Research on the application of Thelephora ganbajun exopolysaccharides in antioxidant, anti-inflammatory and spot-fading cosmetics.

Thelephora ganbajun exopolysaccharides (TGEP) with a "coral-like" branched chain structure (main chain diameter âˆ¼ 80 nm) were prepared by liquid fermentation and fractionated by ion-exchange chromatography. The main fraction (TGEP-2) with the highest in vitro antioxidant capacity was composed of Glc, Man, Gal, GalA, GlcA, Ara, Rha, GlcN, Fuc and Rib in a molar ratio of 465.43:420.43:219.14:188.43:37:35.14:31.43:19.43:11.14:1, with a molecular weight of 1.879 × 104 Da. The sequence of monosaccharide residue release revealed that Gal, Glc and Ara residues were more distributed in the side-branch chains and at their ends, whereas Man and GalA residues were more distributed in the main chains. TGEP-2 contained linear residues (mainly →4)-Glcp-(1 â†’ and →4)-Manp-(1→), branch residues (→3,6)-Glcp-(1→, →4,6)-Glcp-(1 â†’ and →3,6)-Galp-(1→) and terminal residues (Galp-(1→, Manp-(1 â†’ and Glcp-(1→). TGEP-2 consisted of α- and ß-glycosidically linked pyranosides, with a triple helical conformation and many long branches. Zebrafish oxidative stress and inflammation models found that TGEP-2 had antioxidant and anti-inflammatory activities. The zebrafish skin black spot assay showed that TGEP-2 inhibited melanin formation. Therefore, extracellular polysaccharides of T. ganbajun have strong application potential in anti-oxidant, anti-inflammatory and skin spot-fading functions cosmetics.

LEF1 enhances ß-catenin transactivation through IDR-dependent liquid-liquid phase separation.

Wnt/ß-catenin signaling plays a crucial role in cancer development, primarily activated by ß-catenin forming a transcription complex with LEF/TCF in the nucleus and initiating the transcription of Wnt target genes. Here, we report that LEF1, a member of the LEF/TCF family, can form intrinsically disordered region (IDR)-dependent condensates with ß-catenin both in vivo and in vitro, which is required for ß-catenin-dependent transcription. Notably, LEF1 with disrupted IDR lost its promoting activity on tumor proliferation and metastasis, which can be restored by substituting with FUS IDR. Our findings provide new insight into the essential role of liquid-liquid phase separation in Wnt/ß-catenin signaling and present a potential new target for cancer therapy.

Preparation and structure-activity relationship of highly active black garlic polysaccharides.

The aim of this study was to establish a method to improve the biological activity of polysaccharides. Three acid-treated polysaccharides (BGPS-2, BGPS-3 and BGPS-4) were obtained by treating black garlic polysaccharides (BGPS-1) with sulfuric acid at different intensities. The structure was characterized using the sulfuric acid-carbazole assay, IC, HPSEC-MALLS and FT-IR. The biological functions were evaluated using antioxidant and melanin biosynthesis inhibition assays. Compared with BGPS-1, the molecular weight of acid-treated polysaccharides significantly decreased, and the uronic acid content significantly increased. Antioxidant capacity negatively correlated with molecular weight, whereas melanin inhibition activity positively correlated with uronic acid content. BGPS-4 had the highest antioxidant capacity and the lowest molecular weight (1.25 × 103 Da), 79.41 % lower than that of BGPS-1. BGPS-3 was the strongest inhibitor of melanin formation and had the highest uronic acid content (50.73 %), 238.2 % higher than that of BGPS-1. Molecular weight and uronic acid content were the main structural characteristics that affected the antioxidant and melanin biosynthesis inhibition activities, respectively. BGPS-1, BGPS-2, BGPS-3, and BGPS-4 all had ß-linked pyranose, multi-branched, and non-triple helical spiral structures. Therefore, the acid hydrolysis method markedly modified the structural characteristics of black garlic polysaccharides, and increased their antioxidant capacity and melanin biosynthesis inhibition activity.

MBD3 promotes hepatocellular carcinoma progression and metastasis through negative regulation of tumour suppressor TFPI2.

