Morphology and chemical composition of Taiwan oil millet (Eccoilopus formosanus) epicuticular wax.

MAIN CONCLUSION: Taiwan oil millet has two types of epicuticular wax: platelet wax composed primarily of octacosanol and filament wax constituted essentially by the singular compound of octacosanoic acid. Taiwan oil millet (TOM-Eccoilopus formosanus) is an orphan crop cultivated by the Taiwan indigenous people. It has conspicuous white powder covering its leaf sheath indicating abundant epicuticular waxes, that may contribute to its resilience. Here, we characterized the epicuticular wax secretion in TOM leaf blade and leaf sheath using various microscopy techniques, as well as gas chromatography to determine its composition. Two kinds of waxes, platelet and filaments, were secreted in both the leaf blades and sheaths. The platelet wax is secreted ubiquitously by epidermal cells, whereas the filament wax is secreted by a specific cell called epidermal cork cells. The newly developed filament waxes were markedly re-synthesized by the epidermal cork cells through papillae protrusions on the external periclinal cell wall. Ultrastructural images of cork cell revealed the presence of cortical endoplasmic reticulum (ER) tubules along the periphery of plasma membrane (PM) and ER-PM contact sites (EPCS). The predominant wax component was a C28 primary alcohol in leaf blade, and a C28 free fatty acid in the leaf sheath, pseudopetiole and midrib. The wax morphology present in distinct plant organs corresponds to the specific chemical composition: platelet wax composed of alcohols exists mainly in the leaf blade, whereas filament wax constituted mainly by the singular compound C28 free fatty acids is present abundantly in leaf sheath. Our study clarifies the filament wax composition in relation to a previous study in sorghum. Both platelet and filament waxes comprise a protection barrier for TOM.

Maize Golden2-like transcription factors boost rice chloroplast development, photosynthesis, and grain yield.

Chloroplasts are the sites for photosynthesis, and two Golden2-like factors act as transcriptional activators of chloroplast development in rice (Oryza sativa L.) and maize (Zea mays L.). Rice OsGLK1 and OsGLK2 are orthologous to maize ZmGLK1 (ZmG1) and ZmGLK2 (ZmG2), respectively. However, while rice OsGLK1 and OsGLK2 act redundantly to regulate chloroplast development in mesophyll cells, maize ZmG1 and ZmG2 are functionally specialized and expressed in different cell-specific manners. To boost rice chloroplast development and photosynthesis, we generated transgenic rice plants overexpressing ZmG1 and ZmG2, individually or simultaneously, with constitutive promoters (pZmUbi::ZmG1 and p35S::ZmG2) or maize promoters (pZmG1::ZmG1, pZmG2::ZmG2, and pZmG1::ZmG1/pZmG2::ZmG2). Both ZmG1 and ZmG2 genes were highly expressed in transgenic rice leaves. Moreover, ZmG1 and ZmG2 showed coordinated expression in pZmG1::ZmG1/pZmG2::ZmG2 plants. All Golden2-like (GLK) transgenic plants had higher chlorophyll and protein contents, Rubisco activities and photosynthetic rates per unit leaf area in flag leaves. However, the highest grain yields occurred when maize promoters were used; pZmG1::ZmG1, pZmG2::ZmG2, and pZmG1::ZmG1/pZmG2::ZmG2 transgenic plants showed increases in grain yield by 51%, 47%, and 70%, respectively. In contrast, the pZmUbi::ZmG1 plant produced smaller seeds without yield increases. Transcriptome analysis indicated that maize GLKs act as master regulators promoting the expression of both photosynthesis-related and stress-responsive regulatory genes in both rice shoot and root. Thus, by promoting these important functions under the control of their own promoters, maize GLK1 and GLK2 genes together dramatically improved rice photosynthetic performance and productivity. A similar approach can potentially improve the productivity of many other crops.

3D Imaging of Tapetal Mitochondria Suggests the Importance of Mitochondrial Fission in Pollen Growth.

