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In green plants, photosystem I (PSI) and photosystem II (PSII) bind to their respective light-harvesting complexes (LHCI and LHCII) to form the PSI−LHCI supercomplex and the PSII−LHCII supercomplex, respectively. These supercomplexes further form megacomplexes, like PSI−PSII and PSII−PSII in Arabidopsis (Arabidopsis thaliana) and spinach to modulate their light-harvesting properties, but not in the green alga Chlamydomonas reinhardtii. Here, we fractionated and characterized the stable rice PSI−PSII megacomplex. The delayed fluorescence from PSI (lifetime ∼25 ns) indicated energy transfer capabilities between the two photosystems (energy spillover) in the rice PSI−PSII megacomplex. Fluorescence lifetime analysis revealed that the slow PSII to PSI energy transfer component was more dominant in the rice PSI−PSII supercomplexes than in Arabidopsis ones, suggesting that PSI and PSII in rice form a megacomplex not directly but through LHCII molecule(s), which was further confirmed by the negatively stained electron microscopy analysis. Our results suggest species diversity in the formation and stability of photosystem megacomplexes, and the stable PSI–PSII supercomplex in rice may reflect its structural adaptation.

As a ubiquitous picophytoplankton in the ocean and an early-branching green alga, Ostreococcus tauri is a model prasinophyte species for studying the functional evolution of the light-harvesting systems in photosynthesis. Here, we report the structure and function of the O. tauri photosystem I (PSI) supercomplex in low light conditions, where it expands its photon-absorbing capacity by assembling with the light-harvesting complexes I (LHCI) and a prasinophyte-specific light-harvesting complex (Lhcp). The architecture of the supercomplex exhibits hybrid features of the plant-type and the green algal-type PSI supercomplexes, consisting of a PSI core, an Lhca1-Lhca4-Lhca2-Lhca3 belt attached on one side and an Lhca5-Lhca6 heterodimer associated on the other side between PsaG and PsaH. Interestingly, nine Lhcp subunits, including one Lhcp1 monomer with a phosphorylated amino-terminal threonine and eight Lhcp2 monomers, oligomerize into three trimers and associate with PSI on the third side between Lhca6 and PsaK. The Lhcp1 phosphorylation and the light-harvesting capacity of PSI were subjected to reversible photoacclimation, suggesting that the formation of OtPSI-LHCI-Lhcp supercomplex is likely due to a phosphorylation-dependent mechanism induced by changes in light intensity. Notably, this supercomplex did not exhibit far-red peaks in the 77 K fluorescence spectra, which is possibly due to the weak coupling of the chlorophyll a603-a609 pair in OtLhca1-4.

As a ubiquitous picophytoplankton in the ocean and an early-branching green alga,Ostreococcus tauriis a model prasinophyte species for studying the functional evolution of the light-harvesting systems in photosynthesis. Here, we report the structure and function of theO. tauriphotosystem I (PSI) supercomplex in the low light, where it expands its photon-absorbing capacity by assembling with the light-harvesting complexes I (LHCI) and a prasinophyte-specific light-harvesting complex (Lhcp). Its architecture exhibits hybrid features of the plant-type and the green algal-type PSI supercomplexes, consisting of a PSI core, a Lhca1-Lhca4-Lhca2-Lhca3 belt attached on one side and a Lhca5-Lhca6 heterodimer associated on the other side between PsaG and PsaH. Interestingly, nine Lhcp subunits, including one Lhcp1 monomer with a phosphorylated amino-terminal threonine and eight Lhcp2 monomers, oligomerize into three trimers and associate with PSI on the third side between Lhca6 and PsaK. The Lhcp1 phosphorylation and the light-harvesting capacity of PSI were subjected to reversible photoacclimation, suggesting that the formation ofOtPSI–LHCI–Lhcp supercomplex is likely due to a state transition-like mechanism induced by light intensity change. Notably, this supercomplex did not exhibit far-red peaks in the 77 K fluorescence spectra, which is possibly due to weak coupling of the chlorophylla603-a609 pair inOtLhca1-4.