BACKGROUND: The mechanism of recurrence and metastasis of hepatocellular carcinoma (HCC) is complex and challenging. Methyl-CpG binding domain protein 3 (MBD3) is a key epigenetic regulator involved in the progression and metastasis of several cancers, but its role in HCC remains unknown. METHODS: MBD3 expression in HCC was detected by immunohistochemistry and its association with clinicopathological features and patient's survival was analysed. The effects of MBD3 on hepatoma cells growth and metastasis were investigated, and the mechanism was explored. RESULTS: MBD3 is significantly highly expressed in HCC, associated with the advanced tumour stage and poor prognosis in HCC patients. MBD3 promotes the growth, angiogenesis and metastasis of HCC cells by inhibiting the tumour suppressor tissue factor pathway inhibitor 2 (TFPI2). Mechanistically, MBD3 can inhibit the TFPI2 transcription via the Nucleosome Remodeling and Deacetylase (NuRD) complex-mediated deacetylation, thus reactivating the activity of matrix metalloproteinases (MMPs) and PI3K/AKT signaling pathway, leading to the progression and metastasis of HCC CONCLUSIONS: Our results unravel the novel regulatory function of MBD3 in the progression and metastasis of HCC and identify MBD3 as an independent unfavourable prognostic factor for HCC patients, suggesting its potential as a promising therapeutic target as well.

Corn Steep Liquor: Green Biological Resources for Bioindustry.

Corn steep liquor (CSL) is a by-product of the wet milling process and contains mostly crude proteins, amino acids, minerals, vitamins, reducing sugars, organic acids, enzymes and other nutrients. The concentration of organic matter in the CSL is high and the yield is large. If directly discharged into the integrated wastewater treatment system, the load and cost of wastewater treatment will be greatly increased. On the other hand, most of the organic matter in the CSL is a valuable resource that can be reused and recovered, and has a significant resource potential. How to develop and utilize CSL has become a major problem faced by enterprises and society. In recent years, people have done a lot of research on the comprehensive utilization of CSL. CSL is commonly used as an inexpensive source of nitrogen, carbon or vitamins in the production of glutamate, antibiotics, lactic acid and other biotechnologies. This article reviews the active ingredients of CSL and their analytical methods, as well as its use for microbial culture medium, low-cost animal feed, biosurfactant, and biostimulant.

Whole genome sequencing identifies a deletion mutation in the unknown-functional KCNG2 from familial sick sinus syndrome.

Sick sinus syndrome (SSS) is a term used for a variety of disorders defined by abnormal cardiac impulse formation and by abnormal propagation from the heart's sinoatrial node. In this study, we present a case from a Chinese family in which two closely related individuals had the symptoms and electrocardiographic evidence of SSS. We hypothesized that multiple individuals affected by the disease in the family was an indication of its genetic predisposition, and thus performed high-throughput sequencing for the participants from the family to detect potential disease-associated variants. One of the potential variants that was identified was a KCNG2 gene variant (NC_000018.9: g.77624068_77624079del). Further bioinformatic analysis showed that the observed variant may be a pathogenic mutation. The results of protein-protein docking and whole cell patch-clamp measurements implied that the deletion variant in KCNG2 could affect its binding the KV2.1 protein, and finally affect the function of Kv channel, which is an important determinant in regulation of heartbeat. Therefore, we inferred that the variable KCNG2 gene may affect the function of Kv channel by changing the binding conformation of KCNG2 and KV2.1 proteins and then adversely affect propagation from the sinoatrial node and cardiac impulse formation by changing the action potential repolarization of heart cells. In summary, our findings suggested that the dominant KCNG2 deletion variant in the examined Chinese family with SSS may be a potential disease-associated variant.

A glucose biosensor based on glucose oxidase fused to a carbohydrate binding module family 2 tag that specifically binds to the cellulose-modified electrode.