Mitochondrial fission occurs frequently in plant cells, but its biological significance is poorly understood because mutants specifically impaired in mitochondrial fission do not show obvious defects in vegetative growth. Here, we revealed that the production of viable pollen was reduced in mutants lacking one of the three main proteins involved in mitochondrial fission in Arabidopsis (Arabidopsis thaliana), DYNAMIN-RELATED PROTEIN3A (DRP3A)/Arabidopsis DYNAMIN-LIKE PROTEIN2A, DRP3B, and ELONGATED MITOCHONDRIA1 (ELM1). In drp3b and elm1, young microspores contained an abnormal number of nuclei, and mature pollen had aberrant accumulation of lipids in their coat and an irregular pollen outer wall. Because the formation of the pollen wall and coat is mainly associated with tapetal function, we used 3D imaging to quantify geometric and textural features of cells and mitochondria in the tapetum at different stages, using isolated single tapetal cells in which the in vivo morphology and volume of cells and mitochondria were preserved. Tapetal cells and their mitochondria changed in the volume and morphology at different developmental stages. Defective mitochondrial fission in the elm1 and drp3b mutants caused changes in mitochondrial status, including mitochondrial elongation, abnormal mitochondrial ultrastructure, a decrease in cross-sectional area, and a slight alteration of mitochondrial distribution, as well as a large reduction in mitochondrial density. Our studies suggest that mitochondrial fission is required for proper mitochondrial status in the tapetum and possibly in pollen as well and therefore plays an important role for the production of viable pollen.

VPS36-Dependent Multivesicular Bodies Are Critical for Plasmamembrane Protein Turnover and Vacuolar Biogenesis.

Most eukaryotic cells target ubiquitinated plasma membrane (PM) proteins for vacuolar degradation in response to environmental and developmental cues. This process involves endosomal sorting complexes required for transport (ESCRT). However, little is known about the cellular mechanisms of ESCRTs in plants. Here, we studied the function of one ESCRT-II component, VPS36, which shows ubiquitin-binding activity and may form a putative ESCRT-II with VPS22 and VPS25 in Arabidopsis (Arabidopsis thaliana). Recessive mutation of the ubiquitously expressed VPS36 causes multiple defects, including delayed embryogenesis, defective root elongation, and limited expansion of cotyledons, and these effects can be complemented by its genomic DNA. Abnormal intracellular compartments containing several membrane transporters, including members of the PIN-FORMEDs, AUXIN RESISTANT 1, and PIP1 families, were found in vps36-1 plants. Employing a genetic approach to cross vps36-1/+ with transgenic plants harboring various fluorescent protein-tagged organelle markers, as well as fluorescent probe and ultrastructural approaches, revealed PM proteins in microsomal fractions from vps36-1 seedlings and demonstrated that VPS36 is critical for forming multivesicular bodies and vacuolar biogenesis for protein degradation. Our study shows that functional VPS36 is essential for a proper endosomal sorting pathway and for vacuolar biogenesis in Arabidopsis.

Tandem oleosin genes in a cluster acquired in Brassicaceae created tapetosomes and conferred additive benefit of pollen vigor.

During evolution, genomes expanded via whole-genome, segmental, tandem, and individual-gene duplications, and the emerged redundant paralogs would be eliminated or retained owing to selective neutrality or adaptive benefit and further functional divergence. Here we show that tandem paralogs can contribute adaptive quantitative benefit and thus have been retained in a lineage-specific manner. In Brassicaceae, a tandem oleosin gene cluster of five to nine paralogs encodes ample tapetum-specific oleosins located in abundant organelles called tapetosomes in flower anthers. Tapetosomes coordinate the storage of lipids and flavonoids and their transport to the adjacent maturing pollen as the coat to serve various functions. Transfer-DNA and siRNA mutants of Arabidopsis thaliana with knockout and knockdown of different tandem oleosin paralogs had quantitative and correlated loss of organized structures of the tapetosomes, pollen-coat materials, and pollen tolerance to dehydration. Complementation with the knockout paralog restored the losses. Cleomaceae is the family closest to Brassicaceae. Cleome species did not contain the tandem oleosin gene cluster, tapetum oleosin transcripts, tapetosomes, or pollen tolerant to dehydration. Cleome hassleriana transformed with an Arabidopsis oleosin gene for tapetum expression possessed primitive tapetosomes and pollen tolerant to dehydration. We propose that during early evolution of Brassicaceae, a duplicate oleosin gene mutated from expression in seed to the tapetum. The tapetum oleosin generated primitive tapetosomes that organized stored lipids and flavonoids for their effective transfer to the pollen surface for greater pollen vitality. The resulting adaptive benefit led to retention of tandem-duplicated oleosin genes for production of more oleosin and modern tapetosomes.

