The second anti-lipopolysaccharide factor (EsALF-2) with antimicrobial activity from Eriocheir sinensis

The anti-lipopolysaccharide factor CALF) is a small basic protein that can bind and neutralize lipopolysaccharide (LPS), mediating degranulation and activation of an intracellular coagulation cascade. In the present study, cDNA of the second Eriocheir sinensis ALF (designated as EsALF-2) was cloned and the full-length cDNA of EsALF-2 was of 724 bp, consisting of an open reading frame (ORF) of 363 bp encoding a polypeptide of 120 amino acids. The deduced amino acid of EsALF-2 shared 82% similarity with EsALF-1 from E. sinensis and about 53-65% similarity with ALFs from other crustaceans. The potential tertiary structures of EsALF-1 and EsALF-2 contained two highly conserved-cysteine residues to define the LPS binding site, but the N-terminal of EsALF-1 formed a single additional alpha-helix compared to EsALF-2, implying that EsALF-1 and EsALF-2 might represent different biological functions in E. sinensis. The mRNA transcript of EsALF-2 was detected in all examined tissues of healthy crabs, including haemocytes, hepatopancreas, gill, muscle, heart and gonad, which suggested that EsALF-2 could be a multifunctional molecule for the host immune defense responses and thereby provided systemic protection against pathogens. The mRNA expression of EsALF-2 was up-regulated after Listonelln anguillarum and Pichia pastoris challenge and the recombinant protein of EsALF-2 showed antimicrobial activity against L. anguillarum and P. pastoris. indicating that EsALF-2 was involved in the immune defense responses in Chinese mitten crab against L. anguillarum and P. pastoris. These results together indicated that there were abundant and diverse ALFs in E. sinensis with various biological functions and these ALFs would provide candidate promising therapeutic or prophylactic agents in health management and diseases control of crab aquaculture. (C) 2010 Elsevier Ltd. All rights reserved.

Cflec-5, a pattern recognition receptor in scallop Chlamys farreri agglutinating yeast Pichia pastoris

C-type lectins are a superfamily of carbohydrate-recognition proteins which play crucial roles as pattern recognition receptors (PRRs) in the innate immunity. In this study, the full-length cDNA of a C-type lectin was cloned from scallop Chlamys farreri (designated as Cflec-5) by expression sequence tag (EST) analysis and rapid amplification of cDNA ends (RACE) approach The full-length cDNA of Cflec-5 was of 1412 bp. The open reading frame encoded a polypeptide of 153 amino acids, including a signal sequence and a conserved carbohydrate-recognition domain with the EPN motif determining the mannose-binding specificity The deduced amino acid sequence of Cflec-5 showed high similarity to members of C-type lectin superfamily. The quantitative real-time PCR was performed to investigate the tissue distribution of Cflec-5 mRNA and its temporal expression profiles in hemocytes post pathogen-associated molecular patterns (PAMPs) stimulation. In healthy scallops, the Cflec-5 mRNA was mainly detected in gill and mantle, and marginally in other tissues The mRNA expression of Cflec-5 could be significantly induced by lipopolysaccharide (LPS) and glucan stimulation and reached the maximum level at 6 h and 12 h, respectively But its expression level did not change significantly during peptidoglycan (PGN) stimulation The function of Cflec-5 was investigated by recombination and expression of the cDNA fragment encoding its mature peptide in Escherichia coli Rosetta Gami (DE3) The recombinant Cflec-5 agglutinated Pichia pastoris in a calcium-independent way The agglutinating activity could be inhibited by D-mannose. LPS and glucan, but not by D-galactose or PGN. These results collectively suggested that Cflec-5 was involved in the innate Immune response of scallops and might contribute to nonself-recognition through its interaction with various PAMPs (C) 2010 Elsevier Ltd All rights reserved

Identification and molecular analysis of a novel C-type lectin from Scophthalmus maximus

