Curdepsidone A Induces Intrinsic Apoptosis and Inhibits Protective Autophagy via the ROS/PI3K/AKT Signaling Pathway in HeLa Cells.

Among female oncology patients, cervical cancer stands as the fourth most prevalent malignancy, exerting significant impacts on their health. Over 600,000 women received the diagnosis of cervical cancer in 2020, and the illness claimed over 300,000 lives globally. Curdepsidone A, a derivative of depsidone, was isolated from the secondary metabolites of Curvularia sp. IFB-Z10. In this study, we revised the molecular structure of curdepsidone A and investigated the fundamental mechanism of the anti-tumor activity of curdepsidone A in HeLa cells for the first time. The results demonstrated that curdepsidone A caused G0/G1 phase arrest, triggered apoptosis via a mitochondrial apoptotic pathway, blocked the autophagic flux, suppressed the PI3K/AKT pathway, and increased the accumulation of reactive oxygen species (ROS) in HeLa cells. Furthermore, the PI3K inhibitor (LY294002) promoted apoptosis induced by curdepsidone A, while the PI3K agonist (IGF-1) eliminated such an effect. ROS scavenger (NAC) reduced curdepsidone A-induced cell apoptosis and the suppression of autophagy and the PI3K/AKT pathway. In conclusion, our results revealed that curdepsidone A hindered cell growth by causing cell cycle arrest, and promoted cell apoptosis by inhibiting autophagy and the ROS-mediated PI3K/AKT pathway. This study provides a molecular basis for the development of curdepsidone A as a new chemotherapy drug for cervical cancer.

Phragmalin-Type Limonoids from the Fruits of Chukrasia tabularis and Their Anti-Inflammatory Activity.

Phytochemical investigation on the fruits of C. tabularis led to the isolation of five new phragmalin-type limonoids (1-5) and four known ones (6-9). The structures of the new compounds 1-5, named chuktabamalins A-E, were elucidated via spectroscopic techniques (HRESIMS, 1D and 2D NMR) and were comparable with the literature data of known compounds. In addition, new compounds were evaluated for in vitro anti-inflammatory activity. Compounds 1, 2, 3 and 5 showed moderate anti-inflammatory activity with IC50 values of 21.72 ± 2.79, 23.29 ± 1.00, 47.08 ± 3.47 and 66.67 ± 2.89 µM, respectively.

Adsorption Characteristics and Enrichment of Emodin from Marine-Derived Aspergillus flavipes HN4-13 Extract by Macroporous Resin XAD-16.

Emodin, a hydroxyanthraquinone derivative, has been used as medicine for more than 2000 years due to its extensive pharmacological activities. Large-scale production of emodin has been achieved by optimizing the fermentation conditions of marine-derived Aspergillus flavus HN4-13 in a previous study. However, the fermentation broth contained complex unknown components, which adversely affected the study of emodin. Herein, the conditions for the enrichment of emodin from A. flavipes HN4-13 extract using XAD-16 resin were optimized, and a separation method with high efficiency, simple operation, a low cost, and a large preparative scale was established. The adsorption process of emodin on the XAD-16 resin conformed to pseudo-second-order kinetics and Langmuir models. The optimal conditions for the adsorption process were as follows: An emodin concentration, flow rate, and loading volume of 0.112 mg/mL, 2 BV/h, and 10 BV, respectively. For desorption, 50% ethanol was used to elute impurities and 80% ethanol was used to desorb emodin. After enrichment with XAD-16 resin, the emodin content increased from 1.16% to 11.48%, and the recovery rate was 75.53% after one-step treatment. These results demonstrate the efficiency of the simple adsorption-desorption strategy, using the XAD-16 resin for emodin enrichment.

Enhancement of Emodin Production by Medium Optimization and KH2PO4 Supplementation in Submerged Fermentation of Marine-Derived Aspergillus favipes HN4-13.

Emodin is a widely distributed anthraquinone derivative with a variety of biological activities, one that can be efficiently produced by marine-derived fungus Aspergillus favipes HN4-13. However, its relatively low fermentation yield limits further development and pharmaceutical research work. In this study, Plaekett-Burman design and central composite design were adopted to optimize the fermentation conditions of A. favipes HN4-13. Optimal fermentation conditions in a 250-mL Erlenmeyer flask with 50 mL of medium were 59.3 g/L soluble starch, 10 g/L yeast extract paste, 30 g/L seawater salt, 1.04 g/L KH2PO4, 0.05 g/L MgSO4·7H2O, 0.01 g/L FeSO4·7H2O, seed culture 24 h, pH 5, inoculum size 18%, culture temperature 32 °C, and shaking at 160 rpm/min for 7 days. The production of emodin could achieve 132.40 ± 3.09 mg/L, with no significant difference from the predicted value (132.47 mg/L). Furthermore, KH2PO4 supplementation strategy was employed to regulate the mycelial morphology, upregulate the transcriptional level of biosynthesis gene cluster, and enhance emodin production (185.56 ± 4.39 mg/L).