Transplastomic Nicotiana benthamiana plants expressing multiple defence genes encoding protease inhibitors and chitinase display broad-spectrum resistance against insects, pathogens and abiotic stresses.

Plastid engineering provides several advantages for the next generation of transgenic technology, including the convenient use of transgene stacking and the generation of high expression levels of foreign proteins. With the goal of generating transplastomic plants with multiresistance against both phytopathogens and insects, a construct containing a monocistronic patterned gene stack was transformed into Nicotiana benthamiana plastids harbouring sweet potato sporamin, taro cystatin and chitinase from Paecilomyces javanicus. Transplastomic lines were screened and characterized by Southern/Northern/Western blot analysis for the confirmation of transgene integration and respective expression level. Immunogold localization analyses confirmed the high level of accumulation proteins that were specifically expressed in leaf and root plastids. Subsequent functional bioassays confirmed that the gene stacks conferred a high level of resistance against both insects and phytopathogens. Specifically, larva of Spodoptera litura and Spodoptera exigua either died or exhibited growth retardation after ingesting transplastomic plant leaves. In addition, the inhibitory effects on both leaf spot diseases caused by Alternaria alternata and soft rot disease caused by Pectobacterium carotovorum subsp. carotovorum were markedly observed. Moreover, tolerance to abiotic stresses such as salt/osmotic stress was highly enhanced. The results confirmed that the simultaneous expression of sporamin, cystatin and chitinase conferred a broad spectrum of resistance. Conversely, the expression of single transgenes was not capable of conferring such resistance. To the best of our knowledge, this is the first study to demonstrate an efficacious stacked combination of plastid-expressed defence genes which resulted in an engineered tolerance to various abiotic and biotic stresses.

Role of the putative osmosensor Arabidopsis histidine kinase1 in dehydration avoidance and low-water-potential response.

The molecular basis of plant osmosensing remains unknown. Arabidopsis (Arabidopsis thaliana) Histidine Kinase1 (AHK1) can complement the osmosensitivity of yeast (Saccharomyces cerevisiae) osmosensor mutants lacking Synthetic Lethal of N-end rule1 and SH3-containing Osmosensor and has been proposed to act as a plant osmosensor. We found that ahk1 mutants in either the Arabidopsis Nossen-0 or Columbia-0 background had increased stomatal density and stomatal index consistent with greater transpirational water loss. However, the growth of ahk1 mutants was not more sensitive to controlled moderate low water potential (ψ(w)) or to salt stress. Also, ahk1 mutants had increased, rather than reduced, solute accumulation across a range of low ψ(w) severities. ahk1 mutants had reduced low ψ(w) induction of Δ(1)-Pyrroline-5-Carboxylate Synthetase1 (P5CS1) and 9-cis-Epoxycarotenoid Dioxygenase3, which encode rate-limiting enzymes in proline and abscisic acid (ABA) synthesis, respectively. However, neither Pro nor ABA accumulation was reduced in ahk1 mutants at low ψ(w). P5CS1 protein level was not reduced in ahk1 mutants. This indicated that proline accumulation was regulated in part by posttranscriptional control of P5CS1 that was not affected by AHK1. Expression of AHK1 itself was reduced by low ψ(w), in contrast to previous reports. These results define a role of AHK1 in controlling stomatal density and the transcription of stress-responsive genes. These phenotypes may be mediated in part by reduced ABA sensitivity. More rapid transpiration and water depletion can also explain the previously reported sensitivity of ahk1 to uncontrolled soil drying. The unimpaired growth, ABA, proline, and solute accumulation of ahk1 mutants at low ψ(w) suggest that AHK1 may not be the main plant osmosensor required for low ψ(w) tolerance.