The method of immobilization of glucose oxidase (GOD) on electrodes is especially important for the fabrication and performance of glucose biosensors. In this study, a carbohydrate binding module family 2 (CBM2) was successfully fused to the C terminal of GOD with a natural linker (NL) in endo-ß-xylanase by genetic recombination, and a fusion GOD (GOD-NL-CBM2) was obtained. The CBM2 was used as an affinity adsorption tag for immobilization of the GOD-NL-CBM2 on a cellulose modified electrode. The specific activity of GOD-NL-CBM2 was comparable to that of the wild type GOD. In addition, the CBM2 tag of fusion GOD almost maintained its highest binding capacity under optimal catalytic conditions (pH 5.0, 50 °C). The morphology and composition analysis of the cellulose film reacted with and without GOD or GOD-NL-CBM2 confirmed the immobilization of GOD-NL-CBM2. The electrochemical properties of the GOD-NL-CBM2/cellulose film bioelectrode, with a characteristic peak of H2O2 at +0.6 V in the presence of glucose, revealed the capability of the immobilized GOD-NL-CBM2 to efficiently catalyze glucose and produce H2O2. Additionally, the current signal response of the biosensor to glucose was linear in the concentration range from 1.25 to 40 mM (r2 ≥ 0.99). The sensitivity and detection limit of the GOD-NL-CBM2/cellulose film bioelectrode were 466.7 µA mol-1 L cm-2 and 0.475 mM (S/N = 3), respectively. Moreover, the glucose biosensor exhibited a rapid current change (< 5 s), high reproducibility (Relative standard deviation, RSD < 5%), substrate selectivity and stability, and retained about 80 % of the original current response after 2 months. The affinity adsorption-based immobilization strategy for GOD provides a promising approach to develop a high performance glucose biosensor.

Orientated Immobilization of FAD-Dependent Glucose Dehydrogenase on Electrode by Carbohydrate-Binding Module Fusion for Efficient Glucose Assay.

The discovery or engineering of fungus-derived FAD-dependent glucose 1-dehydrogenase (FAD-GDH) is especially important in the fabrication and performance of glucose biosensors. In this study, a novel FAD-GDH gene, phylogenetically distantly with other FAD-GDHs from Aspergillus species, was identified. Additionally, the wild-type GDH enzyme, and its fusion enzyme (GDH-NL-CBM2) with a carbohydrate binding module family 2 (CBM2) tag attached by a natural linker (NL), were successfully heterogeneously expressed. In addition, while the GDH was randomly immobilized on the electrode by conventional methods, the GDH-NL-CBM2 was orientationally immobilized on the nanocellulose-modified electrode by the CBM2 affinity adsorption tag through a simple one-step approach. A comparison of the performance of the two electrodes demonstrated that both electrodes responded linearly to glucose in the range of 0.12 to 40.7 mM with a coefficient of determination R2 > 0.999, but the sensitivity of immobilized GDH-NL-CBM2 (2.1362 × 10-2 A/(M*cm2)) was about 1-fold higher than that of GDH (1.2067 × 10-2 A/(M*cm2)). Moreover, a lower detection limit (51 µM), better reproducibility (<5%) and stability, and shorter response time (≈18 s) and activation time were observed for the GDH-NL-CBM2-modified electrode. This facile and easy immobilization approach used in the preparation of a GDH biosensor may open up new avenues in the development of high-performance amperometric biosensors.

Increased antioxidant activity and improved structural characterization of sulfuric acid-treated stepwise degraded polysaccharides from Pholiota nameko PN-01.

The aim of this study was to investigate sulfuric acid degradation of the Pholiota nameko polysaccharide (AIPS-1). Three stepwise degraded polysaccharides (AIPS-2, AIPS-3, and AIPS-4) were obtained by sequentially increasing the strength of sulfuric acid treatment. Structural characterization showed that sulfuric acid treatment significantly decreased molecular weight, increased the content of uronic acid and changed the molar ratio of monosaccharide composition, while the major functional groups and the triple helical conformation of polysaccharides did not change significantly. In vitro experiments proved that the antioxidation ability of the stepwise degraded polysaccharides gradually increased (AIPS-1 < AIPS-2 < AIPS-3 < AIPS-4). An oxidative stress zebrafish model was established, which demonstrated that the ability of AIPS-3 and AIPS-4 to scavenge free radicals in zebrafish was significantly improved compared to AIPS-1. In conclusion, sulfuric acid treatment is an effective method for improving the antioxidant activity of polysaccharides, and increased antioxidant activity was closely related to the changes in their structural characteristics.

Distribution of Zinc in Mycelial Cells and Antioxidant and Anti-Inflammatory Activities of Mycelia Zinc Polysaccharides from Thelephora ganbajun TG-01.