Bioactive Indole Diketopiperazine Alkaloids from the Marine Endophytic Fungus Aspergillus sp. YJ191021.

Six new prenylated indole diketopiperazine alkaloids, asperthrins A-F (1-6), along with eight known analogues (7-14), were isolated from the marine-derived endophytic fungus Aspergillus sp. YJ191021. Their planar structures and absolute configurations were elucidated by HR-ESI-MS, 1D/2D NMR data, and time-dependent density functional theory (TDDFT)/ECD calculation. The isolated compounds were assayed for their inhibition against three agricultural pathogenic fungi, four fish pathogenic bacteria, and two agricultural pathogenic bacteria. Compound 1 exhibited moderate antifungal and antibacterial activities against Vibrioanguillarum, Xanthomonas oryzae pv. Oryzicola, and Rhizoctoniasolani with minimal inhibitory concentration (MIC) values of 8, 12.5, and 25 µg/mL, respectively. Furthermore, 1 displayed notable anti-inflammatory activity with IC50 value of 1.46 ± 0.21 µM in Propionibacteriumacnes induced human monocyte cell line (THP-1).

DMC triggers MDA-MB-231 cells apoptosis via inhibiting protective autophagy and PI3K/AKT/mTOR pathway by enhancing ROS level.

DMC, a kind of compound derived from the dry flower buds of Cleistocalyx operculatus, has been shown to inhibit the growth of various cancer cells, but research on triple-negative breast cancer cells remains scarce. To explore this issue, MDA-MB-231 cells were selected, and the results showed that DMC has strong proliferation inhibit effects on this kind of cells. The inhibit rate of 30 µM DMC incubated for 24 h was 56.25%, and 40.6% cells were arrested under the G2/M phase. The levels of pro-apoptosis protein Bax and active caspase-3, cleaved PARP and cell cycle related proteins, such as p21 and p27 increased, but apoptosis regulators, like Bcl-2, Cdc 2, Cyclin B1, and LC3 II decreased dramatically. In addition, DMC induced the accumulation of autophagosomes and autophagic substrates, and the combination of DMC with CQ promoted apoptosis of MDA-MB-231 cells, which suggested that DMC induced apoptosis partly by blocking autophagy flow. Moreover, the phosphorylation levels of phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and its mechanistic target of rapamycin kinase (mTOR) were also decreased after 30 µM DMC incubating for 24 h. The proteins play a critical role in cell proliferation, apoptosis, and autophagy modulation. The inhibition of autophagy flow and PI3K/AKT/mTOR pathway could be reversed after being treated with ROS scavenger NAC. Altogether, the results of the present study suggest that DMC effectively induces apoptosis and growth inhibition in MDA-MB-231 cells through blocking autophagy flow and regulating the PI3K/AKT/mTOR pathway by increasing ROS level.