C-type lectins are calcium-dependent carbohydrate-binding proteins that play Important roles in innate immunity In this study, a C-type lectin homologue (SmLec1) was identified from turbot (Scophthalmus maximus) and analyzed at expression and functional levels. The open reading frame of SmLec1 is 504 bp, with a 5'-untranslated region (UTR) of 101 bp and a 3'-UTR of 164 bp The deduced amino acid sequence of SmLec1 shares 34%-38% overall identities with the C-type lectins of several fish species In silico analysis identified in SmLec1 conserved C-type lectin features, including a carbohydrate-recognition domain, four disulfide bond-forming cysteine residues, and the mannose-type carbohydrate-binding motif In addition, SmLec1 possesses a putative signal peptide sequence and is predicted to be localized in the extracellular. Expression of SmLec1 was highest in liver and responded positively to experimental challenges with fish pathogens Recombinant SmLec1 (rSmLec1) purified from yeast was able to agglutinate the Gram-negative fish pathogen Listonella anguillarum but not the Gram-positive pathogen Streptococcus uncle The agglutinating ability of rSmLec1 was abolished in the presence of mannose and ethylenediaminetetraacetic acid and by elevated temperature (65 degrees C) Further analysis showed that rSmLec1 could stimulate kidney lymphocyte proliferation and enhance the killing of bacterial pathogen by macrophages Taken together, these results suggest that SmLec1 is a unique mannose-binding C-type lectin that possesses apparent immunomodulating property and is likely to be involved in host defense against bacterial infection (C) 2010 Elsevier Ltd. All rights reserved

Identification and analysis of a Scophthalmus maximus ferritin that is regulated at transcription level by oxidative stress and bacterial infection

Ferritins are conserved Iron storage proteins that exist in most living organisms and play an essential role in Iron homeostasis. In this study, we reported the identification and analysis a ferritin M subunit, SmFerM, from turbot Scophthalmus maximus. The full length cDNA of SmFerM contains a 5'-untranslated region (UTR) of 232 bp, an open reading frame (ORF) of 531 bp, and a 3'-UTR of 196 bp The ORF encodes a putative protein of 176 amino acids, which shares extensive sequence identities with the M terrains of several fish species. In silico analysis identified in SmFerM both the ferroxidase center of mammalian H ferritins and the iron nucleation site of mammalian L ferritins. Quantitative real time reverse transcriptase-PCR analysis indicated that SmFerM expression was highest in muscle and lowest in heart and responded positively to experimental challenges with bacterial pathogens and poly(I center dot C) Exposure of cultured turbot hepatocytes to treatment of stress inducers (iron, copper, and H2O2) significantly upregulated the expression of SmFerM in a dose dependent manner. Iron chelating analysis showed that recombinant SmFerM purified from Escherichia coli exhibited apparent iron binding activity. These results suggest that SmFerM is a functional M ferritin and is likely to play a role in iron sequestration and protection against oxidative stress and microbial infection (C) 2010 Elsevier Inc All rights reserved

Molecular cloning, characterization and expression of a novel serine proteinase inhibitor gene in bay scallops (Argopecten irradians, Lamarck 1819)

Serine protease inhibitors, critical regulators of endogenous proteases, are found in all multicellular organisms and play crucial roles in host physiological and immunological effector mechanisms. The first mollusk serine proteinase inhibitor (designated AISPI) cDNA was obtained from the bay scallop Argopecten irradians by randomly sequencing a whole tissue cDNA library and rapid amplification of cDNA ends (RACE). The full-length cDNA of the scallop serine protease inhibitor was 1020 bp, consisting of a 5'-terminal untranslated region (UTR) of 39 bp, a 3'-terminal UTR of 147 bp with a canonical polyadenylation signal sequence AATAAA and a poly(A) tail, and an open reading frame of 834 bp. The AISPI cDNA encoded a polypeptide of 278 amino acids with a putative signal peptide of 22 amino acids and a mature protein of 256 amino acids. The deduced amino-acid sequence of AISPI contained six tandem and homologous domains similar to that of Kazal-type serine protease inhibitors, including the conserved sequence C-X(7)-C-X(6)-Y-X(3)-C-X(2,3)-C and six cysteine residues responsible for the formation of disulfide bridges, indicating that the AISPI protein from bay scallop should be a member of the Kazal-type serine protease inhibitor family. The temporal expression of AISPI was measured by semi-quantitative RT-PCR after injury or bacterial challenge. After the adductor muscle was wounded or injected with Vibrio anguillarum, the expression of AISPI mRNA in hemolymph was up-regulated and reached the maximum level at 8 and 16 h, respectively, and then progressively dropped back to the original level. The results indicated that AISPI could play an important role in injury healing and immune response in mollusks as it could be induced by injury and bacterial challenge. (c) 2005 Elsevier Ltd. All rights reserved.

