Sea Urchin Extracts Research Articles

Tropomyosin induces the synthesis of magnesian calcite in sea urchin spines

Calcium carbonate is present in many biominerals, including in the exoskeletons of crustaceans and shells of mollusks. High Mg-containing calcium carbonate was synthesized by high temperatures, high pressures or high molecular organic matter. For example, biogenic high Mg-containing calcite is synthesized under strictly controlled Mg concentration at ambient temperature and pressure. The spines of sea urchins consist of calcite, which contain a high percentage of magnesium. In this study, we investigated the factors that increase the magnesium content in calcite from the spines of the sea urchin, Heliocidaris crassispina. X-ray diffraction and inductively coupled plasma mass spectrometry analyses showed that sea urchin spines contain about 4.8% Mg. The organic matrix extracted from the H. crassispina spines induced the crystallization of amorphous phase and synthesis of magnesium-containing calcite, while amorphous was synthesized without SUE (sea urchin extract). In addition, aragonite was synthesized by SUE treated with protease-K. HC tropomyosin was specifically incorporated into Mg precipitates. Recombinant HC-tropomyosin induced calcite contained 0.1–2.5% Mg synthesis. Western blotting of sea urchin spine extracts confirmed that HC tropomyosin was present in the purple sea urchin spines at a protein weight ratio of 1.5%. These results show that HC tropomyosin is one factor that increases the magnesium concentration in the calcite of H. crassispina spines.

Sea urchin (Diadema savignyi) extract as a novel protective agent against cisplatin induced neurotoxicity in rats

Neurotoxicity is a severe side effect of platinum compounds used for cancer chemotherapy such as Cisplatin. This neurotoxicity leads to severe cognitive and nervous dysfunction, therefore, limiting the dose of Cisplatin and compromising the treatment protocol.The present study investigates the neuroprotective effect of Sea Urchins which is a marine animal known for its rich bioactive compounds. Male Sprague Dawley rats received Cisplatin (2 mg/kg body weight) for 4 weeks, two times per week, followed by Sea Urchin extracts (50 and 100 mg/kg body weight) twice weekly for 4 weeks.Results show that rats treated with Urchin’s extracts showed a significant improvement in the thermal (heat and cold) sensitivity compared to untreated rats. Liver enzymes Alanine Aminotransferase (ALT) and Aspartate Aminotransferase (AST) and Urea levels were also significantly decreased back to normal following treatment with sea urchin extracts. Brain tissue oxidative stress marker Nitric oxide (NO) and lipid peroxidation marker Malondialdehyde (MDA) increased significantly in the cisplatin-treated rats while the reduced glutathione levels (GSH) and catalase activity (CAT) showed a significant decrease. Treatment with sea Urchin extracts reversed these changes.Histological and immunohistochemical examination of the cerebral cortex reveled degenerative changes such as karyopyknosis and shrunken necrotic ghost like neurons in the cisplatin treated groups. There was also strong positive Glial fibrillary acidic protein (GFAP) reactivity and a negative B-cell leukemia/lymphoma 2 protein (Bcl2) reaction in most apparent neurons, indicating strong apoptotic changes. Treatment with Urchin extracts reversed these changes. Quantification of cerebral cortex neurons also revealed the strong effect of the extracts. Cisplatin treated groups showed 3708 cells/ mm3 compared to 8091 cells/mm3 in the normal rats. Extract treatment increased the neuronal numbers to almost normal levels. Quantification of the Immuno-histochemical expression of GFAP showed an increase by 10-folds after cisplatin administration. A remarkable decline from the cisplatin group was seen in the extract treated groups.In Conclusion, Sea Urchins extracts possess a strong neuroprotective activity and could provide a novel therapeutic method to prevent Cisplatin-induced neurotoxicity.