Octahydroindolizine alkaloid Homocrepidine A from Dendrobium crepidatum attenuate P. acnes-induced inflammatory in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Dendrobium crepidatum Lindl. ex Paxton is a perennial epiphyte of Dendrobium genus, distributed in southern China, and utilized as the traditional Chinese medicine "Shihu" in Yunnan Province. Due to its heat-clearing and detoxicating properties, it is formulated as the "XiaoCuoWan" as recorded in the China Pharmacopoeia, and specially used to treat chronic skin inflammatory diseases, such as acne. AIM OF THE STUDY: This research aimed to estimate impact of the octahydroindoline alkaloid Homocrepidine A (HCA), isolated from D. crepidatum, on acne inflammation using both human THP-1 cells and mouse models. Furthermore, the potential anti-inflammatory mechanism of HCA has been analyzed through molecular biology methods and computer simulation. MATERIALS AND METHODS: THP-1 cells and mouse models induced by live Propionibacterium acnes (P. acnes) were employed to evaluate the anti-inflammatory properties of crude extract of D. crepidatum (DCE) and HCA. ELISA was utilized to detect the release of inflammatory cytokines in both cellular and murine ear tissues. RNAseq was used to screen the pathways associated with HCA-mediated inflammatory inhibition, while Western blot, RT-qPCR, and immunofluorescence were utilized to detect the expression of relevant proteins. Additionally, molecular docking simulations and cellular thermal shift assays were employed to confirm the target of HCA. RESULTS: Our research shows that DCE and HCA can effectively alleviate acne inflammation. HCA inhibits TLR2 expression by interacting with amino acid residues in the TIR domain of hTLR2, including Pro-681, Asn-688, Trp-684, and Ile-685. Moreover, HCA disrupts inflammatory signal transduction mediated by MAPK and NF-κB pathways through MyD88-dependent pathway. Additionally, HCA treatment facilitates Nrf2 nuclear translocation and upregulates HO-1 expression, thereby inhibiting NLRP3 inflammasomes activation. In vivo experiments further revealed that HCA markedly attenuated erythema and swelling caused by P. acnes in mice ears, while also decreasing the expression of pro-inflammatory cytokines IL-1ß and IL-8. CONCLUSIONS: Our research highlights the protective effects of D. crepidatum and its bioactive compound HCA against acne inflammation, marking the first exploration of its potential in this context. The discoveries indicate that HCA treatment may represent a promising functional approach for acne therapy.

Four previously undescribed diketopiperazines from marine fungus Aspergillus puniceus FAHY0085 and their effects on liver X receptor α.

Four previously undescribed diketopiperazine-type alkaloids including one oxepin-containing diketopiperazine-type alkaloid, oxepinamide L (1), three 4-quinazolinone alkaloids, puniceloids E-G (10-12), together with 12 known analogues, protuboxepin D (2), oxepinamides D-G, J-K and I (3-9), puniceloids B-D (13-15) and protubonine B (16), were isolated from the culture of the marine-derived fungus Aspergillus puniceus FAHY0085. The structures of the previously undescribed compounds were comprehensively elucidated by detailed interpretation of their NMR and HRESIMS data. Their absolute configurations were unambiguously determined by ROESY experiments, Marfey's method, calculated ECD experiments and single-crystal X-ray diffraction analysis. Compounds (3-4, 6-8, 14-15) were evaluated for their cytotoxic activity against HepG2, MCF-7, SW1116 and HeLa cells and compound 6 and 14 showed moderate cytotoxic activity against HeLa cells with IC50 49.61 ± 2.91 and 28.38 ± 1.57 µM, respectively. Compounds (1-8, 11-15) were screened for their transcriptional activation of liver X receptor α and compound 11 with known compounds 13-15 showed significant transcriptional activation of liver X receptor α with EC50 values in the range 2-50 µM.

Efficient ilamycins production utilizing Enteromorpha prolifera by metabolically engineered Streptomyces atratus.

With the invasion of green tides and the increase of urban green areas worldwide, multimillion tons of Enteromorpha need to be reutilized. In this study, Enteromorpha prolifera powder is considered a promising biomass resource for the production of commercial chemical products production. Ilamycins, novel cyclic heptapeptides with significant anti-TB activities, are isolated from Streptomyces atratus SCSIO ZH16, a deep-sea-derived strain. Using EP powder as a nitrogen source, the production of ilamycins reached 709.97 mg/L through optimization of the nitrogen source using the engineered strain S. atratus SCSIO ZH16 ΔR. After mutant strain constructions and tests, strain S. atratus SCSIO ZH16 ΔR::bldD EP powder achieved a higher production titer of ilamycins. Furthermore, the production titer of ilamycins and ilamycin E reached 1561.77 mg/L and 745.44 mg/L, respectively, in a 5 L bioreactor. This study suggests that E. prolifera is a promising and eco-friendly nitrogen source for the production of ilamycins.

Hypericin-mediated photodynamic therapy enhances gemcitabine induced Capan-2 cell apoptosis via inhibiting NADPH level.

