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Protistology ■ 9
NUCLEUS-ASSOCIATED ACTIN IN DIFFERENT STAGES OF AMOEBA PROTEUS CELL CYCLE
Berdieva M.A., Bogolyubov D.S., Demin S.Y.,
Podlipaeva Y.I., Goodkov A.V.
Institute of Cytology of the Russian Academy of
Sciences, St. Petersburg, Russia
maria.berd4@yandex.ru
Despite of plenty of the reports and reviews there are still a lot of blank spots in the matter of the organization and functioning of Amoeba proteus nuclear apparatus. According to our 3D-reconstruction of its chromatin compartment, it has a complicated and highly dynamic structure (Demin et al., 2016). Actin was shown to be a key protein actor in providing different nuclear processes. Immunocytochemical study has shown an actin meshwork strongly co-localized with chromatin fibrils. On the other hand, the amoeba nucleus appears embraced by a basket-like structure formed by F-actin that in turn is connected with cytoplasmic actin filaments. 3D-observations of the nucleus during the cell cycle indicate a regular process of the chromatin extrusion/elimination ofthe part ofthe chromatin "excess" to the cytoplasm. Based on the 3D-reconstruction ofphalloidin-stained cells, we suppose the existence of specific chromatin-binding sites interacting with actin filaments meshwork. We also propose a hypothesis that the chromatin extrusion is provided by actin filaments which may pull out the chromatin fibrils from the nucleus. Funded by the Russian Foundation for Basic Research, projects 15-0403451, 15-04-01857, and the granting program "Molecular and Cell Biology" of the Presidium of RAS.
INFLUENCE OF SALINITY STRESS ON DNA SYNTHESIS AND CHROMOSOME FINE STRUCTURE OF DINOFLAGELLATES PRO-ROCENTRUM MINIMUM (PAVILLARD) SCHILLER
Berdieva M.A.1, Filatova N.A.1, Knyazev N.A.12, Skarlato S.O.1
1 - Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia
2 - Nanotechnology Research and Education Centre, Russian Academy of Sciences, St. Petersburg, Russia maria.berd4@yandex.ru
Prorocentrum minimum is a common planktonic potentially toxic bloom-forming dinoflagellate. Being widely distributed, this species demonstrates high adaptive capacity to different factors, particularly salinity. We studied cellular and biochemical aspects of physiological adaptation of this dino-
flagellate. The basic culture of P. minimum, isolated from the Black Sea, was grown in 17 psu f/2 medium (cultivation conditions described in Pozdnyakov et al., 2014). Experiments were carried out by inoculation of cells into 4, 8, 35 psu f/2 medium and 17 psu as a control for 30 min. Then cells were returned to the basic medium, incubated for 24 h and fixed for flow cytometry or transmission electron microscopy. We detected DNA concentration value (1.2 pg/cell) and fine structure of chromosomes in control series. After inoculation of cells into 8 psu f/2 medium we observed the highest DNA concentration (1.5 pg/cell) and the lowest cell mortality rates. Chromosomes became more condensed, local unwound sites increased in number and appeared more distinct. Transfer into the medium with native for these organisms salinity 35 psu (the World ocean level) did not caused any reliable differences in DNA concentration (1,3 pg/ cell). At the ultrastructural level, we observed total splitting of chromosomes in most cases. Inoculation of cells into 4 psu f/2 did not caused any significant changes in DNA concentration value, but led to high cell mortality rates. Our results are in agreement with the protistan species maximum concept for the horohalinicum. Funded by the Russian Science Foundation, project 16-14-10116.
FINDING AND ANALYSIS OF AMOEBO-ZOA-SPECIFIC GENES TO STUDY ENVIRONMENTAL DIVERSITY OF AMOEBAE Bondarenko N.I., Smirnov A.V. Department of Invertebrate Zoology, Faculty of Biology, Saint-Petersburg State University, Saint-Petersburg, Russia natalya.lannik@gmail.com
Amoebozoa is a one of the supergroups of eukaryo-tes, which includes naked and testate lobose amoebae, pelobionts, mycetozoa, and several groups of flagellated organisms. In environmental DNA surveys done on traditional DNA barcodes (SSU-rDNA, Cox I gene) amoebozoan genes normally constitutes a minor part of the total gene diversity and represent only the most abundant lineages. To resolve this problem, we attempted to find Amoebozoa-specific genes and gene families with low level of paralogy appropriate for the application as a DNA barcodes for this group of protists. We analyzed the Amoebozoa RNA-Seq data which were available from MMETSP (http://www.moore.org/). First, we analyzed assembled transcriptomes from MMETSP and found transcripts with unusually big size (more than 50 000 bp) in several assemblies. This led us to the decision to repeat data assembly de novo.