Identification of sea urchin gonads chemical compounds using thin-layer chromatography from Bokory island, Southeast Sulawesi

Southeast Sulawesi is one area that has a diversity of potential marine biota as medicinal ingredients, including the sea urchin gonads. Empirically coastal communities have been hereditary, consuming sea urchin gonads as a source of high protein. Considering its potential as a raw material for medicines, it is necessary to conduct a series of tests to guarantee its safety and efficacy. This study aims to determine the chemical compounds of sea urchin gonad extract (Diadema setosum L) by Thin-Layer Chromatography (TLC). The research method uses a Pre-experimental design with a one-shot case study model. Identification of metabolites of sea urchin gonad extracts was carried out in two stages: phytochemical screening and identification by TLC method by UV-Vis. Samples were dried by the Freeze-drying method to get 335.11 g of dry samples and 58.06 g of extracts. The results of the test of the content of water-soluble and ethanol-soluble sea urchin extract were 5.73% and 3.72%. The results showed that in phytochemical screening, ethyl acetate extracts of the positive marine gonads contained alkaloids, saponins, and steroids. In addition, UV-vis 254 nm and 366 nm TLC methods showed alkaloids and saponins. The Rf values are 0.4 and 0.68, respectively.

In vitro characterization of bioactive compounds extracted from sea urchin (Stomopneustes variolaris) using green and conventional techniques

This study investigated the in vitro bioactivities of extracts obtained from viscera, spines, shells, and gonads of Stomopneustes variolaris using subcritical water extraction (SWE) at 110 °C, 150 °C, 190 °C, and 230 °C and Soxhlet extraction. The highest amounts of phenolics (22.68 ± 0.05 mg GAE/g), flavonoids (27.11 ± 0.10 mg RE/g), and proteins (40.25 ± 0.84 mg BSA/g) were recorded from gonads at 230 °C, whereas maximum sugar content (23.38 ± 1.30 mg glucose/g) was in viscera at 150 °C. Gonads at 230 °C exhibited the highest DPPH activity (78.68 ± 0.18%), whereas viscera at 150 °C exhibited the highest ABTS+ (98.92 ± 1.27%) and protein denaturation inhibition activity (37.13 ± 9.94%). Viscera at 110 °C claimed the highest amylase inhibition (42.46 ± 0.83%), and spines at 150 °C had the highest anticancer activity (IC50 = 767.47 μg/mL). SWE achieved superior results in bioactive compound recovery and detected higher levels of bioactivities (p < 0.05). Results suggest processing sea urchin extracts via SWE has potential application to the food and pharmaceutical industries.

STANDARDISASI EKSTRAK ETANOL 96% BULU BABI Echinometra mathaei DARI PERAIRAN BANGKALAN

Echinometra mathaei sea urchins are known to contain chemical compounds that can be used to overcome health problems, one of which has antioxidant activity. Some previous studies have explained the benefits of sea urchins but to ensure the quality assurance of sea urchins extracts, efforts should be made to determine the quality standards of sea urchins extract. This study aims to standardize specific parameters and non-specific parameters on Echinometra mathaei sea urchins extract to ensure the quality of 96% ethanol extract. Samples were obtained from Rongkang Beach, Kwanyar District, Bangkalan District, Madura which were then extracted using maceration method with 96% ethanol solvent. The results of the study for specific parameters showed that organoleptically and macroscopically the extracts were thick, blackish brown in color and characteristic of sea urchins; levels of dissolved compounds in ethanol of 78.37%; levels of dissolved compounds in water of 93.09%; and chemical content tests showed that the extract contained steroid compounds, tannins, flavonoids, saponins, alkaloids and terpenoids. Determination of non-specific parameters showed that Echinometra mathaei sea urchin extract had a drying shrinkage of 3.03%; specific gravity of 0.8411 g/mL; water content of 5.32%; Pb heavy metal contamination was 7.28 mg/kg, Cd was &lt;0.0024 mg/kg, Hg was &lt;0.0002 mg / kg, Mg was 347.83 mg / kg; microbial contamination of &lt;10 colonies / mL; and yeast mold contamination of &lt;10 colonies/mL. The conclusion is Echinometra mathaei sea urchin extract has the potential to be developed into a medicinal preparation. Key word: Echinometra mathaei, Specific standardize parameters, non-specific standardize parameters, Extract thanol 96%.