This study demonstrates that Thelephora ganbajun had a strong ability to absorb zinc, and zinc can be compartmentally stored in the small vesicles and mainly accumulated in the form of zinc-enriched polysaccharides (zinc content was 25.0 ± 1.27 mg/g). Mycelia zinc polysaccharides (MZPS) and its fractions were isolated. The main fraction (MZPS-2) with the highest antioxidant activity in vitro was composed of mannose : galacturonic acid : glucose : galactose in a molar ratio of 61.19 : 1 : 39.67 : 48.67, with a weight-averaged molecular weight of 5.118 × 105 Da. MZPS-2 had both α-pyranose and ß-pyranose configuration and had a triple helical conformation. By establishing zebrafish models, we found that MZPS-2 can significantly scavenge free radicals, reduce the generation of reactive oxygen species caused by inflammation, and inhibit the recruitment of neutrophils toward the injury site. Therefore, MZPS-2 exhibited antioxidant and anti-inflammatory effects and can be used as a zinc supplement with specific biological activities to alleviate zinc deficiency complications, such as chronic oxidative stress or inflammation.

Integrated Functional-Omics Analysis of Thermomyces lanuginosus Reveals its Potential for Simultaneous Production of Xylanase and Substituted Xylooligosaccharides.

Thermophiles have several beneficial properties for the conversion of biomass at high temperatures. Thermomyces lanuginosus is a thermophilic filamentous fungus that was shown to secrete 40 glycoside hydrolases and 25 proteases when grown on different carbon sources. Among the 13 identified glycoside hydrolases with high expression levels, 9 were reduced sugar glycosidases (RSGs) belonging to seven GH families, and 7 of the 10 identified proteases were exopeptidases belonging to six different protease families. High expression of RSGs and exopeptidases may allow the fungus to efficiently degrade oligosaccharides and oligopeptides in saprophytic habitats. There were no xylan side chain-degrading enzymes predicted in the genome of T. lanuginosus, and only one thermophilic GH11 xylanase (g4601.t1) and one GH43 xylosidase (g3706.t1) were detected by liquid chromatography-mass spectrometry/mass spectrometry when T. lanuginosus grown on xylan, which led to the accumulation of substituted xylooligosaccharides (SXOS) during corncob xylan degradation where SXOS output made up more than 8% of the total xylan. The SXOS are beneficial prebiotics and important inducers for enzymes secretion of microorganisms. Thus, T. lanuginosus exhibits distinct advantages in utilizing cheap raw materials producing one thermostable xylanase and the high value-added SXOS as well as microbial inoculants to compost by batch fermentation.

A Highly Efficient Xylan-Utilization System in Aspergillus niger An76: A Functional-Proteomics Study.

Xylan constituted with ß-1,4-D-xylose linked backbone and diverse substituted side-chains is the most abundant hemicellulose component of biomass, which can be completely and rapidly degraded into fermentable sugars by Aspergillus niger. This is of great value for obtaining renewable biofuels and biochemicals. To clarify the underlying mechanisms associated with highly efficient xylan degradation, assimilation, and metabolism by A. niger, we utilized functional proteomics to analyze the secreted proteins, sugar transporters, and intracellular proteins of A. niger An76 grown on xylan-based substrates. Results demonstrated that the complete xylanolytic enzyme system required for xylan degradation and composed of diverse isozymes was secreted in a sequential order. Xylan-backbone-degrading enzymes were preferentially induced by xylose or other soluble sugars, which efficiently produced large amounts of xylooligosaccharides (XOS) and xylose; however, XOS was more efficient than xylose in triggering the expression of the key transcription activator XlnR, resulting in higher xylanase activity and shortening xylanase-production time. Moreover, the substituted XOS was responsible for improving the abundance of side-chain-degrading enzymes, specific transporters, and key reductases and dehydrogenases in the pentose catabolic pathway. Our findings indicated that industries might be able to improve the species and concentrations of xylan-degrading enzymes and shorten fermentation time by adding abundant intermediate products of natural xylan (XOS) to cultures of filamentous fungi.

Extracellular Enzyme Composition and Functional Characteristics of Aspergillus niger An-76 Induced by Food Processing Byproducts and Based on Integrated Functional Omics.

Byproducts of food processing can be utilized for the production of high-value-added enzyme cocktails. In this study, we utilized integrated functional omics technology to analyze composition and functional characteristics of extracellular enzymes produced by Aspergillus niger grown on food processing byproducts. The results showed that oligosaccharides constituted by arabinose, xylose, and glucose in wheat bran were able to efficiently induce the production of extracellular enzymes of A. niger. Compared with other substrates, wheat bran was more effective at inducing the secretion of ß-glucosidases from GH1 and GH3 families, as well as >50% of proteases from A1-family aspartic proteases. Compared with proteins induced by single wheat bran or soybean dregs, the protein yield induced by their mixture was doubled, and the time required to reach peak enzyme activity was shortened by 25%. This study provided a technical platform for the complex formulation of various substrates and functional analysis of extracellular enzymes.