Molecular cloning and expression analysis of Crustin-like gene from Chinese shrimp Fenneropenaeus chinensis

A new member of antimicrobial protein genes of the Crustin family was cloned from haemocytes of the Chinese shrimp Fenneropenaeus chinensis by 3' and 5' RACE. The full-length cDNA of Crustin-like gene contains a 390 bp open reading frame, encoding 130 amino acids. The deduced peptide contains a putative signal peptide of 17 amino acids and mature peptide of 113 amino acids. The molecular mass of the deduced mature peptide is 12.3 ku. It is highly cationic with a theoretical isoelectric point of 8.5. The deduced amino acids sequence of this Crustin showed high homology with those of Penaeus (Litopenaeas) setferus. Northern blotting showed that the cloned Crustin gene was mainly expressed in haemocytes, gill, intestine, and RNA in situ hybridization indicated that the Crustin gene was constitutively expressed exclusively in haemocytes of these tissues. Capillary electrophoresis RT-PCR analysis showed that Crustin was up-regulated dramatically from 12 to 48 h after a brief decrease of mRNA during first 6 h in response to microbe infection. The level of Crustin mRNA began to restore at 72 h post-challenge. This indicated that Crustin gene might play an important role when shrimps are infected by bacterial pathogen.

Formation and evolution of the modern warm current system in the East China Sea and the Yellow Sea since the last deglaciation

To reconstruct the formation and evolution process of the warm current system within the East China Sea (ECS) and the Yellow Sea (YS) since the last deglaciation, the paleoceangraphic records in core DGKS9603, core CSH1 and core YSDP102, which were retrieved from the mainstream of the Kuroshio Current (KC), the edge of the modern Tsushima Warm Current (TWC) and muddy region under cold waters accreted with the Yellow Sea Warm Current (YSWC) respectively, were synthetically analyzed. The results indicate that the formation and evolution of the modern warm current system in the ECS and the YS has been accompanied by the development of the KC and impulse rising of the sea level since the last deglaciation. The influence of the KC on the Okinawa Trough had enhanced since 16 cal kyr BP, and synchronously the modern TWC began to develop with the rising of sea level and finally formed at about 8.5 cal kyr BP. The KC had experienced two weakening process during the Heinrich event 1 and the Younger Drays event from 16 to 8.5 cal kyr BP. The period of 7-6 cal kyr BP was the strongest stage of the KC and the TWC since the last deglaciation. The YSWC has appeared at about 6.4 cal kyr BP. Thus, the warm current system of the ECS and the YS has ultimately formed. The weakness of the KC, indicated by the occurrence of Pulleniatina minimum event (PME) during the period from 5.3 to 2.8 cal kyr BP, caused the main stream of the TWC to shift eastward to the Pacific Ocean around about 3 cal kyr BP. The process resulted in the intruding of continent shelf cold water mass with rich nutrients. Synchronously, the strength of the YSWC was relatively weak and the related cold water body was active at the early-mid stage of its appearance against the PME background, which resulted in the quick formation of muddy deposit system in the southeastern YS. The strength of the warm current system in the ECS and the YS has enhanced evidently, and approached to the modern condition gradually since 3 cal kyr BP.

Expression and localization of a novel phosducin-like protein from amphioxus Branchiostoma belcheri

A full length amphioxus cDNA, encoding a novel phosducin-like protein (Amphi-PhLP), was identified for the first time from the gut cDNA library of Branchiostoma belcheri. It is comprised of 1 550 bp and an open reading frame (ORF) of 241 amino acids, with a predicted molecular mass of approximately 28 kDa. In situ hybridization histochemistry revealed a tissue-specific expression pattern of Amphi-PhLP with the high levels in the ovary, and at a lower level in the hind gut and testis, hepatic caecum, gill, endostyle, and epipharyngeal groove, while it was absent in the muscle, neural tube and notochord. In the Chinese Hamster Ovary (CHO) cells transfected with the expression plasmid pEGFP-N1/Amphi-PhLP, the fusion protein was targeted in the cytoplasm of CHO cells, suggesting that Amphi-PhLP is a cytosolic protein. This work may provide a framework for further understanding of the physiological function of Amphi-PhLP in B. belcheri.

Benthic foraminifera and bottom water evolution in the middle-southern Okinawa Trough during the last 18 ka