A novel lily anther-specific gene encodes adhesin-like proteins associated with exine formation during anther development.

The anther-specific gene LLA1271 isolated from lily (Lilium longiflorum Thunb.) anthers is novel and exists in two forms. The protein encoded by LLA1271 may represent an adhesin-like protein first found in higher plants. The protein contains a typical N-terminal signal peptide followed by a highly conserved repeat domain. The LLA1271 gene is temporally expressed at the phase of microspore development. RNA blot and RNA in situ hybridization analyses demonstrated that the gene was expressed both in the tapetum and in the microspore. The gene is endo- and exogenously induced by gibberellin. Studies with the gibberellin biosynthesis inhibitor uniconazole and an inhibitor of ethylene activity, 2,5-norbornadien (NBD), revealed that LLA1271 is negatively regulated by ethylene, and a cross-talk of regulation between gibberellin and ethylene occurs in young anthers. The treatment with NBD caused the tapetum to become densely cytoplasmic and highly polarized, whereas uniconazole arrested tapetal development in a state close to that of a tapetum without treatment. The LLA1271 protein is heat stable and heterogeneous. An immunoblot of separated protein fractions of the anther revealed that the LLA1271 protein was detected in protein fraction of the microspore released from the cell wall by treatment with either 0.5% or 2% Triton X-100. Ectopic expression of LLA1271 resulted in impaired stamen and low pollen germination. Scanning electron microscopy of TAP::LLA1271 pollen showed distorted exine formation and patterning. The LLA1271 protein once synthesized in both the tapetum and microspore is secreted and deposited on the surface of microspores, moderately affecting exine formation and patterning.

Arabidopsis thaliana CENTRORADIALIS homologue (ATC) acts systemically to inhibit floral initiation in Arabidopsis.

Floral initiation is orchestrated by systemic floral activators and inhibitors. This remote-control system may integrate environmental cues to modulate floral initiation. Recently, FLOWERING LOCUS T (FT) was found to be a florigen. However, the identity of systemic floral inhibitor or anti-florigen remains to be elucidated. Here we show that Arabidopsis thaliana CENTRORADIALIS homologue (ATC), an Arabidopsis FT homologue, may act in a non-cell autonomous manner to inhibit floral initiation. Analysis of the ATC null mutant revealed that ATC is a short-day-induced floral inhibitor. Cell type-specific expression showed that companion cells and apex that express ATC are sufficient to inhibit floral initiation. Histochemical analysis showed that the promoter activity of ATC was mainly found in vasculature but under the detection limit in apex, a finding that suggests that ATC may move from the vasculature to the apex to influence flowering. Consistent with this notion, Arabidopsis seedling grafting experiments demonstrated that ATC moved over a long distance and that floral inhibition by ATC is graft transmissible. ATC probably antagonizes FT activity, because both ATC and FT interact with FD and affect the same downstream meristem identity genes APETALA1, in an opposite manner. Thus, photoperiodic variations may trigger functionally opposite FT homologues to systemically influence floral initiation.

Association of a D-alanyl-D-alanine carboxypeptidase gene with the formation of aberrantly shaped cells during the induction of viable but nonculturable Vibrio parahaemolyticus.

Vibrio parahaemolyticus is a halophilic Gram-negative bacterium that causes human gastroenteritis. When the viable but nonculturable (VBNC) state of this bacterium was induced by incubation at 4°C in Morita minimal salt solution containing 0.5% NaCl, the rod-shaped cells became coccoid, and various aberrantly shaped intermediates were formed in the initial stage. This study examined the factors that influence the formation of these aberrantly shaped cells. The proportion of aberrantly shaped cells was not affected in a medium containing D-cycloserine (50 µg/ml) but was lower in a medium containing cephalosporin C (10 µg/ml) than in the control medium without antibiotics. The proportion of aberrantly shaped cells was higher in a culture medium that contained 0.5% NaCl than in culture media containing 1.0 or 1.5% NaCl. The expression of 15 of 17 selected genes associated with cell wall synthesis was enhanced, and the expression of VP2468 (dacB), which encodes D-alanyl-D-alanine carboxypeptidase, was enhanced the most. The proportion of aberrantly shaped cells was significantly lower in the dacB mutant strain than in the parent strain, but the proportion was restored in the presence of the complementary dacB gene. This study suggests that disturbance of the dynamics of cell wall synthesis by enhanced expression of the VP2468 gene is associated with the formation of aberrantly shaped cells in the initial stage of induction of VBNC V. parahaemolyticus cells under specific conditions.