OBJECTIVES: The combination of gemcitabine (Gem) and hypericin (HY) enhances the apoptosis of Capan-2 cells, providing a promising option for the treatment of pancreatic cancer. Our study further explored the cytotoxic mechanism of HY combined with chemotherapy drugs on pancreatic cancer. METHODS: The proliferation rate of the cells assayed with the MTT method. The ROS (reactive oxygen species) levels of each treatment were evaluated by DCFH-DA oxidisation methods. The activity of glutathione reductase and the levels of reduced glutathione (GSH) and oxidised glutathione (GSSG) were assessed using assay kits. The expression levels of apoptosis-related proteins were analysed by western blotting. KEY FINDINGS: The activity of glucose-6-phosphate dehydrogenase (G6PDH), a key enzyme of the pentose phosphate pathway, significantly decreased in Gem + HY groups, however, the ROS level enhanced accompanying with GSH depleting, mitochondrial membrane depolarisation and cytochrome C release. Gem + HY inhibits the expression of Bcl-2 but stimulates Bax level, triggering caspase activation and PARP cleavage and thus promoted apoptosis of Capan-2 cells. CONCLUSIONS: We demonstrated that Gem combined HY-PDT could inhibit the proliferation of Capan-2 cells and induce cell apoptosis. HY-PDT combined with Gem had a great potential on pancreatic cancer treatment clinically.

Development of a High-Titer Culture Medium for the Production of Cholesterol by Engineered Saccharomyces cerevisiae and Its Fed-Batch Cultivation Strategy.

Steroids are a class of compounds with cyclopentane polyhydrophenanthrene as the parent nucleus, and they usually have unique biological and pharmacological activities. Most of the biosynthesis of steroids is completed by a series of enzymatic reactions starting from cholesterol. Synthetic biology can be used to synthesize cholesterol in engineered microorganisms, but the production of cholesterol is too low to further produce other high-value steroids from cholesterol as the raw material and precursor. In this work, combinational strategies were established to increase the production of cholesterol in engineered Saccharomyces cerevisiae RH6829. The basic medium for high cholesterol production was selected by screening 8 kinds of culture media. Single-factor optimization of the carbon and nitrogen sources of the culture medium, and the addition of calcium ions, zinc ions and citric acid, further increased the cholesterol production to 192.53 mg/l. In the 5-L bioreactor, through the establishment of strategies for glucose and citric acid feeding and dissolved oxygen regulation, the cholesterol production was further increased to 339.87 mg/l, which was 734% higher than that in the original medium. This is the highest titer of cholesterol produced by microorganisms currently reported. The fermentation program has also been conducted in a 50-L bioreactor to prove its stability and feasibility.

Combinatorial strategies for production improvement of anti-tuberculosis antibiotics ilamycins E1/E2 from deep sea-derived Streptomyces atratus SCSIO ZH16 ΔilaR.

Ilamycins E1/E2 are novel cyclic heptapeptides from Streptomyces atratus SCSIO ZH16, which have the MIC value of 9.8 nM against Mycobacterium tuberculosis H37Rv. However, the lower fermentative titer of ilamycins E1/E2 cut off further development for novel anti-TB lead drugs. In order to break the obstacle, the combinatorial strategy of medium optimization, fermentative parameters optimization, exogenous addition of metal ions, precursors, and surfactants was developed to promoted the production of ilamycins E1/E2. Addition of 1 mM ZnCl2 at 0 h, 1 g/L tyrosine at 96 h, and 2 g/L shikimic acid at 48 h increased the production of ilamycins E1/E2 from 13.51 to 762.50 ± 23.15, 721.39 ± 19.13, and 693.83 ± 16.86 mg/L, respectively. qRT-PCR results showed that the transcription levels of key genes in Embden-Meyerhof-Parnas pathway, hexose phosphate shunt pathway, and shikimic acid pathway were upregulated. In addition, the production of ilamycins E1/E2 reached 790.34 mg/L in a 5-L bioreactor by combinatorial strategy. Combinatorial strategies were used for improving ilamycins E1/E2 production in S. atratus ΔilaR and provided a sufficient basis on further clinic development.

Bioactive indole alkaloids from insect derived endophytic Aspergillus lentulus.

As part of our program to discover new bioactive agents from endophytic fungi, three new indole alkaloids (1-2, 4) along with twelve known compounds were isolated from an inset derived endophytic strain Aspergillus lentulus. Their structures were determined by comprehensive spectroscopic analyses of 1D/2D NMR and HR-ESI-MS data. The absolute configurations were confirmed by ECD calculation using Time-dependent Density functional theory (TD-DFT) at the B3LYP/6-31 + g (d, p) level and Rh2(OCOCF3)4-induced ECD experiments. Compounds 2, 4, 5, 13 and 15 exhibited moderate cytotoxic effects on A549 cell line with IC50 in the range of 17.92-48.29 µM. Compounds 1, 2 and 13-15 displayed the anti-bacterial activity against Xanthomonas oryzae pv. oryzae and Xanthomonas oryzae pv. oryzicola with MIC values ranging from 25 to 100 µg/mL.

