Identification of hepta-histidine as a candidate drug for Huntington's disease by in silico-in vitro- in vivo-integrated screens of chemical libraries.

We identified drug seeds for treating Huntington's disease (HD) by combining in vitro single molecule fluorescence spectroscopy, in silico molecular docking simulations, and in vivo fly and mouse HD models to screen for inhibitors of abnormal interactions between mutant Htt and physiological Ku70, an essential DNA damage repair protein in neurons whose function is known to be impaired by mutant Htt. From 19,468 and 3,010,321 chemicals in actual and virtual libraries, fifty-six chemicals were selected from combined in vitro-in silico screens; six of these were further confirmed to have an in vivo effect on lifespan in a fly HD model, and two chemicals exerted an in vivo effect on the lifespan, body weight and motor function in a mouse HD model. Two oligopeptides, hepta-histidine (7H) and Angiotensin III, rescued the morphological abnormalities of primary neurons differentiated from iPS cells of human HD patients. For these selected drug seeds, we proposed a possible common structure. Unexpectedly, the selected chemicals enhanced rather than inhibited Htt aggregation, as indicated by dynamic light scattering analysis. Taken together, these integrated screens revealed a new pathway for the molecular targeted therapy of HD

Distinct hormonal and morphological control of dormancy and germination in Chenopodium album dimorphic seeds

Dormancy and heteromorphism are innate seed properties that control germination timing through adaptation to the prevailing environment. The degree of variation in dormancy depth within a seed population differs considerably depending on the genotype and maternal environment. Dormancy is therefore a key trait of annual weeds to time seedling emergence across seasons. Seed heteromorphism, the production of distinct seed morphs (in color, mass or other morphological characteristics) on the same individual plant, is considered to be a bet-hedging strategy in unpredictable environments. Heteromorphic species evolved independently in several plant families and the distinct seed morphs provide an additional degree of variation. Here we conducted a comparative morphological and molecular analysis of the dimorphic seeds (black and brown) of the Amaranthaceae weed Chenopodium album. Freshly harvested black and brown seeds differed in their dormancy and germination responses to ambient temperature. The black seed morph of seedlot #1 was dormant and 2/3rd of the seed population had non-deep physiological dormancy which was released by after-ripening (AR) or gibberellin (GA) treatment. The deeper dormancy of the remaining 1/3rd non-germinating seeds required in addition ethylene and nitrate for its release. The black seeds of seedlot #2 and the brown seed morphs of both seedlots were non-dormant with 2/3rd of the seeds germinating in the fresh mature state. The dimorphic seeds and seedlots differed in testa (outer seed coat) thickness in that thick testas of black seeds of seedlot #1 conferred coat-imposed dormancy. The dimorphic seeds and seedlots differed in their abscisic acid (ABA) and GA contents in the dry state and during imbibition in that GA biosynthesis was highest in brown seeds and ABA degradation was faster in seedlot #2. Chenopodium genes for GA and ABA metabolism were identified and their distinct transcript expression patterns were quantified in dry and imbibed C. album seeds. Phylogenetic analyses of the Amaranthaceae sequences revealed a high proportion of expanded gene families within the Chenopodium genus. The identified hormonal, molecular and morphological mechanisms and dormancy variation of the dimorphic seeds of C. album and other Amaranthaceae are compared and discussed as adaptations to variable and stressful environments

Redox feedback regulation of ANAC089 signaling alters seed germination and stress response

21 p.-4 fig.-2 tab. 1 graph. abst.The interplay between the phytohormone abscisic acid (ABA) and the gasotransmitter nitric oxide (NO) regulates seed germination and post-germinative seedling growth. We show that GAP1 (germination in ABA and cPTIO 1) encodes the transcription factor ANAC089 with a critical membrane-bound domain and extranuclear localization. ANAC089 mutants lacking the membrane-tethered domain display insensitivity to ABA,salt, and osmotic and cold stresses, revealing a repressor function. Whole-genome transcriptional profiling and DNA-binding specificity reveals that ANAC089 regulates ABA- and redox-related genes. ANAC089 truncated mutants exhibit higher NO and lower ROS and ABA endogenous levels, alongside an altered thiol and disulfide homeostasis. Consistently, translocation of ANAC089 to the nucleus is directed by changes in cellular redox status after treatments with NO scavengers and redox-related compounds. Our results reveal ANAC089 to be a master regulator modulating redox homeostasis and NO levels, able to repress ABA synthesis and signaling during Arabidopsis seed germination and abiotic stress.Wethank the Spanish networks BIO2015-68957-REDT and RED2018-102397-T for stimulating discussions, as well as Dr. José M. Carrasco and Dr. Pablo Vera (IBMCP-CSIC) for help with the protein-expression experiments of the PBM. This work was financed by grants EcoSeed Impacts of Environmental Conditions on Seed Quality ‘‘EcoSeed-311840’’ ERC.KBBE.2012.1.1-01;BIO2017-85758-R and CSD2007-00057 (TRANSPLANTA) from the Ministerio de Ciencia, Innovación y Universidades (MICIU) (Spain); SA313P18 and SA137P20 from Junta de Castilla y León; Escalera de Excelencia CLU-2018-04 co-funded by the P.O. FEDER of Castilla y León 2014–2020 Spain (to O.L.); and the PhD and University Teacher Training Fellowship, Spanish Ministry of Science and Education (to P.A.).Peer reviewe

Reproduction and population structure of the sea urchin Heliocidaris crassispina in its newly extended range: The Oga Peninsula in the Sea of Japan, northeastern Japan.