Determination of the modes of action and synergies of xylanases by analysis of xylooligosaccharide profiles over time using fluorescence-assisted carbohydrate electrophoresis.

The structure of xylan, which has a 1,4-linked ß-xylose backbone with various substituents, is much more heterogeneous and complex than that of cellulose. Because of this, complete degradation of xylan needs a large number of enzymes that includes GH10, GH11, and GH3 family xylanases together with auxiliary enzymes. Fluorescence-assisted carbohydrate electrophoresis (FACE) is able to accurately differentiate unsubstituted and substituted xylooligosaccharides (XOS) in the heterogeneous products generated by different xylanases and allows changes in concentrations of specific XOS to be analyzed quantitatively. Based on a quantitative analysis of XOS profiles over time using FACE, we have demonstrated that GH10 and GH11 family xylanases immediately degrade xylan into sizeable XOS, which are converted into smaller XOS in a much lower speed. The shortest substituted XOS produced by hydrolysis of the substituted xylan backbone by GH10 and GH11 family xylanases were MeGlcA(2) Xyl3 and MeGlcA(2) Xyl4 , respectively. The unsubstituted xylan backbone was degraded into xylose, xylobiose, and xylotriose by both GH10 and GH11 family xylanases; the product profiles are not family-specific but, instead, depend on different subsite binding affinities in the active sites of individual enzymes. Synergystic action between xylanases and ß-xylosidase degraded MeGlcA(2) Xyl4 into xylose and MeGlcA(2) Xyl3 but further degradation of MeGlcA(2) Xyl3 required additional enzymes. Synergy between xylanases and ß-xylosidase was also found to significantly accelerate the conversion of XOS into xylose.

Draft Genome Sequence of Aspergillus niger Strain An76.

The filamentous fungus Aspergillus niger has become one of the most important fungi in industrial biotechnology, and it can efficiently secrete both polysaccharide-degrading enzymes and organic acids. We report here the 6,074,961,332-bp draft sequence of A. niger strain An76, and the findings provide important information related to its lignocellulose-degrading ability.

Comparative Secretome Analysis of Aspergillus niger, Trichoderma reesei, and Penicillium oxalicum During Solid-State Fermentation.

Filamentous fungi such as Aspergillus spp., Trichoderma spp., and Penicillium spp. are frequently used to produce high concentrations of lignocellulosic enzymes. This study examined the discrepancies in the compositions and dynamic changes in the extracellular enzyme systems secreted by Aspergillus niger ATCC1015, Trichoderma reesei QM9414, and Penicillium oxalicum 114-2 cultured on corn stover and wheat bran. The results revealed different types and an abundance of monosaccharides and oligosaccharides were released during incubation, which induced the secretion of diverse glycoside hydrolases. Both the enzyme activities and isozyme numbers of the three fungal strains increased with time. A total of 279, 161, and 183 secretory proteins were detected in A. niger, T. reesei, and P. oxalicum secretomes, respectively. In the A. niger secretomes, more enzymes involved in the degradation of (galacto)mannan, xyloglucan, and the backbone of pectin distributed mostly in dicots were detected. In comparison, although P. oxalicum 114-2 hardly secreted any xyloglucanases, the diversities of enzymes involved in the degradation of xylan and ß-(1,3;1,4)-D-glucan commonly found in monocots were higher. The cellulase system of P. oxalicum 114-2 was more balanced. The degradation preference provided a new perspective regarding the recomposition of lignocellulosic enzymes based on substrate types.

[Response rules of pressing pain on back section in Governor Vessel in patients with gastroesophageal reflux disease].