Genital reversal of betta fish by immersion using steroid extract of sea urchins

Sea urchin, Diadema setosum is a potential fishery commodity with high economic value. It contains a steroid compound as an aphrodisiac in male monosexes (masculinization). In aquaculture usually used 17α-methyl testosterone, but it was difficult to decompose or it contaminated with carcinogens and pollutants. Therefore, it needs to be replaced with safer natural hormones from sea urchin. Betta sp. is an ornamental fish with high demand because of its colorful, diverse tail shapes, and price of male is higher than female, so that male cultivation is beneficial. This study aimed to utilize steroid compounds of sea urchin gonad extracts in masculinization of betta fish. In particular, it is to examine the steroid compounds by providing different doses and soaking times to the formation of male. The study was conducted using a completely randomized design method which was further classified into two stages. In first stage, the soaking dose was 0, 2, 4, 6 and 8 mg L-1 for 12 h, while in stage II, the immersion duration was 0, 12, 18, 24 and 30 h at a dose of 4 mg L-1 in larvae aged 2 weeks. Each treatment was repeated 3 times, and data were analyzed using Anova as well as with the LSD test at the level of 5%. The results showed that on the immersion in dose of 4 mg L-1, the highest male individual of 84.10% was formed. Also, a dose of 4 mg L-1 was significantly different from the control and 6 mg L-1, but it was not significantly different from that of 2 and 8 mg L-1. The difference in immersion time significantly affected the success of male monosex formation and the duration of 12 h in dose of 4 mg L-1 sea urchin extracts showed the highest percentage of 84.00%.

Behavioral analysis of rock bream Oplegnathus fasciatus reveals a strong attraction potential for sea urchin extracts

Monitoring fish movement is important to understand how physiology adapts to environmental change. To explore the applicability of a video tracking system for determining if chemical cues attract or repel aquatic animals, the movement patterns of rock bream, Oplegnathus fasciatus, were analyzed upon exposure to various materials, including extracts of sea urchin, rock worm, bait worm, krill, barley kernel, and commercial fish feed. Pellets were prepared by mixing freeze-dried tissues with a cellulose and corn flour mixture. Behavioral analysis was carried out with five fish that had been acclimated in the adaptation zone of a Y-shaped tank. Preference toward chemical cues was quantified by assessing the frequency rock bream were observed in a discrete zone around the test material located at the end of each arm and the duration each fish stayed in each zone. The analysis of fish movement upon exposure to commercial feed and barley kernel at each end, respectively, indicated a clear preference toward the feed relative to the barley kernel. Movement responses were further tested with pellets containing extracts of sea urchin, one of the species collected on a large scale, and other materials including krill and worms. A stronger preference toward sea urchin (100%) was observed based on the duration of stay in the test zone, compared to krill (90.1 ± 44.2%), bait worm (81.1 ± 39.1%), rock worm (73.7 ± 28.9%), and barley (63.9 ± 25.9%), under the conditions tested. A detailed comparison of rock bream movements toward each material revealed significant differences in frequency and duration, respectively, between pairs of test materials including krill (74 ± 29.8 and 375.6 ± 118.9) vs. rock worm (41.5 ± 18.7 and 160.2 ± 42.6), krill (86.3 ± 22.9 and 477.1 ± 84) vs. bait worm (36.2 ± 5.5 and 166.1 ± 50.7), and rock worm (45.9 ± 26.2 and 213.7 ± 100.1) vs. bait worm (34.6 ± 21.7 and 159.5 ± 98.5). Rock bream exhibited preference for the test materials in the following order: commercial fish feed &gt; sea urchin &gt; krill &gt; rock worm &gt; bait worm &gt; barley. The results suggest a higher potency of sea urchin extract as a rock bream fishing bait compared to the other materials that are used as commercial bait.