Enhancing polyhydroxyalkanoate production in Cupriavidus sp. L7L through wcaJ gene deletion.

Cupriavidus sp. L7L synthesizes a high content of ductile polyhydroxyalkanoate. However, during fermentation, the medium's viscosity gradually increases, eventually reaching a level similar to 93 % glycerol, leading to fermentation termination and difficulties in cell harvest. A non-mucoid variant was isolated from a mini-Tn5 mutant library with the transposon inserted at the promoter sequence upstream of the wcaJ gene. Deletion of wcaJ eliminated the mucoid-colony appearance. The complementation experiment confirmed the association between wcaJ gene expression and mucoid-colony formation. Additionally, the wild-type strain exhibited a faster specific growth rate than the deletion strain using levulinate (Lev) as a carbon source. In fed-batch fermentation, Cupriavidus sp. L7L∆wcaJ showed similar PHA content and monomer composition to the wild-type strain. However, the extended fermentation time resulted in a 42 % increase in PHA concentration. After fed-batch fermentation, the deletion strain's medium had only 8.75 % of the wild-type strain's extracellular polymeric substance content. Moreover, the deletion strain's medium had a much lower viscosity (1.04 mPa·s) than the wild-type strain (194.7 mPa·s), making bacterial cell collection easier through centrifugation. In summary, Cupriavidus sp. L7L∆wcaJ effectively addressed difficulties in cell harvest, increased PHA production, and Lev-to-PHA conversion efficiency, making these characteristics advantageous for industrial-scale PHA production.

The use of immobilised bacteria cross-linked within magnetic alginate beads enhances the treatment of benzophenone-type UV filter chemicals by the SBR system.

Benzophenone-n compounds (BPs) are applied in a broad spectrum of commercial products, one of which is sunscreen. These chemicals are frequently detected in a variety of environmental matrices worldwide, especially water bodies. BPs are defined as emerging contaminants as well as endocrine-disrupting contaminants; thus, it has become necessary to develop aggressive and green treatments to remove BPs. In this study, we used immobilised BP-biodegrading bacteria linked to reusable magnetic alginate beads (MABs). The MABs were added to a sequencing batch reactor (SBR) system to enhance the removal of 2,4-dihydroxybenzophenone (BP-1) and oxybenzone (BP-3) from sewage. The BP-1 and BP-3 biodegrading bacteria in the MABs consisted of strains from up to three genera to allow for efficient biodegradation. The strains used were Pseudomonas spp., Gordonia sp., and Rhodococcus sp. The optimal composition of the MABs consisted of 3% (w/v) alginate and 10% (w/v) magnetite. The MABs resulted in 60.8%-81.7% recovery by weight after 28 days, and there was a continuous release of bacteria. Moreover, the biological treatment of the BPs sewage improved after adding 100 g of BP1-MABs (1:27) and also 100 g BP3-MABs (1:27) into the SBR system at a hydraulic retention time (HRT) of 8 h. Compared with the SBR system without MABs, the removal rates of BP-1 and BP-3 increased from 64.2% to 71.5% and from 78.1% to 84.1%, respectively. Furthermore, the COD removal increased from 36.1% to 42.1%, and total nitrogen increased from 30.5% to 33.2%. Total phosphorus remained constant at 29%. The bacterial community analysis showed that the Pseudomonas population was <2% before the MAB addition, but increased to 56.1% by day 14. In contrast, the Gordonia sp. And Rhodococcus sp. Populations (<2%) remained unchanged throughout the 14-day treatment period.

Similar but different: Characterization of dddD gene-mediated DMSP metabolism among coral-associated Endozoicomonas.

