurturing tissue whose main function is to deliver nutrients from the mother plant to the embryo; and the seed coat (or testa), a tissue that is derived from the maternal integuments and which provides support and protection to the growing embryo. All three seed components need to exchange signals to ensure co-ordinated growth and development. The present review discusses the structure of the seed coat, its interaction with the endosperm, and bidirectional signalling events between endosperm and seed coat that co-ordinate growth of both tissues. Angiosperm seeds are not only of evolutionary significance, but also of major agronomic importance, demanding a thorough understanding of the events governing seed growth and development.

An epiQTL underlying asexual seed formation in Arabidopsis.

KEY MESSAGE: The DNA methylation status at an epigenetic quantitative trait locus in the Arabidopsis chromosome 2 is linked to the formation of apomictic-like endosperms. Seed development in most angiosperms is coupled to fertilization of the maternal gametes by two sperm cells. However, apomictic species can reproduce asexually via seeds. This trait is of great agricultural interest, as it would fix complex genotypes and allow for pollen-independent seed production. However, engineering full apomixis requires three independent processes: apomeiosis, parthenogenesis and autonomous endosperm development. While the first two have been successfully engineered in some crops, the formation of autonomous endosperms remains a challenge. Although it is known that this trait is under epigenetic control, such as of DNA methylation, the underlying mechanisms remain mostly undiscovered. Here, using epigenetic recombinant inbred lines, we identified an epigenetic quantitative trait locus in the Arabidopsis chromosome 2, which correlates with permissiveness for the formation of asexual seeds: hypomethylation at this genomic region allows the formation of larger autonomous endosperms. Importantly, the methylation at this locus only correlates with asexual seed size, and not to the size of sexual seeds or that of other organs. With this, we aim to show that screening for epialleles is a promising strategy to uncover loci underlying relevant traits and could pave the way to identifying genes necessary for the engineering of apomixis.

Isolation and characterization of rice (Oryza sativa L.) E3-ubiquitin ligase OsHOS1 gene in the modulation of cold stress response.

Plants can cope with adverse environmental conditions through the activation of stress response signalling pathways, in which the proteasome seems to play an important role. However, the mechanisms underlying the proteasome-mediated stress response in rice are still not fully understood. To address this issue, we have identified a rice E3-ubiquitin ligase, OsHOS1, and characterized its role in the modulation of the cold stress response. Using a RNA interference (RNAi) transgenic approach we found that, under cold conditions, the RNAi::OsHOS1 plants showed a higher expression level of OsDREB1A. This was correlated with an increased amount of OsICE1, a master transcription factor of the cold stress signalling. However, the up-regulation of OsDREB1A was transient and the transgenic plants did not show increased cold tolerance. Nevertheless, we could confirm the interaction of OsHOS1 with OsICE1 by Yeast-Two hybrid and bi-molecular fluorescence complementation in Arabidopsis protoplasts. Moreover, we could also determine through an in vitro degradation assay that the higher amount of OsICE1 in the transgenic plants was correlated with a lower amount of OsHOS1. Hence, we could confirm the involvement of the proteasome in this response mechanism. Taken together our results confirm the importance of OsHOS1, and thus of the proteasome, in the modulation of the cold stress signalling in rice.

OsRMC, a negative regulator of salt stress response in rice, is regulated by two AP2/ERF transcription factors.

High salinity causes remarkable losses in rice productivity worldwide mainly because it inhibits growth and reduces grain yield. To cope with environmental changes, plants evolved several adaptive mechanisms, which involve the regulation of many stress-responsive genes. Among these, we have chosen OsRMC to study its transcriptional regulation in rice seedlings subjected to high salinity. Its transcription was highly induced by salt treatment and showed a stress-dose-dependent pattern. OsRMC encodes a receptor-like kinase described as a negative regulator of salt stress responses in rice. To investigate how OsRMC is regulated in response to high salinity, a salt-induced rice cDNA expression library was constructed and subsequently screened using the yeast one-hybrid system and the OsRMC promoter as bait. Thereby, two transcription factors (TFs), OsEREBP1 and OsEREBP2, belonging to the AP2/ERF family were identified. Both TFs were shown to bind to the same GCC-like DNA motif in OsRMC promoter and to negatively regulate its gene expression. The identified TFs were characterized regarding their gene expression under different abiotic stress conditions. This study revealed that OsEREBP1 transcript level is not significantly affected by salt, ABA or severe cold (5 °C) and is only slightly regulated by drought and moderate cold. On the other hand, the OsEREBP2 transcript level increased after cold, ABA, drought and high salinity treatments, indicating that OsEREBP2 may play a central role mediating the response to different abiotic stresses. Gene expression analysis in rice varieties with contrasting salt tolerance further suggests that OsEREBP2 is involved in salt stress response in rice.