SKRINING FITOKIMIA DAN UJI AKTIVITAS ANTIBAKTERI EKSTRAK ETANOL BULU BABI (Diadema setosum) TERHADAP PERTUMBUHAN Staphylococcus aureus DI PERAIRAN DESA PELAUW

Sea urchin (Diadema setosum) is an aquatic biota that has high nutritional value. Bioactive compounds produced by sea urchins have the potential to be used as natural antibacterial compounds. This study aims to identify chemical compounds and determine the antibacterial activity of ethanol extract of sea urchins (Diadema setosum) against the growth of Staphylococcus aureus bacteria. The research method used is the agar diffusion method. The results of phytochemical screening of sea urchin extract (Diadema setosum) showed the presence of alkaloids and saponins. By using variations in the concentration of ethanol extract of sea urchin (Diadema setosum) at a concentration of 80% it had an inhibitory power of 18 mm, a concentration of 90% had an inhibitory power of 23 mm, a concentration of 100% had an inhibitory power of 28 mm and for a concentration of 110% had an inhibitory power of 35 mm. The positive control has a resistance of 30 mm and the negative control is 0 mm. The results showed that the ethanol extract of sea urchins (Diadema setosum) had a strong bacterial inhibitory activity against Staphylococcus aureus bacteria.

In vitro antibacterial activity and potential applications in food of sea urchin (Diadema setosum) from Cape of Palette, South Sulawesi

Marine invertebrates in support of his life's defence of sea predatory use an innate immune mechanism, namely the cellular component hemocytes by secreting the dissolved antimicrobial and cytotoxic substances. It shows that marine invertebrates are potential sources and promising antimicrobial compounds. This research was aimed at determining the antibacterial activity of sea urchin (Diadema setosum) extract against some bacterial isolates and its application to foodstuffs. The gonad and shell of sea urchin extracted by methanol and ethyl acetate and then separated by ultrasound-assisted extraction. Screening of antibacterial compound use gas chromatography-mass spectrometry (GC-MS), and disc diffusion method was followed to determine the antimicrobial activity against Salmonella, Escherichia coli and Staphylococcus aureus. The results of this study showed antibacterial activity against one or more strains. The gonad of sea urchin from methanol extract exhibited significant inhibitory effect and effective against Salmonella, E. coli and S. aureus. Majority of gonad and shell of sea urchin extract showed antibacterial activity against the tested strains. However, gonad of methanol extract was found to be inhibiting microorganisms gram-negative (E. coli) and gram-positive (Salmonella and S. aureus). Gonad extract can be a good source of antibacterial agents.

Preparation of the Persian Gulf Echinometra mathaei Organic Extracts and Investigation of Their Antibacterial Activity

Background: Marine organisms such as Echinoderms have secondary metabolites, which are antimicrobial naturally. Various extracts of echinoderms organs possess pharmacological activities, including anticancer, antimicrobial, antifungal, antiviral, and anti-inflammatory functions. Objectives: The purpose of this research was to assay the Persian Gulf sea urchin secondary metabolites for antimicrobial effectiveness. Methods: Sea urchins (Echinometra mathaei) were collected from the Boushehr tide coasts, Persian Gulf. Gonads, gut, tests, shell, and spines were carefully dissected from sea urchins, washed with tap water, and separated for the extraction procedure. All organs were extracted with 1:3 volumes (v/w) of methanol, chloroform, and n-hexane by maceration method for 72 hours at room temperature. The antimicrobial activity of the sea urchin tissue extracts was tested by well diffusion agar method. Results: All extracts of the sea urchin E. mathaei at 50 mg/mL concentration exhibited in vitro antimicrobial activity against Gram-negative and Gram-positive bacteria, including Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Bacillus subtilis, and two fungi, including Candida albicans and Aspergillus niger. The antibacterial activity of the sea urchin extracts varied with the solvent used for the bacterial strains. Conclusions: The results clearly showed the high antimicrobial activity of test and spines of the Persian Gulf sea urchin extracts against various Gram-positive and Gram-negative bacteria.