Endozoicomonas are often predominant bacteria and prominently important in coral health. Their role in dimethylsulfoniopropionate (DMSP) degradation has been a subject of discussion for over a decade. A previous study found that Endozoicomonas degraded DMSP through the dddD pathway. This process releases dimethyl sulfide, which is vital for corals coping with thermal stress. However, little is known about the related gene regulation and metabolic abilities of DMSP metabolism in Endozoicomonadaceae. In this study, we isolated a novel Endozoicomonas DMSP degrader and observed a distinct DMSP metabolic trend in two phylogenetically close dddD-harboring Endozoicomonas species, confirmed genetically by comparative transcriptomic profiling and visualization of the change of DMSP stable isotopes in bacterial cells using nanoscale secondary ion spectrometry. Furthermore, we found that DMSP cleavage enzymes are ubiquitous in coral Endozoicomonas with a preference for having DddD lyase. We speculate that harboring DMSP degrading genes enables Endozoicomonas to successfully colonize various coral species across the globe.

Degradation of Decabromodiphenyl Ether in an Aerobic Clay Slurry Microcosm Using a Novel Immobilization Technique.

A novel chitosan immobilization technique that entraps photocatalyst and microbes was developed and applied to decompose decabromodiphenyl ether (BDE-209) in a clay slurry microcosm. The optimized conditions for immobilization were obtained by mixing 1.2% (w/v) chitosan dissolved in 1% (v/v) acetic acid with nano-TiO2 particles and the BDE-209-degrading bacterial mixed culture. This aqueous mixture was injected into 1% (w/v) water solution containing sodium tripolyphosphate to form spherical immobilized beads. The surface of the immobilized beads was reinforced by 0.25% (v/v) glutaraldehyde cross-linking. These beads had enough mechanical strength during BDE-209 degradation to maintain their shape in the system at a stirring rate of 200-rpm, while undergoing continuous 365 nm UVA irradiation. This novel TiO2-Yi-Li immobilized chitosan beads system allowed a successful simultaneous integration of photolysis, photocatalysis and biodegradation to remove BDE-209. The remaining percentage of BDE-209 was 41% after 70 days of degradation using this system. The dominant bacteria in the BDE-209-degrading bacterial mixed culture during remediation were Chitinophaga spp., Methyloversatilis spp., Terrimonas spp. and Pseudomonas spp. These bacteria tolerated the long-term UVA irradiation and high-level free radicals present, while utilizing BDE-209 as their primary carbon resource. This new method has great potential for the treatment of a range of pollutants.

Synthesis of cross-linked magnetic chitosan beads immobilised with bacteria for aerobic biodegrading benzophenone-type UV filter.

Bacterial immobilisation is a technique by which bacteria are embedded into or adsorbed onto a carrier material thereby increasing bacterial tolerance to harsh environments. This technique can be used to enhance bacterial activity and to degrade pollutants. Immobilised bacterial beads that contain nanomagnetic particles allow bead recycling and reuse. In this study, our objective was to produce cross-linked nanomagnetic chitosan beads (MCBs) for the biodegradation of benzophenone-type UV filter chemicals such as 2,4-dihydroxybenzophenone (BP-1) and oxybenzone (BP-3). We found that the optimal concentration for creating these MCBs to be 1.2% by weight chitosan and 10% by weight nano-magnetite. We selected and isolated six benzophenone-n (BPs)-biodegrading bacteria identified to be various Pseudomonas spp., a Gordonia sp., and Rhodococcus zopfii; these were used to create MCBs that were able to effectively biodegrade BP-1 or BP-3 as a sole carbon source. Both BPs were effectively biodegraded and mineralised over 8 days in the presence of the selected MCB-immobilised bacterial strains. The highest pseudo-first-order constant rates for BP biodegradation were 8.7 × 10-3 h-1 for BP-1 (strain BP1-D) and 1.02 × 10-3 h-1 for BP-3 (strain BP3-1). The mechanical strength of the MCBs was measured to be above 90% based on recovered weight. The MCBs released their bacteria at rates in the range of 104-105 CFU/day. We also determined the pathway through which the BPs were being aerobically biodegraded based on the GC/MS profiles of the intermediates. Our findings provide a novel strategy for treating BPs via the use of reusable and recyclable MCBs that are cheap, easy and fast to synthesise.