10 • "PROTIST—2016
During this work we developed pipeline, based on publically available bioinformatics tools and our own scripts written in Python for transcriptome assembly and annotation. We have found 300 groups ofgenes, which not found outside Amoebozoa or were highly derived within this group of protists. Among them we selected 15 groups of genes with low level of paralogy and performed phylogenetic analysis and primers construction. These genes are promising DNA barcodes for studies of environmental diversity of Amoebozoa. Supported with MK-4853.2015.4 President grant, RFBR 16-34-60111 and SPSU grant 1.38.251.2014.
BURIED BUT NOT DEAD: INSIGHTS INTO THE DIVERSITY, PHYSIOLOGY, FUNCTIONS AND ECOLOGICAL ROLES OF DEEP SUBSEAFLOOR FUNGI USING AN INTEGRATED APPROACH
Burgaud G.1, Rédou V.1, Pachiadaki M.G23, Navarri M.1, Fleury Y.1, Barbier G.1, Edgcomb V.P.2
1 - Université de Brest, EA 3882, Laboratoire Universitaire de Biodiversité et Ecologie Microbienne, ESIAB, Technopôle de Brest Iroise, 29280Plouzané, France
2 - Woods Hole Oceanographic Institution, Department ofGeology and Geophysics, Woods Hole, Massachusetts, United States of America
3 - Bigelow Laboratories, East Boothbay, Maine, USA gaetan.burgaud@univ-brest.fr
Bacteria and Archaea are the most commonly studied microorganisms in the marine environment, and habitats such as deep subseafloor ecosystems are no exception. However, recent studies strongly support the idea that deep subseafloor microbial communities include Fungi, which seem to dominate those micro-eukaryotic communities. Using sediment samples from the IODP Expedition 317 as a model, our aims were (i) to better understand the diversity, physiology and functions of deep subseafloor fungi and (ii) to provide clues about how they interact with other microbial populations in those communities. Using a record depth sediment core, fungal molecu-lar signatures and fungal cultures were obtained from samples as deep as 1740mbsf (Rédou et al. 2014) and 1884mbsf (Rédou et al. 2015), respectively. In spite of the fact that those complementary approaches revealed low diversity of higher fungal lineages, DNA and rRNA signatures as well as almost 200 cultured isolates provide direct evidence that fungi persist in this challenging habitat. Consistent with this idea, physiological analyses indicate some deep subseafloor fungal isolates appear well-adapted to in situ conditions.
Metatranscriptome analysis provided an examination of the functional repertoire of deep subseafloor fungi. Gene expression was assigned to metabolic and biosynthetic processes, responses to stress, cell and membrane functions, conidiogenesis and biosynthesis of secondary metabolites (Pachiadaki et al., in revision). These results all provide further support for the notion offungal presence and activity in the deep subseafloor biosphere, with the ability to interact with other microbial populations by synthesizing antimicrobial compounds (Navarri et al. 2016).
THE GREEN ALGA AND THE SALAMANDER:
A SUFFOCATING LOVE STORY
Burns J.A.1, Zhang H.2, Hill E.2, Kerney R.2, Kim E.1
1 - American Museum of Natural History
2 - Gettysburg College jburns@amnh.org
The recently discovered endosymbiosis between the green alga Oophila amblystomatis and the salamander Ambystoma maculatum is a unique relationship among the chloroplastida and vertebrates. Using a dual RNA-seq approach, we assembled novel transcriptomes ofthese two organisms and identified differentially expressed transcripts between ecto-and endo-symbiotic algae as well as between salamander cells with and without endosymbiotic algae. The results offer a glimpse at the changes in both organisms that take place during this novel endosymbiosis. We found that the intracellular algae downregulate nutrient transporters related to phosphate and nitrogen acquisition from the environment. They also exhibit hallmarks of cellular stress, especially related to osmotic stress, sulfur starvation, and hypoxia. Further, the results suggest that the alga undergoes a large scale metabolic shift from oxidative metabolism to fermentation with the potential evolution ofhydrogen gas. The salamander cells exhibit milder differences, including changes in gene expression indicating the initiation of an innate immune response to the alga, and alterations in nutrient sensing related to insulin sensitivity. The salamander cells do not exhibit large scale stress or apoptotic responses suggesting that intracellular algae are not a big drain on the salamander cell's resources.
GENOME AND TRANSCRIPTOME OF HEMI-STASIA PHAEOCYSTICOLA, A FLAGELLATE RELATED TO A NOVEL HYPER-DIVERSE CLADE OF MARINE PROTISTS Butenko A.1, Yabuki A.2, Flegontova O.34, Horak A.3, Flegontov P.135, Lukes J.346 1 - Life Science Research Centre, Faculty ofScience,