Bioactive potential of Diadema sp. from the South East Coast of Mauritius

As a consequence of the alarming rise in antimicrobial resistance, this study aimed at evaluating the bioactive potential of sea urchin extract, Diadema sp. Two different tissues; shells and gonads were used and two drying methods were compared; freeze drying (FD) and non-freeze drying (NFD). Statistically significant interaction was observed between the drying methods and solvents used. Compared to the gonadal tissue, NFD methanolic shell extract reported higher extraction yield, 9.13 ± 0.87%. Phenolic content assessed using the Folin-Ciocalteau method was higher in non-freeze dried (NFD) gonadal extract, 688.1 mg GAE/g DW. Promising antimicrobial activity was recorded using agar disc diffusion and results confirmed using MIC. Among the bacteria used, Escherichia coli and Staphylococcus aureus were more vulnerable, reporting inhibition zone of 16.3 ± 1.37 mm and 17.1 ± 0.23 mm using NFDSM and FDGM extract respectively. DPPH antioxidant assay showed NFDSM having better activity than the positive control, IC50 of 3.77 µg/ml. No significant larvicidal activity was recorded. Diadema sp are a potential source of novel antimicrobial compounds and suggest its possible use as natural food-grade antioxidant. This study also shows non-freeze drying method and methanol as the recommended solvent for maximal bioactivity.

Determination of In Vitro Antioxidant Properties, Anti-inflammatory Effects and A-Amylase Inhibition of Purple Sea Urchin Extract of Echinometra mathaei from the Persian Gulf

Background: Sea urchins contain high values of antioxidant substances such as polyphenols. Objectives: In the present study, the antioxidant activities, α-amylase inhibition, anti-inflammatory activity and the amounts of phenols and flavonoids of purple sea urchin Echinometra mathaei tissues obtained from Persian Gulf were examined. Methods: In December 2012, the purple sea urchin E. mathaei used for this study was harvested in the intertidal zone of Qeshm Island, Hormozgan Province, Iran. In the experimental design, extract isolation of different tissues of sea urchin such as spines, shells, gonads and Aristotle’s lantern was performed with the application of n-hexane, ethyl acetate and methanol solvents. Antioxidant properties of the samples were evaluated using different antioxidant tests such as total antioxidant capacity (TAC) and reducing power. The anti-diabetic and anti-inflammatory effects of samples were examined by inhibition of α-amylase and retardation of serum albumin denaturation, respectively. Results: With respect to antioxidant properties, the methanol extract of shell containing 40 μg/mL concentration possessed the maximum reducing power (OD = 0.475 ± 0.0049 and, ethyl acetate extract of Aristotle’s lantern at 1 mg/mL concentration presented the highest TAC (OD = 0.439 ± 0.0018). The n-hexane extract of gonad at a concentration of 1 mg/mL revealed the highest α-amylase inhibition (84.7 ± 2.92%). The ethyl acetate extract of Aristotle’s lantern at 20 μg/mL concentration possessed the highest anti-inflammatory property (69.78 ± 1.61 μg/mL). In biochemical analysis, the methanol extract of spine and ethyl acetate extract of Aristotle’ lantern had the highest phenolic (0.33 ± 0.017 mg/g) and flavonoid compounds (24.6 ± 0.72 mg/g), respectively. Conclusions: The obtained results revealed that sea urchin extracts could be used in pharmaceutical industries. Furthermore, its application in food industries as a supplementary food can be of great value.