A piston sediment core E017 from the middle-southern Okinawa Trough was investigated. A preliminary study of the deep-water evolution since 18 cal. ka BP was performed based on the quantitative census data of benthic foraminiferal fauna, together with planktonic foraminiferal oxygen and carbon isotope, AMS(14)C dating, and the previous results achieved in the southern Okinawa Trough. The result shows that the benthic fauna was dominated by Bulimina aculeata (d'Orbigny), Uvigerina peregrina (Cushman), Hispid Uvigerina and Uvigerina dirupta (Todd) during the glaciation-deglaciation before 9.2 cal. ka BP, while Epistominella exigua (Brady), Pullenia bulloides (d'Orbigny), Cibicidoides hyalina (Hofker), Sphaeroidina bulloides (d'Orbigny) and Globocassidulina subglobosa (Brady) predominated the fauna in the post-glacial period after 9.2 cal. ka BP. The benthic foraminifera accumulation rate (BFAR), paleoproductivity estimates and benthic foraminiferal assemblage conformably indicate that surface water paleoproductivity and organic matter flux during the glaciation-deglaciation were higher than those of the post-glacial period in the middle-southern Okinawa Trough, and gradually enhanced from the southern to the central Okinawa Trough during the glaciation-deglaciation, which could be caused by the discrepancy of the terrigenous nutrients supply. High abundances of E exigua, an indicator of pulsed organic matter input, after 9.2 cal. ka. BP may indicate that the intensity of seasonally riverine pulsed flux during the post-glacial period was stronger than that of the glaciation-deglaciation period, and the seasonal influx in the central trough might be stronger than in the south. The temporal distributions of the typical species indicating bottom water oxygen content and ventilation condition show that the ventilation of the bottom water during the post-glacial period is more active than the glaciation-deglaciation, which reflects that the evolution of the intermediate and deep waters of the northwestern Pacific during the last glaciation has no evident influence on the deep-water of the middle-southern Okinawa Trough. Additionally, the variations in agglutinated benthic foraminiferal abundance and other carbonate dissolution proxies indicate that carbonate dissolution gradually increased since the last 18 ka in the Okinawa Trough and rapidly enhanced at 9.2 cal. ka BP. The modern shallow carbonate lysocline could form at 3 cal. ka BP.

Recent 2000-year geological records of mud in the inner shelf of the East China Sea and their climatic implications

AXIS(14)C dating and grain-size analysis for Core DD2, located at the north of the Yangtze River-derived mud off the Zhejiang-Fujian coasts in the inner shelf of the East China Sea, provide us a high-resolution grain-size distribution curve varying with depth and time. Data in the upper mud layer of Core DD2 indicate that there are at least 9 abrupt grain-size increasing in recent 2000 years, with each corresponding very well with the low-temperature events in Chinese history, which might result from the periodical strengthening of the East Asian Winter Monsoon (EAWM), including the first-revealed maximum temperature lowering event at around 990 a BP. At the same time, the finer grain size section in Core DD2 agrees well with the Sui-Tang Warming Period (600-1000 a AD) defined previously by Zhu Kezhen, during which the climate had a warm, cold and warm fluctuation, with a dominated cooling period of 750-850 a AD. The Little Ice Age (LIA) can also be identified in the core. It starts around 1450 a AD and was followed by a subsequent cooling events at 1510, 1670 and 1840 a AD. Timing of these cold events revealed here still needs to be further verified owing to some current uncertainty of dating we used in this study.

Heinrich event imprints in the Okinawa Trough: evidence from oxygen isotope and planktonic foraminifera

Planktonic foraminiferal faunas, oxygen isotope and modern analog technique sea surface temperature records were obtained in piston core DGKS9603 (28degrees08.869'N, 127degrees16.238'E, water depth 1100 in) collected from the middle Okinawa Trough. During the last glaciation, four cold events were identified and correlate Heinrich events (HE) H2-5 of the last 45 ka. During the last deglaciation, core DGKS9603 has begun to be influenced by the Kuroshio since about 16 cal ka BP. Three weakenings of this warm current occurred at about 2.8-5.3, 11.4 and 15.5 cal ka BP respectively. Among the three fluctuations, the oldest one is synchronous with HE1 and could be a response to the strong cooling observed in the North Atlantic Ocean. The fluctuation occurring at about 11.4 cal ka ago corresponds to the Younger Dryas within the age error bars. Our observations provide new evidence that the HEs documented from Greenland and the northern North Atlantic had a global climatic impact. Changes in the intensity of the East Asian monsoon could be the main mechanism responsible for the paleoccanographic variations observed in the Okinawa Trough. (C) 2001 Elsevier Science B.V. All rights reserved.

PALEOMONSOONS OF CHINA OVER THE LAST 130,000 YEARS - PALEOMONSOON VARIATION

Based upon the effect of land-sea interaction on the paleomonsoon variation and the time series of climatic proxy-indicators, the historical Asian monsoon variation over the last 130,000 and 18,000 years has been reconstructed with an emphasis on the basic characteristics of summer monsoon circulation. The monsoon-climatic cycles and associated model of environmental development over the central and eastern China are proposed and the mechanism of paleomonsoon variation of China preliminarily discussed. The variation of East Asian monsoon circulation should be regarded as a regional result of both solar-radiation changes and the global glacial-interglacial cycles. The episodic uplifting of the Qinghai-Xizang Plateau since the late Miocene has to a large extent controlled the forming and evolution of the paleomonsoon circulation of China.