Endoparasites of marsupials in fragments of the Atlantic rainforest, western Paraná State, Brazil.

Knowledge of taxonomy and biodiversity of parasites is fundamental to better understand ecosystem dynamics. The objective of this study was to describe the helminth fauna of two species of marsupials in five fragments of the Atlantic rainforest in the western region of Paraná State, Brazil. In a total of 4050 trap-nights, the animals were captured using Sherman, Tomahawk, and Pitfall traps, euthanized, necropsied, and their organs inspected for helminths. After identification of the parasites, descriptors of infection, such as prevalence, mean abundance, mean intensity, and range of intensity, were calculated. Collectively, six helminth species were observed in 18 animals. The following five species were observed in Marmosa paraguayana: Viannaia hamata (58.8%), Gracilioxyuris agilisis (52.9%), Travassostrongylus sextus (17.6%), Oncicola luehei (5.9%), and Pritchardia boliviensis (5.9%). Whereas the following two species were observed in Monodelphis dimidiata: Trichohelix tuberculata (100%) and Travassostrongylus sextus (100%). This study represents a new locality record for all helminths described herein, and a new host for four helminth species. This is the first report on the helminth fauna of Monodelphis dimidiata, expanding knowledge about marsupials in the Brazilian Atlantic Forest.

Seven zinc-finger transcription factors are novel regulators of the stress responsive gene OsDREB1B.

Plants have evolved several mechanisms in order to cope with adverse environmental conditions. The transcription factors (TFs) belonging to the DREB1/CBF subfamily have been described as major regulators of the plant responses to different abiotic stresses. This study focused on the rice gene OsDREB1B, initially described as highly and specifically induced by cold. However, here it is shown that OsDREB1B is not only induced by low temperatures, but also by drought and mechanical stress. In order to identify novel TFs that bind to its promoter, a yeast one-hybrid system was used to screen a cold-induced cDNA expression library. Thereby seven novel Zn-finger TFs were identified that bind to the promoter of OsDREB1B. Among them, there were four Zn-finger homeodomain (ZF-HD) and three C(2)H(2)-type Zn-finger TFs. Gene expression studies showed that these TFs are differentially regulated at transcriptional level by different abiotic stress conditions, which is illustrative of the crosstalk between stress signalling pathways. Protein-protein interaction studies revealed the formation of homo- and heterodimers among the ZF-HD TFs identified, but not for the C(2)H(2)-type. Using a transactivation assay in Arabidopsis protoplasts, all the TFs identified repressed the expression of the reporter gene, driven by the promoter of OsDREB1B. This assay also showed that the dimerization observed between the ZF-HD TFs may play a role on their transactivation activity. The results here presented suggest a prominent role of Zn-finger TFs in the regulation of OsDREB1B.

Rodent helminths in fragmented Atlantic Forest areas in the western region of the state of Paraná.

Rodents are small mammals that can be parasitized by various helminths. This study aimed to identify and describe the ecological indicators of infection in rodents captured in fragments of the Atlantic Forest in the western region of Paraná State, Brazil. Sixty-eight specimens of five rodent species were collected, necropsied, and inspected in search of helminths. The parasites were stored in 70% ethanol, morphologically identified, and counted for calculation of infection indicators. Fourteen species of helminths and one species of Crustacea were recorded: ten in Akodon montensis, four in Mus musculus, two in Thaptomys nigrita, two in Oligoryzomys nigripes, and one in Euryoryzomys russatus. The registered species of parasites were: Rodentolepis akodontis, Angiostrongylus sp., Protospirura numidica criceticola, Trichuris navonae, Syphacia alata, Syphacia criceti, Syphacia evaginata, Trichofreitasia lenti, Stilestrongylus aculeata, Stilestrongylus eta, Stilestrongylus gracielae, Stilestrongylus franciscanus, Stilestrongylus moreli, Stilestrongylus sp., and Pentastomida gen. sp. A positive correlation between the intensity of infection of T. navonae and T. lenti was observed with the body condition index of the host A. montensis. For all species, this study represents a new register of locality, and for eight of them a new host.