Determination of In Vitro Antioxidant Properties, Anti-inflammatory Effects and A-Amylase Inhibition of Purple Sea Urchin Extract of Echinometra mathaei from the Persian Gulf

Determination of In Vitro Antioxidant Properties, Anti-inflammatory Effects and A-Amylase Inhibition of Purple Sea Urchin Extract of Echinometra mathaei from the Persian Gulf

Evaluation of antioxidant properties and biofunctions of polar, nonpolar, and water-soluble fractions extracted from gonad and body wall of the sea urchin Tripneustes gratilla

This study was performed to compare the various bioactivities of water-soluble, polar, and nonpolar extracts from the edible part (gonad) and waste (body wall) of the sea urchin Tripneustes gratilla. During antioxidant activity screening, these three extracts from both gonad and body wall showed various antioxidant activities in different tests. During human skin fibroblast (CCD966SK) viability and collagen screening, only water-soluble and polar extracts from gonad and body wall were observed to promote cell viability, while the water-soluble gonad extract promoted the collagen-generating activity of CCD966SK cells. The water-soluble and polar extracts from gonad and body wall showed proliferative activity towards different cell types. In contrast, the nonpolar extracts of gonad and body wall exerted antiproliferative effects on most tumor cells. These results indicate that the bioactivities of sea urchin extracts depend on the part of the urchin that the extract was taken from as well as the extraction method used.

Calcium- and polyphosphate-containing acidic granules of sea urchin eggs are similar to acidocalcisomes, but are not the targets for NAADP

Acidocalcisomes are acidic calcium-storage compartments described from bacteria to humans and characterized by their high content in poly P (polyphosphate), a linear polymer of many tens to hundreds of Pi residues linked by high-energy phosphoanhydride bonds. In the present paper we report that millimolar levels of short-chain poly P (in terms of Pi residues) and inorganic PPi are present in sea urchin extracts as detected using 31P-NMR, enzymatic determinations and agarose gel electrophoresis. Poly P was localized to granules randomly distributed in the sea urchin eggs, as shown by labelling with the poly-P-binding domain of Escherichia coli exopolyphosphatase. These granules were enriched using iodixanol centrifugation and shown to be acidic and to contain poly P, as determined by Acridine Orange and DAPI (4′,6′-diamidino-2-phenylindole) staining respectively. These granules also contained large amounts of calcium, sodium, magnesium, potassium and zinc, as detected by X-ray microanalysis, and bafilomycin A1-sensitive ATPase, pyrophosphatase and exopolyphosphatase activities, as well as Ca2+/H+ and Na+/H+ exchange activities, being therefore similar to acidocalcisomes described in other organisms. Calcium release from these granules induced by nigericin was associated with poly P hydrolysis. Although NAADP (nicotinic acid–adenine dinucleotide phosphate) released calcium from the granule fraction, this activity was not significantly enriched as compared with the NAADP-stimulated calcium release from homogenates and was not accompanied by poly P hydrolysis. GPN (glycyl-L-phenylalanine-naphthylamide) released calcium when added to sea urchin homogenates, but was unable to release calcium from acidocalcisome-enriched fractions, suggesting that these acidic stores are not the targets for NAADP.

Identification of a Major Yolk Protein as an Allergen in Sea Urchin Roe

Anaphylaxis after eating sea urchin roe has been reported. However, its major allergens have not yet been identified. The aim of this study was to identify the major allergens of sea urchin roe. Proteins of sea urchin roe were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and two-dimensional electrophoresis (2-DE). An immunoglobulin (Ig)E-binding protein was detected by immunoblotting using the patient's serum. An allergen isolated from 2DE-gel was identified by peptide mass fingerprinting using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry. Immunoblot analysis of sea urchin extracts showed that a 160-kDa protein at pI 6-7 was recognized by the patient's IgE. Peptide mass fingerprint analysis revealed that the protein was the major yolk protein (152 kDa, pI 6.9) of sea urchins. The results show that a major allergen of sea urchin roe is the major yolk protein.