Ocean warming has facilitated the range expansion of commercially important sea urchin species to higher latitudes. Heliocidaris crassispina was recorded to extend northward to Toga Bay along the Oga Peninsula, Japan following an increase in seawater temperatures, and replacement of local sea urchin species Mesocentrotus nudus. In order to identify evidence of adaptation occurring in response to a range extension of H. crassispina to the newly extended environments, we randomly collected 106 H. crassispina in August 2014 in Toga Bay, determined the growth and age composition and examined gonad traits (size, color and development). To confirm the gonad development, 30 H. crassispina with > 30 mm diameter were collected in July, August and September 2017. We found slower growth in the extended range than the central range. More delayed gonad development of males than those of females and a large variety of developmental stages in the acini of testis indicated that the spawning of both sexes of the sea urchins were asynchronous. In terms of gonad color, L* (lightness) values increased with increasing GI, while b* (yellowness) values decreased with increasing age. The population consisted of seven year-classes from 2006 to 2012, suggesting persistent juvenile recruitment. Long-term water temperature data indicated that the range extension of H. crassispina was due to ocean warming, in particular during the summer spawning season

CHOTTO1, a Putative Double APETALA2 Repeat Transcription Factor, Is Involved in Abscisic Acid-Mediated Repression of Gibberellin Biosynthesis during Seed Germination in Arabidopsis1[W][OA]

The phytohormones abscisic acid (ABA) and gibberellins (GAs) are the primary signals that regulate seed dormancy and germination. In this study, we investigated the role of a double APETALA2 repeat transcription factor, CHOTTO1 (CHO1), in seed dormancy, germination, and phytohormone metabolism of Arabidopsis (Arabidopsis thaliana). Wild-type seeds were dormant when freshly harvested seeds were sown, and these seeds were released from dormancy after a particular period of dry storage (after-ripening). The cho1 mutant seeds germinated easily even in a shorter period of storage than wild-type seeds. The cho1 mutants showed reduced responsiveness to ABA, whereas transgenic plants constitutively expressing CHO1 (p35S∷CHO1) showed an opposite phenotype. Notably, after-ripening reduced the ABA responsiveness of the wild type, cho1 mutants, and p35S∷CHO1 lines. Hormone profiling demonstrated that after-ripening treatment decreased the levels of ABA and salicylic acid and increased GA4, jasmonic acid, and isopentenyl adenine when wild-type seeds were imbibed. Expression analysis showed that the transcript levels of genes for ABA and GA metabolism were altered in the wild type by after-ripening. Hormone profiling and expression analyses indicate that cho1 seeds, with a short period of storage, resembled fully after-ripened wild-type seeds. Genetic analysis showed that the cho1 mutation partially restored delayed seed germination and reduced GA biosynthesis activity in the ABA-overaccumulating cyp707a2-1 mutant background but did not restore seed germination in the GA-deficient ga1-3 mutant background. These results indicate that CHO1 acts downstream of ABA to repress GA biosynthesis during seed germination

Seed Dormancy in Arabidopsis Requires Self-Binding Ability of DOG1 Protein and the Presence of Multiple Isoforms Generated by Alternative Splicing

<div><p>The Arabidopsis protein DELAY OF GERMINATION 1 (DOG1) is a key regulator of seed dormancy, which is a life history trait that determines the timing of seedling emergence. The amount of DOG1 protein in freshly harvested seeds determines their dormancy level. DOG1 has been identified as a major dormancy QTL and variation in <i>DOG1</i> transcript levels between accessions contributes to natural variation for seed dormancy. The <i>DOG1</i> gene is alternatively spliced. Alternative splicing increases the transcriptome and proteome diversity in higher eukaryotes by producing transcripts that encode for proteins with altered or lost function. It can also generate tissue specific transcripts or affect mRNA stability. Here we suggest a different role for alternative splicing of the <i>DOG1</i> gene. <i>DOG1</i> produces five transcript variants encoding three protein isoforms. Transgenic <i>dog1</i> mutant seeds expressing single <i>DOG1</i> transcript variants from the endogenous <i>DOG1</i> promoter did not complement because they were non-dormant and lacked DOG1 protein. However, transgenic plants overexpressing single DOG1 variants from the 35S promoter could accumulate protein and showed complementation. Simultaneous expression of two or more <i>DOG1</i> transcript variants from the endogenous <i>DOG1</i> promoter also led to increased dormancy levels and accumulation of DOG1 protein. This suggests that single isoforms are functional, but require the presence of additional isoforms to prevent protein degradation. Subsequently, we found that the DOG1 protein can bind to itself and that this binding is required for DOG1 function but not for protein accumulation. Natural variation for DOG1 binding efficiency was observed among Arabidopsis accessions and contributes to variation in seed dormancy.</p></div

The accumulation of DOG1 requires the presence of multiple isoforms.

<p>Germination profiles of control lines (<b>A</b>), single (<b>B</b>), double (<b>C</b>) and triple (<b>D</b>) <i>pDOG1_Cvi</i>:<i>DOG1-α</i>, <i>DOG1-β</i>, and <i>DOG1-δ</i> transformants in <i>dog1-1</i> after different periods of dry storage. Error bars represent S.E.M. of at least three biological replicates. w, week. <i>DOG1</i> overall mRNA (<b>E</b>) and protein (<b>F</b>) levels in transgenic lines. (<b>E</b>) <i>DOG1</i> mRNA level was normalised to <i>ACT8</i> mRNA level. (<b>F</b>) The top panel shows DOG1 protein, and the bottom panel a nonspecific band around 60 kD that is used as loading control (LC) [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005737#pgen.1005737.ref018" target="_blank">18</a>]. An asterisk on the left of the top panel shows molecular mass marker around 36 kD. The <i>dog1-1</i> mutant produces only truncated protein and serves as a negative control. Individual line names were indicated at the bottom for (E) and (F). The line colours beneath the transgenic line names correspond to the line colours in (A-D).</p