Summary In the seminiferous tubules of mouse testes, a population of glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRα1)-positive spermatogonia harbors the stem cell functionality and supports continual spermatogenesis, likely independent of asymmetric division or definitive niche control. Here, we show that activation of Wnt/β-catenin signaling promotes spermatogonial differentiation and reduces the GFRα1+ cell pool. We further discovered that SHISA6 is a cell-autonomous Wnt inhibitor that is expressed in a restricted subset of GFRα1+ cells and confers resistance to the Wnt/β-catenin signaling. Shisa6+ cells appear to show stem cell-related characteristics, conjectured from the morphology and long-term fates of T (Brachyury)+ cells that are found largely overlapped with Shisa6+ cells. This study proposes a generic mechanism of stem cell regulation in a facultative (or open) niche environment, with which different levels of a cell-autonomous inhibitor (SHISA6, in this case) generates heterogeneous resistance to widely distributed differentiation-promoting extracellular signaling, such as WNTs., Graphical Abstract, Highlights • Wnt/β-catenin signaling promotes the differentiation of GFRα1+ spermatogonia • SHISA6 is a cell-autonomous Wnt inhibitor expressed in subset GFRα1+ cells • SHISA6 confers resistance to differentiation induction by Wnt/β-catenin signaling • SHISA6+ spermatogonia show stem cell-related properties, In mouse testicular seminiferous tubules, Yoshida and colleagues questioned how sperm stem cells follow different fates (to differentiate or to self-renew) in response to homogeneously distributed extracellular signals. They showed that heterogeneous expression of Shisa6, a cell-autonomous Wnt inhibitor, confers resistance to homogeneously distributed differentiation-promoting Wnt signaling. This may be generic for stem cell regulation in a facultative niche.

Accumulating evidence suggests that exosomes are heterogeneous in molecular composition and physical properties. Here we examined whether epithelial cells secrete a heterogeneous population of exosomes, and if that is the case, whether epithelial cell polarity affects release of different populations of exosomes, especially that of those carrying Wnt. Sucrose-density ultracentrifugation and molecular marker analysis revealed that different populations of exosomes or exosome-like vesicles were released from MDCK cells depending on the cell polarity. Wnt3a associated with these vesicles were detectable in culture media collected from both apical and basolateral sides of the cells. Basolaterally secreted Wnt3a were co-fractionated with a typical exosomal protein TSG101 in fractions having typical exosome densities. In contrast, most of apically secreted Wnt3a, as well as Wnt11, were co-fractionated with CD63 and Hsp70, which are also common to the most exosomes, but recovered in higher density fractions. Wnt3a exhibiting similar floatation behavior to the apically secreted ones were also detectable in the culture media of Wnt3a-expressing L and HEK293 cells. The lipidation of Wnt3a was required for its basolateral secretion in exosomes but was dispensable for the apical one. Thus, epithelial cells release Wnt via distinct populations of vesicles differing in secretion polarity and lipidation dependency.

In many animal species, germ-line progenitors associate with gonadal somatic cells to form the embryonic gonads (EGs) that later develop into functional organ producing gametes. To explore the genetic regulation of the germ-line development, we initiated a comprehensive identification and functional analysis of the genes expressed within the EGs. First, we generated a cDNA library from gonads purified from Drosophila embryos by FACS. Using this library, we catalogued the genes expressed in the gonad by EST analysis. A total of 17,218 high-quality ESTs representing 3,051 genes were obtained, corresponding to 20% of the predicted genes in the genome. The EG transcriptome is unexpectedly distinct from that of adult gonads and includes an extremely high proportion of retrotransposon-derived transcripts. We verified 101 genes preferentially expressed in the EGs by whole-mount in situ hybridization. Within this subset, 39 and 58 genes were expressed predominantly in germ-line and somatic cells, respectively, whereas four genes were expressed in the both cell lineages. The gonad-enriched genes encompassed a variety of predicted functions. However, genes implicated in SUMOylation and protein translation, including germ-line-specific ribosomal proteins, are preferentially expressed in the germ line, whereas the expression of various retrotransposons and RNAi-related genes are more prominent in the gonadal soma. These transcriptome data are a resource for understanding the mechanism of various cellular events during germ-line development.

Primordial germ cells (PGC) are the earliest identifiable germ cells in the embryo. To understand the molecular basis of germline development, isolation of pure PGC is required. We report here the use of fluorescence-activated cell sorting (FACS) to isolate pure populations of Drosophila pole cells, which are the presumptive primordial germ cells in flies. In order to fluorescently mark pole cells, we used an EGFP-vasa transgenic line that expresses green fluorescent protein (GFP) specifically and continuously in the germ line throughout the life cycle. The purity of FACS-sorted pole cells from embryos was confirmed by microscopic inspection and quantitative polymerase chain reaction. Moreover, by optimizing the sample preparation and the sorting protocol, embryonic gonads could also be isolated. This technique opens the way for genome-wide transcriptome analysis of germline cells. In a pilot experiment, we generated a cDNA library from purified embryonic gonad and identified a novel germline-specific gene, RpL22-like.