Food allergy to Paracentrotus lividus (sea urchin roe)

Sea urchins are increasingly used as culinary products in Japanese and Korean cuisine throughout the world. To investigate a possible IgE-mediated allergy in a patient who experienced an anaphylactic reaction after eating sea urchin roe and mussels. A 40-year-old man experienced generalized pruritus and urticaria, shortness of breath, and wheezing 10 minutes after eating steamed mussels and boiled sea urchin roe. Investigations included skin prick tests, serum specific IgE determinations, oral challenge tests, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and immunoblotting. Results of skin prick tests with common aeroallergens were negative. Positive prick test responses were obtained with boiled and raw sea urchin extracts, boiled and raw sea urchins themselves, and boiled sea urchin water. Results of an oral challenge test with steamed mussels were negative. Five minutes after the oral mucosa contacted the boiled sea urchin extract, lip pruritus was present. Specific IgE concentrations to boiled sea urchin water and boiled and raw sea urchins surpassed the criterion for a positive result (1.1, 0.6, and 0.4 kU/L, respectively). In boiled sea urchin extract, a 118-kDa band appeared as the unique relevant IgE-binding antigen. In this patient with IgE-mediated food allergy to sea urchin roe, a 118-kDa protein seemed to be the antigen responsible for the reaction.

Transcription of the sea urchin U6 gene in vitro requires a TATA-like box, a proximal sequence element, and sea urchin USF, which binds an essential E box.

The tandemly repeated gene set encoding the sea urchin U6 gene has been cloned from the sea urchin Strongylocentrotus purpuratus. The U6 gene is transcribed by RNA polymerase III in a sea urchin nuclear extract. Like that of the vertebrate U6 genes, transcription of the sea urchin U6 gene does not require any internal sequences or 3' sequences but requires only 5' flanking sequences. Only 88 nucleotides of 5' flanking sequence are required for maximal expression in vitro. Mutagenesis experiments demonstrated the requirement for three elements, a CACGTG element at -80, a proximal sequence element at about -55, and the TATA-like box at -25. The major protein in sea urchin extracts that interacts with the CACGTG element is sea urchin USF, and immunodepletion of sea urchin USF greatly reduces transcription. The USF binding site in the U6 gene is highly homologous (11 of 13 nucleotides) with the USF binding sites found in the promoter of the S. purpuratus spec genes.

Transcription of the sea urchin U6 gene in vitro requires a TATA-like box, a proximal sequence element, and sea urchin USF, which binds an essential E box.

The tandemly repeated gene set encoding the sea urchin U6 gene has been cloned from the sea urchin Strongylocentrotus purpuratus. The U6 gene is transcribed by RNA polymerase III in a sea urchin nuclear extract. Like that of the vertebrate U6 genes, transcription of the sea urchin U6 gene does not require any internal sequences or 3' sequences but requires only 5' flanking sequences. Only 88 nucleotides of 5' flanking sequence are required for maximal expression in vitro. Mutagenesis experiments demonstrated the requirement for three elements, a CACGTG element at -80, a proximal sequence element at about -55, and the TATA-like box at -25. The major protein in sea urchin extracts that interacts with the CACGTG element is sea urchin USF, and immunodepletion of sea urchin USF greatly reduces transcription. The USF binding site in the U6 gene is highly homologous (11 of 13 nucleotides) with the USF binding sites found in the promoter of the S. purpuratus spec genes.

Sea urchin USF: A helix-loop-helix protein active in embryonic ectoderm cells

We previously characterized a DNA-binding factor in nuclear extracts of Strongylocentrotus purpuratus embryos that bound Spec gene promoters, was ectoderm specific, and had properties similar to the vertebrate transcription factor USF. Here we describe a cDNA clone, suUSF, isolated from an S. purpuratus cDNA library, with sequence homology to human USF. Spec gene promoter fragments formed sequence-specific complexes with suUSF, and antibodies against suUSF inhibited binding activity in nuclear extracts. Reaction of USF-site containing probes with filter-bound nuclear proteins demonstrated that suUSF binding activity was enriched in ectoderm cells, and immunoblotting showed a similar ectoderm enrichment. These data demonstrated that suUSF was responsible for the ectoderm-specific activity observed in sea urchin extracts.