The response rules of pressing pain on the back section in the Governor Vessel in patients with gastro-esophageal reflux disease (GERD) were studied to provide references for the diagnosis and treatment of GERD. Seventy-six cases of GERD were included into an observation group while 30 healthy volunteers were recruited into a control group. A mechanical measurement device of pressing pain that could measure the pain threshold was adapted to observe the pressing pain on the back section in the Governor Vessel in GERD patients and healthy volunteers. The test area is from spinous process of the 1st thoracic vertebra to that of the 12th thoracic vertebra (T1 -T12), including acupoints and non-acupoints on the Governor Vessel. As a result, in the observation group the pain threshold of T5-T7 spinous process clearance, which was the location of Shendao (GV 11), Lingtai (GV 10) and Zhiyang (GV 9), was lower than that in the control group (all P < 0.05). This result indicated that there was significant pressing pain in T5-T7 spinous process clearance in patients with GERD, which could be taken as an important auxiliary diagnosis and a new thinking method in the treatment of GERD with acupuncture.

NEDD4 ubiquitinates TRAF3 to promote CD40-mediated AKT activation.

CD40, a member of tumour necrosis factor receptor (TNFR) superfamily, has a pivotal role in B-cell-mediated immunity through various effector pathways including AKT kinase, but the signal transduction of CD40-meidated AKT activation is poorly understood. Here we report that the neural precursor cell expressed developmentally downregulated protein 4 (NEDD4), homologous to E6-AP Carboxyl Terminus family E3 ubiquitin ligase, is a novel component of the CD40 signalling complex. It has a key role in CD40-mediated AKT activation and is involved in modulating immunoglobulin class switch through regulating the expression of activation-induced cytidine deaminase. NEDD4 constitutively interacts with CD40 and mediates K63-linked ubiquitination of TNFR-associated factor3 (TRAF3). The ubiquitination of TRAF3 by NEDD4 is critical for CD40-mediated AKT activation. Thus, NEDD4 is a previously unknown component of the CD40 signalling complex necessary for AKT activation.

Dysregulation of the miR-324-5p-CUEDC2 axis leads to macrophage dysfunction and is associated with colon cancer.

CUEDC2, a CUE-domain-containing protein, modulates inflammation, but its involvement in tumorigenesis is still poorly understood. Here, we report that CUEDC2 is a key regulator of macrophage function and critical for protection against colitis-associated tumorigenesis. CUEDC2 expression is dramatically upregulated during macrophage differentiation, and CUEDC2 deficiency results in excessive production of proinflammatory cytokines. The level of CUEDC2 in macrophages is modulated by miR- 324-5p. We find that Cuedc2 KO mice are more susceptible to dextran-sodium-sulfate-induced colitis, and macrophage transplantation results suggest that the increased susceptibility results from the dysfunction of macrophages lacking CUEDC2. Furthermore, we find that Cuedc2 KO mice are more prone to colitis-associated cancer. Importantly, CUEDC2 expression is almost undetectable in macrophages in human colon cancer, and this decreased CUEDC2 expression is associated with high levels of interleukin-4 and miR-324-5p. Thus, CUEDC2 plays a crucial role in modulating macrophage function and is associated with both colitis and colon tumorigenesis.

[Effects of knocking down Gankyrin on breast cancer metastasis in mice].

OBJECTIVE: To establish Gankyrin knocking down 4T1-luc cell model and detect the effects of Gankyrin expression on breast cancer metastasis. METHODS: 4T1-luc cells carrying shGankyrin construct were established by lentivirus infection and antibiotic screening. Western blotting and real-time PCR were used to check the expression levels of Gankyrin. In vivo imaging system was used to monitor the effects of Gankyrin knocked down on cell growth and tumor metastasis after the in situ implantation of Gankyrin knocking down 4T1-luc cells in BALB/c mice. RESULTS: The cell expression decreased at the protein and mRNA levels. Gankyrin mRNA expression in different shGankyrin 4T1-luc cells was respectively 4.9%, 25.1% and 69.8% versus the control cells. ShGankyrin#2 4T1-luc cells were chosen for in situ implantation into BAL/c mice because luminescent intensity was consistent with cell numbers. The photon flux of lung metastatic tumor induced by Gankyrin knocking down 4T1-luc cell was 3.02 × 10(6), while that of lung metastasis induced by control cells was 10.9 × 10(6). The differences between two groups were significant. In pathology, Gankyrin was detected positive in lung metastasis tumors induced by control group. However, Gankyrin was negative in the Gankyrin knockdown group. CONCLUSIONS: Lentivirus infection may be effectively used to establish Gankyrin knocking down 4T1-luc cell model. Because of its involvement in the in vivo pulmonary metastasis of breast cancers, Gankyrin should be a novel target for tumor therapy.