The MADS-box transcription factor PHERES1 controls imprinting in the endosperm by binding to domesticated transposons.

MADS-box transcription factors (TFs) are ubiquitous in eukaryotic organisms and play major roles during plant development. Nevertheless, their function in seed development remains largely unknown. Here, we show that the imprinted Arabidopsis thaliana MADS-box TF PHERES1 (PHE1) is a master regulator of paternally expressed imprinted genes, as well as of non-imprinted key regulators of endosperm development. PHE1 binding sites show distinct epigenetic modifications on maternal and paternal alleles, correlating with parental-specific transcriptional activity. Importantly, we show that the CArG-box-like DNA-binding motifs that are bound by PHE1 have been distributed by RC/Helitron transposable elements. Our data provide an example of the molecular domestication of these elements which, by distributing PHE1 binding sites throughout the genome, have facilitated the recruitment of crucial endosperm regulators into a single transcriptional network.

Endoparasites of marsupials in fragments of the Atlantic rainforest, western Paraná State, Brazil / Endoparasitas de marsupiais em fragmentos de Mata Atlântica no oeste do Estado do Paraná, Brasil

Abstract Knowledge of taxonomy and biodiversity of parasites is fundamental to better understand ecosystem dynamics. The objective of this study was to describe the helminth fauna of two species of marsupials in five fragments of the Atlantic rainforest in the western region of Paraná State, Brazil. In a total of 4050 trap-nights, the animals were captured using Sherman, Tomahawk, and Pitfall traps, euthanized, necropsied, and their organs inspected for helminths. After identification of the parasites, descriptors of infection, such as prevalence, mean abundance, mean intensity, and range of intensity, were calculated. Collectively, six helminth species were observed in 18 animals. The following five species were observed in Marmosa paraguayana: Viannaia hamata (58.8%), Gracilioxyuris agilisis (52.9%), Travassostrongylus sextus (17.6%), Oncicola luehei (5.9%), and Pritchardia boliviensis (5.9%). Whereas the following two species were observed in Monodelphis dimidiata: Trichohelix tuberculata (100%) and Travassostrongylus sextus (100%). This study represents a new locality record for all helminths described herein, and a new host for four helminth species. This is the first report on the helminth fauna of Monodelphis dimidiata, expanding knowledge about marsupials in the Brazilian Atlantic Forest.

Resumo Conhecimentos taxonômicos e da biodiversidade parasitária são fundamentais para uma melhor compreensão da dinâmica do ecossistema. O objetivo do estudo foi descrever a helmintofauna de marsupiais em cinco fragmentos de Mata Atlântica, na região Oeste do Estado do Paraná, Brasil. Em um total de 4.050 armadilhas/noite, os animais foram capturados com as do tipo Sherman, Tomahawk e Pitfall, eutanasiados, necropsiados e seus órgãos inspecionados em busca de helmintos. Após a identificação do parasita, foram calculados os indicadores de infecção (prevalência, abundância média, intensidade média e variação de intensidade). Seis espécies de helmintos foram observadas em dezoito animais; cinco em Marmosa paraguayana: Viannaia hamata (58,8%), Gracilioxyuris agilisis (52,9%), Travassostrongylus sextus (17,6%), Oncicola luehei (5,9%) e Pritchardia boliviensis (5,9%), e duas em Monodelphis dimidiata: Trichohelix tuberculata (100%) e Travassostrongylus sextus (100%). Este estudo representa um novo registro de localidade para todos os helmintos descritos e um novo hospedeiro para quatro deles. Trata-se do primeiro relato da helmintofauna de Monodelphis dimidiata, expandindo-se os conhecimentos sobre marsupiais na mata Atlântica brasileira.

Bridging the generation gap: communication between maternal sporophyte, female gametophyte and fertilization products.

In seed plants, as in placental animals, gamete formation and zygotic development take place within the parental tissues. To ensure timely onset and to coordinate the development of the new generation, communication between the parent plant with the filial tissues and its precursors is of utmost importance. During female gametogenesis the maternal tissues tightly regulate megagametophyte formation and the interplay between the sporophyte and the fertilization products, embryo and endosperm, has major implications in the formation of a viable seed. We review the current knowledge on these interactions and highlight the many questions that still remain unanswered, in particular the nature of the pathways involved in these signaling events.