A Holocene paleotemperature record based on radiolaria from the northern Okinawa Trough (East China Sea)

Using a radiolarian-based transfer function, mean annual sea surface temperature (SST) and seasonal temperature range are reconstructed through the last 10,500yrs in the northern Okinawa Trough. Down-core SST estimates reveal that throughout the Holocene the changes of mean annual SST display a three-step trend: (i) an early Holocene continuous warming between 10,500 and 8500yr BP which ends up with a abrupt cooling at about 8200yr BP; (ii) a relatively stable middle Holocene with high SST that lasted until 3200yr BP; and (iii) a late-Holocene distinct SST decline between 3200 and 500yr BP. This pattern is in agreement with the ice core and terrestrial paleoclimatic records in the Chinese continent and other regions of the world. Five cooling events with abrupt mean annual SST drops, which occur at similar to 300-600, 1400, 3100, 4600-5100 and 8200yr BP, are recognized during the last 10,500yrs. Comparison of our results with records of GISP2 ice core and marine sediment in North Atlantic region suggests these cooling events are strongly coupled, which implies a possible significant climatic correlation between high- and low-latitude areas. (C) 2007 Elsevier Ltd and INQUA. All rights reserved.

Evolution and variation of the Tsushima warm current during the late Quaternary: Evidence from planktonic foraminifera, oxygen and carbon isotopes

The evolution and variation history of the Tsushima warm current during the late Quaternary was reconstructed based on the quantitative census data of planktonic foraminiferal fauna, together with oxygen and carbon isotope records of mixed layer dweller a ruber and thermocline dweller N. dutertrei in piston core CSH1 and core DGKS9603 collected separately from the Tsushima warm current and the Kuroshio dominated area. The result showed that the Tsushima warm current vanished in the lowstand period during 40-24 cal ka BP, while the Kuroshio still flowed across the Okinawa Trough, arousing strong upwelling in the northern Trough. Meanwhile, the influence of freshwater greatly increased in the northern Okinawa Trough, as the broad East China Sea continental shelf emerged. The freshwater reached its maximum during the last glacial maximum (LGM), when the upwelling obviously weakened for the lowest sea-level and the depression of the Kuroshio. The modern Tsushima warm current began its development since 16 cal ka BP, and the impact of the Kuroshio increased in the middle and northern Okinawa Trough synchronously during the deglaciation and gradually evolved as the main water source of the Tsushima current. The modern Tsushima current finally formed at about 8.5 cal ka BP, since then the circulation structure has been relatively stable. The water of the modern Tsushima current primarily came from the Kuroshio axis. A short-term wiggle of the current occurred at about 3 cal ka BP, probably for the influences from the enhancement of the winter monsoon and the depression of the Kuroshio. The cold water masses greatly strengthened during the wiggle.

Flux and fate of Yangtze river sediment delivered to the East China Sea

Numerous cores and dating show the Yangtze River has accumulated about 1.16 x 10(12) t sediment in its delta plain and proximal subaqueous delta during Holocene. High-resolution seismic profiling and coring in the southern East China Sea during 2003 and 2004 cruises has revealed an elongated (similar to 800 km) distal subaqueous mud wedge extending from the Yangtze River mouth southward off the Zhejiang and Fujian coasts into the Taiwan Strait. Overlying what appears to be a transgressive sand layer, this distal clinoform thins offshore, from similar to 40 in thickness between the 20 and 30 m water depth to < 1-2 in between 60 and 90 m water depth, corresponding to an across shelf distance of less than 100 km. Total volume of this distal mud wedge is about 4.5 x 10(11) m(3), equivalent to similar to 5.4 x 10(11) t of sediment. Most of the sediment in this mud wedge comes from the Yangtze River, with some input presumably coming from local smaller rivers. Thus, the total Yangtze-derived sediments accumulated in its deltaic system and East China Sea inner shelf have amounted to about 1.7 x 10(12) t. Preliminary analyses suggest this longshore and across-shelf transported clinoform mainly formed in the past 7000 yrs after postglacial sea level reached its mid-Holocene highstand, and after re-intensification of the Chinese longshore current system. Sedimentation accumulation apparently increased around 2000 yrs BP, reflecting the evolution of the Yangtze estuary and increased land erosion due to human activities, such as fanning and deforestation. The southward-flowing China Coastal Current, the northward-flowing Taiwan Warm Current, and the Kuroshio Current appear to have played critical roles in transporting and trapping most of Yangtze-derived materials in the inner shelf, and hence preventing the sediment escape into the deep ocean. (c) 2006 Elsevier B.V. All rights reserved.