日本, 韓国, 中国, 台湾, ブータンおよびネパールにおいて収集された在来アズキ203系統を供試材料としてDNAレベルの遺伝的変異を解析した. RAPD法により48個のプライマーを用いて解析した結果, 全部で413本の明瞭なバンドが検出され, そのうち97本のバンドで多型が認められた. これらのRAPDバンドを用いると, 供試系統のほとんどを識別することができた. そこで, 多型を示したバンドの有無について1-0データマトリクスを作成し, Simple matching coefficientを求めて非類似度を算出した. その結果, 各地域間の非類似度は, 地域内のそれよりも大きな値を示した. 特に, ブータン-ネパール地域の系統群と日本および韓国-中国-台湾地域の系統群との間において, それぞれ0.615および0.598と極めて大きな値が認められた。全系統間の非類似度をもとにUPGMA法によりクラスター分析を行ったところ, ほぼ日本, 韓国-中国-台湾地域およびブータン-ネパール地域に対応したグループに供試系統を分類することができた. これらのことから, 地域間での在来系統の交流があまりなかったことが推察された. また, 本研究では, 在来アズキ系統にはDNAレベルで大きな遺伝的変異が存在し, これらが今後の育種素材として有用であること, さらに, ブータン-ネパール地域の系統と東アジア地域の系統との雑種集団を用いることにより, 種内レベルでの分子地図作成が可能であることが示唆された.

Stem cells ensure tissue homeostasis through the production of differentiating and self-renewing progeny. In some tissues, this is achieved by the function of a definitive stem cell niche. However, the mechanisms that operate in mouse spermatogenesis are unknown because undifferentiated spermatogonia (Aundiff) are motile and intermingle with differentiating cells in an ‘open’ niche environment of seminiferous tubules. Aundiff include glial cell line-derived neurotrophic factor receptor α1 (GFRα1)+ and neurogenin 3 (NGN3)+ subpopulations, both of which retain the ability to self-renew. However, whereas GFRα1+ cells comprise the homeostatic stem cell pool, NGN3+ cells show a higher probability to differentiate into KIT+ spermatogonia by as yet unknown mechanisms. In the present study, by combining fate analysis of pulse-labeled cells and a model of vitamin A deficiency, we demonstrate that retinoic acid (RA), which may periodically increase in concentration in the tubules during the seminiferous epithelial cycle, induced only NGN3+ cells to differentiate. Comparison of gene expression revealed that retinoic acid receptor γ (Rarg) was predominantly expressed in NGN3+ cells, but not in GFRα1+ cells, whereas the expression levels of many other RA response-related genes were similar in the two populations. Ectopic expression of RARγ was sufficient to induce GFRα1+ cells to directly differentiate to KIT+ cells without transiting the NGN3+ state. Therefore, RARγ plays key roles in the differentiation competence of NGN3+ cells. We propose a novel mechanism of stem cell fate selection in an open niche environment whereby undifferentiated cells show heterogeneous competence to differentiate in response to ubiquitously distributed differentiation-inducing signals., Highlighted article: In the open environment of mouse seminiferous tubules, an undifferentiated stem cell pool is maintained by the differential expression of the retinoic acid receptor-γ in spermatogonia.

The seed storage proteins of 434 strains of azuki bean, Vigna angularis (Willd.) Ohwi and Ohashi, from Japan, South Korea, Bhutan, Nepal, China (mainland) and Taiwan, were analyzed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Fifteen protein types (types I to XV) were identified on the basis of the combination of the polymorphic bands. The geographical distribution of total protein types differed among regions from where the strains originated. In Bhutan and Nepal, most strains showed region-specific types I and II, whereas a few strains had type III. Genetic variability (H’) of Bhutan and Nepal strains were 0.502 and 0.636, respectively. Four types (III, IV, V and VII) were detected in germplasm from South Korea. Of these, types III and IV were major types. They showed a genetic variability value of 0.854. All strains from China (mainland) and Taiwan belonged to type III and type IV, respectively. In Japan, 311 strains from 38 prefectures were divided into seven geographical groups, i.e., Hokkaido, North, East, Central and West Honshu, Shikoku and Kyushu. In Hokkaido, only types III and IV were detected with equal frequency, whereas type IV was dominant in other six regions. Type V strains were distributed with a low frequency in East, Central and West Honshu, and Shikoku. A small number of strains with the other nine types (VI, VIII-XV) were uniquely observed in Central Honshu, West Honshu, Shikoku and/or Kyushu. Most of these unique types were detected with the strains in the districts along the coast of the Japan Sea. Consequently, the strains of northeastern and central Japan exhibited a lower genetic variability (0.416-0.694) than those of southwestern Japan (0.619-1.286). The highest genetic variability (1.286) was found in West Honshu where 11 kinds of protein types were observed.