Hypericin Enhances Paclitaxel-Induced B16-F10 Cell Apoptosis by Activating a Cytochrome c Release-Dependent Pathway.

The enhanced inhibitory effect of paclitaxel (PTX) combined with hypericin (HY) on B16-F10 cells may be realized through the ROS-related cytochrome c release pathway. The apoptotic characteristics of the B16-F10 cells, such as DNA fragmentation, chromatin condensation, and apoptotic body formation, were all enhanced in the combined treatment group. Further investigation showed that the combination of paclitaxel and HY could increase the level of mitochondrial damage and the concentration of cytochrome c, causing the expression of caspase-3 and the cleavage of PARP.. Compared with paclitaxel or HY alone, the level of reactive oxygen species (ROS) increased significantly, while glutathione reductase (GR) activity and intracellular glutathione (GSH) levels decreased significantly in the combination group.

Association study of six activity SNPS in adrenal steroid hormone metabolism and IBM related genes with precocious puberty in Chinese girls.

OBJECTIVES: In order to investigate whether the variances in the candidate genes (Insulin receptor substrate IRS-1, beta3-adrenergic receptor ADRB3, 3 beta-hydro-xysteroid dehydrogenase HSD3B2, glucocorticoid receptor GRL and 21-hydroxylase CYP21) which affect the metabolism of adrenal steroids hormone and body composition, are associated with precocious puberty in Chinese girls. METHODS: PCR-RFLP analysis method was applied in the typing of six activity SNPs in five genes in two groups: 176 precocious puberty girls as case and 192 healthy girls as control. RESULTS: The typing results showed that the frequencies of the allele CYP21 L282 polymorphism were 32.7% and 32.6% in case and control groups (P=0.518), respectively. For IRS-1 R972, they were 0.6% in cases and 2.3% in controls (P=0.043), for ADRB3 R64, 13.6% in cases and 12% in controls (P=0. 378), and for GRL S363, 7.4% in cases and 5.73% in controls (P=0.335). No sample in the two groups carries the variation of CYP21 I172N and HSD3B2 L236S. CONCLUSION: Among these six activity SNPs in five candidate genes, IRS-1 972R was statistically associated with the onset time of puberty in Chinese girls. In order to confirm whether the candidate genes have any other activity SNPs that are associated with the onset time of puberty in Chinese girls, resequencing of these candidate genes is needed in following time.

Association study of four activity SNPs of CYP3A4 with the precocious puberty in Chinese girls.

CYP3A4 plays an important role in the metabolism of a variety of endogenous and exogenous compounds. Earlier studies revealed that a high-activity variation of the CYP3A4 gene, CYP3A4*1B is significantly associated with the precocious puberty in girls. Other three variations, CYP3A4*4, CYP3A4*5 and CYP3A4*6, which were found in a study carried out in a Chinese population in Taiwan, were reported to down-regulate the enzymatic activity of CYP3A4. The four activity SNPs were typed in our study in two groups of Chinese girls: 176 girls with precocious puberty as the case group, and 192 normal girls as the control group. No variations of CYP3A4*1B and CYP3A4*4 were found in all the cases and controls. Heterozygous of CYP3A4*5 was found in five subjects of the 192 controls but none in the cases, heterozygous of CYP3A4*6 was found in two subjects of the controls but none in the cases. Fisher's exact test showed that the variation of CYP3A4*5 was associated with the onset of puberty in Chinese girls (P-trend=0.038), while the variation of CYP3A4*6 was not associated with the onset of puberty in Chinese girls (P-trend=0.272). The result suggests that these mutations in the CYP3A4 gene have no contribution to the early onset of puberty in Chinese girls, but are related in some way with the puberty development in Chinese girls.

Improving the bioactivity of rHirudin with boronophenylalanine site-specific modification.