TFEB- and TFE3-dependent autophagy activation supports cancer proliferation in the absence of centrosomes.

Centrosome amplification is a phenomenon frequently observed in human cancers, so centrosome depletion has been proposed as a therapeutic strategy. However, despite being afflicted with a lack of centrosomes, many cancer cells can still proliferate, implying there are impediments to adopting centrosome depletion as a treatment strategy. Here, we show that TFEB- and TFE3-dependent autophagy activation contributes to acentrosomal cancer proliferation. Our biochemical analyses uncover that both TFEB and TFE3 are novel PLK4 (polo like kinase 4) substrates. Centrosome depletion inactivates PLK4, resulting in TFEB and TFE3 dephosphorylation and subsequent promotion of TFEB and TFE3 nuclear translocation and transcriptional activation of autophagy- and lysosome-related genes. A combination of centrosome depletion and inhibition of the TFEB-TFE3 autophagy-lysosome pathway induced strongly anti-proliferative effects in cancer cells. Thus, our findings point to a new strategy for combating cancer.Abbreviations: AdCre: adenoviral Cre recombinase; AdLuc: adenoviral luciferase; ATG5: autophagy related 5; CQ: chloroquine; DAPI: 4',6-diamidino-2-phenylindole; DKO: double knockout; GFP: green fluorescent protein; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LAMP2: lysosomal associated membrane protein 2; LTR: LysoTracker Red; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MITF: melanocyte inducing transcription factor; PLK4: polo like kinase 4; RFP: red fluorescent protein; SASS6: SAS-6 centriolar assembly protein; STIL: STIL centriolar assembly protein; TFEB: transcription factor EB; TFEBΔNLS: TFEB lacking a nuclear localization signal; TFE3: transcription factor binding to IGHM enhancer 3; TP53/p53: tumor protein p53.

Bacteria associated with an encrusting sponge (Terpios hoshinota) and the corals partially covered by the sponge.

Terpios hoshinota, a dark encrusting sponge, is known to be a competitor for space in coral reef environments, and facilitates the death of corals. Although numerous cyanobacteria have been detected in the sponge, little is known of the sponge-associated bacterial community. This study examined the sponge-associated bacterial community and the difference between the bacterial communities in the sponge and the coral partially covered by the sponge by analysis of 16S rRNA gene sequences of samples isolated from the sponge covering the corals Favia complanata, Isopora palifera, Millepora sp., Montipora efflorescens and Porites lutea. The sponge-associated bacterial community was mainly (61-98%) composed of cyanobacteria, with approximately 15% of these alphaproteobacteria and gammaproteobacteria, although the proportions varied in different sponge samples. The dominant cyanobacteria group was an isolated group closely related to Prochloron sp. The comparison of the bacterial communities isolated from sponge-free and the sponge-covered P. lutea showed that covering by the sponge caused changes in the coral-associated bacterial communities, with the presence of bacteria similar to those detected in black-band disease, suggesting the sponge might benefit from the presence of bacteria associated with unhealthy coral, particularly in the parts of the coral closest to the margin of the sponge.

Super-resolution microscopy reveals coupling between mammalian centriole subdistal appendages and distal appendages.

Subdistal appendages (sDAPs) are centriolar elements that are observed proximal to the distal appendages (DAPs) in vertebrates. Despite the obvious presence of sDAPs, structural and functional understanding of them remains elusive. Here, by combining super-resolved localization analysis and CRISPR-Cas9 genetic perturbation, we find that although DAPs and sDAPs are primarily responsible for distinct functions in ciliogenesis and microtubule anchoring, respectively, the presence of one element actually affects the positioning of the other. Specifically, we find dual layers of both ODF2 and CEP89, where their localizations are differentially regulated by DAP and sDAP integrity. DAP depletion relaxes longitudinal occupancy of sDAP protein ninein to cover the DAP region, implying a role of DAPs in sDAP positioning. Removing sDAPs alter the distal border of centrosomal γ-tubulins, illustrating a new role of sDAPs. Together, our results provide an architectural framework for sDAPs that sheds light on functional understanding, surprisingly revealing coupling between DAPs and sDAPs.