To improve the bioactivity of recombinant (r)Hirudin, the orthogonal pair MjBTyrRS/tRNATyr cua (made up of the boronophenylalanine, tRNA and tRNA synthetase), was selected to incorporate boronophenylalanine site­specifically into rHirudin at the 63 sites in an Escherichia coli system in response to the TAG codon. Following fusion with the gIII signal peptide and a hexahistidine tag, the modified protein was secreted into Luria­Bertani culture medium and purified by nickel-nitrilotriacetic acid affinity chromatography following a gel filtration column. In a 200 ml flask, the yield of boronophenylalanine­modified hirudin was 10 mg l­1 and that of rHirudin was 19 mg l­1. The authenticity of the purified proteins was verified using matrix-assisted laser desorption ionization time of flight mass spectroscopy and antithrombin activity assays. The results revealed that the antithrombin activity of the boronophenylalanine­modified hirudin to human thrombin was more enhanced than that of rHirudin. The modified hirudin demonstrated stronger proliferation inhibiting ability on fibroblast L929 cells compared with that of rHirudin.

Association study of TAC3 and TACR3 gene polymorphisms with idiopathic precocious puberty in Chinese girls.

BACKGROUND AND AIMS: The allele variant of neurokinin B and its receptor genes were thought important in regulating the human reproductive axis in many populations. The aim of this study was to investigate whether the variances in TAC3 and TACR3 genes that encoded neurokinin B (NKB) and its receptor (NK3R), individually, were associated with idiopathic precocious puberty in Chinese girls. METHODS: The genotyping method of polymerase chain reaction-restriction fragment length polymorphism was applied in this study; the distribution of five active single nucleon acid polymorphisms (SNPs) in TAC3 (p.G34R, p. A63P, p.Q66*, p.S99P, and a mutation on 5' UTR) and four sites in TACR3 (A29V or G, G59E, S455G, and A449S or T) genes was analyzed in 267 healthy and 186 idiopathic precocious puberty Chinese girls. RESULTS: Among the nine active SNPs, only A63P in TAC3 gene showed statistical differences; the p value was 0.024. CONCLUSION: A63P in TAC3 gene was statistically associated with the puberty onset time in Chinese girls.

Ascorbic acid-2-o-phosphate-6-o-palmitate protecting the human umbilical cord vein endothelial cells against hydrogen peroxide and tert-butyl hydroperoxide induced cytotoxicity.

Reactive oxygen species are believed to play a role in the development of several diseases including vascular diseases and the aging process. It is reported that increased reactive oxygen species were implicated as an important mechanism that contributes to endothelial dysfunction. So, human umbilical cord vein endothelial cells were used to study the antioxidative effect of L-ascorbic acid and its derivative. The study indicated that L-ascorbic acid as a traditional antioxidant was instable and could protect the cells against hydrogen peroxide induced cytotoxicity as its concentration below 50 microg/ml, but hardly could protect the cells against tert-butyl hydroperoxide induced cytotoxicity. Ascorbic acid-2-o-phosphate-6-o-palmitate could effectively protect the cells against hydrogen peroxide and tert-butyl hydroperoxide induced cytotoxicity, and exhibited no cytotoxicity within the tested concentration range. The study indicated that ascorbic acid-2-o-phosphate-6-o-palmitate could not only significantly reduce the intracellular reactive oxygen species level in 3 h culture, but also increase the cell viability in 15 h culture. In addition, ascorbic acid-2-o-phosphate-6-o-palmitate could keep stable in RPMI-1640 medium and water for 4 days, permeate the cell membrane, which in turn may scavenge the intracellular reactive oxygen species, increase the cell viability and the plasminogen activators'activity. All the above results suggested that addition of some hydrophobic groups to the traditional antioxidants could form novel compounds with better properties.

Regulation of the HMOX1 gene by the transcription factor AP-2δ with unique DNA binding site.

AP-2 transcription factors are important sequence-specific DNA-binding regulators that are expressed in the neural crest and other tissues during mammalian development. The human AP-2 family of transcription factors consists of five members, AP-2α, -ß, -γ, -δ and -ε, which have an important role in the regulation of gene expression during development and in the differentiation of multiple organs and tissues. The present study aimed to investigate the mechanism by which AP-2δ mediates heme oxygenase-1 (HMOX1) gene expression. It was identified that the human AP-2δ protein exhibited weak binding to a suboptimal AP-2 sequence, 5'-GCCN3GGC-3', to which all other AP-2 proteins bind in vitro, providing the first example of DNA target specificity amongst the AP-2 family. AP-2δ protein bound to an optimized AP-2 consensus DNA sequence, 5'-GCCTGAGGC-3', in vitro and transactivated gene expression in eukaryotic cells. The transactivation domain of Ap-2δ differs notably from those in the other AP-2 proteins as it lacks the PY motif (XPPXY) and several other conserved residues that are important for the transcriptional activity of AP-2 proteins, yet it functions as an equally strong activator.