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ANKRD11 gene encodes for the large nuclear protein essential for multiple system development including the nervous system. However, the molecular basis for the proper nuclear localization of ANKRD11 has not yet been elucidated. In this study, we have identified a functional bipartite nuclear localization signal (bNLS) between residues 53 and 87 of ANKRD11. Using biochemical approaches, we discovered two major binding sites in this bipartite NLS for Importin α1. Through site-directed mutagenesis and functional analysis, we further found that this bipartite NLS is sufficient for nuclear import of overexpressing GFP in HeLa cells and necessary for nuclear localization of ANKRD11. Importantly, our study provides a possible pathogenic mechanism for certain clinical variants located within the bipartite nuclear localization signal of ANKRD11. • ANKRD11 N-terminus binds to Importin α1 via a bipartite region. • The region is a functional bipartite NLS in ANKRD11 with a long linker. • Disruption of the interaction impairs the nuclear localization of ANKRD11. [ABSTRACT FROM AUTHOR]

Pressure overload induced cardiac remodeling is associated with a complex spectrum of pathophysiological mechanisms. As inflammatory cells, macrophages maintain a critical position in mechanical stress-induced myocardial remodeling. HMGB1 is a highly conserved, ubiquitous protein in various types of cells whose biological roles are closely dependent on subcellular sites. However, whether HMGB1 expressed in macrophages performs the protective or pathological responses in cardiac remodeling is unknown. In this study, we generated the myeloid-specific HMGB1 knockout mice and detected the effects of macrophage HMGB1 in response to pathophysiological stress. Our data showed HMGB1 in macrophages played a protective role against the pressure overload induced cardiac pathophysiology. The deletion of HMGB1 in macrophages gains more differentiation of M1-type pro-inflammatory macrophage during the mechanical stress-induced myocardial remodeling, thereby aggravating the inflammatory response in whole heart, resulting in accelerated deterioration of cardiac function. Moreover, in vitro data also validated HMGB1 got involved in the process of macrophage polarization. Macrophages without HMGB1 are more inclined to differentiate into M1 during the stretch process. In summary, the present results indicated that loss of HMGB1 in macrophages can exacerbate heart failure through increased differentiation of pro-inflammatory macrophages and enhanced inflammatory response. • HMGB1 in macrophage nucleus played a protective role against the pressure overload induced cardiac pathophysiology. • Deleting HMGB1 in macrophages gains more differentiation of M1 macrophage and expression of inflammatory genes after TAC. • Macrophages without nuclear HMGB1 are more inclined to differentiate into M1 macrophage during the in vitro stretch process. [ABSTRACT FROM AUTHOR]

Long noncoding RNAs (lncRNAs) exert crucial roles in hepatocellular carcinoma (HCC) progression. LncRNA EIF3J-AS1 is recently reported to be highly expressed in HCC and correlates with recurrence-free survival. However, the biological function of EIF3J-AS1 and its underlying molecular mechanism are unexplored yet. In the current study, we demonstrated that EIF3J-AS1 expression was obviously upregulated in HCC tissues compared to adjacent noncancerous tissues. Moreover, the elevated levels of EIF3J-AS1 was detected in four HCC cell lines (HepG2, SMMC-7721, MHCC97H, HCCLM3) compared with the normal hepatic cell line LO2. Notably, the expression of EIF3J-AS1 was correlated with prognostic features including tumor size, vascular invasion and tumor stage. TCGA-LIHC data indicated that the upregulated expression of EIF3J-AS1 predicted poor prognosis of HCC. EIF3J-AS1 knockdown remarkably suppressed the proliferation, migration and invasion of HCC cells. Mechanistically, EIF3J-AS1 inversely regulated miR-122–5p expression via acting as a competing endogenous RNA (ceRNA) in HCC cells. Furthermore, catenin delta 2 (CTNND2) was recognized as a novel target of miR-122–5p. CTNND2 restoration partially reversed EIF3J-AS1 knockdown-induced inhibitory effects on HCC cell proliferation, migration and invasion. Importantly, we found that hypoxia induced EIF3J-AS1 and CTNND2 expression, and led to miR-122–5p downregulation in HCC cells. EIF3J-AS1 knockdown partially abolished hypoxia-induced HCC cell proliferation and mobility. In conclusion, our results provide a new insight into the molecular pathogenesis of HCC, and EIF3J-AS1 may be a potential therapeutic target for HCC. • EIF3J-AS1 expression is obviously up-regulated in HCC. • The elevated expression of EIF3J-AS1 indicates poor prognosis of HCC. • EIF3J-AS1 knockdown inhibits HCC cell proliferation, migration and invasion. • EIF3J-AS1 promotes CTNND2 expression via sponging miR-122–5p. • Hypoxia-induced EIF3J-AS1 facilitates HCC progression via regulating CTNND2. [ABSTRACT FROM AUTHOR]

Post-MI heart failure is characterized by structural remodeling, in which intramyocardial fibrosis takes a important part. Poly(ADP-ribose) polymerase 1 (PARP-1) is a extensive nuclear enzyme and plays a critical role in various diseases. It was shown that PARP-1 inhibition could alleviate heart failure and dowregulate autophagy, but whether PARP-1 regulates autophagy and thus impacts the activities of CFs remain unknown. We transfected cultured cardiac fibroblasts (CFs) with small interfere RNA-PARP-1 (siPARP-1) to downregulate PARP-1 and analyzed the ability of proliferation, migration, differentiation, and autophagy levels of CFs under different treatments using CCK8 assays, transwell migration assays, immunofluorescence assays detecting expression of α-SMA, western blot assays detecting autophagy-related proteins respectively. Furthermore, rat models of myocardial infarction (MI) were induced by ligation of left anterior descending coronary artery and PARP-1 inhibitor, 4-aminobenzamide (4-AB), was injected intraperitoneally after MI, followed by echocardiography detection, masson assays, immunohistochemistry assays detecting expression of α-SMA and western blot assays detecting autophagy-related proteins to investigate whether PARP-1 inhibition could regulate autophagy, alleviate cardiac fibrosis and improve cardiac function in vivo. In cultured CFs, siPARP-1 repressed TGF-β1-induced proliferation, migration, and differentiation through regulating autophagic levels. The in vitro results was verified by the in vivo study, indicating that PARP-1 inhibition partially decreased autophagy, abrogated cardiac fibrosis and significantly improved cardiac function post-MI. In conclusion, this work demonstrated the vital connection of PARP-1 and autophagy in the activation of CFs, and provided solid evidence supporting PARP-1 inhibition as a feasible strategy for the treatment of post-MI heart failure. [ABSTRACT FROM AUTHOR]

Abstract Inflammatory bowel disease (IBD) is a chronic autoimmune disease, and its pathogenesis remains mostly unknown. MicroRNAs (miRs) has drawn much attention as a crucial regulator of autoimmune diseases. In this study, we demonstrated, for the first time, that miR-425 was significantly up-regulated in peripheral blood mononuclear cells (PBMC) and mucosa of patients with IBD. In note, T helper (Th) 17 cells were found to be the major source of miR-425 expression. Using gain-of-function approaches, we demonstrated that miR-425 could facilitate the differentiation of CD4+ T cells into Th17 lineage. In addition, forkhead box O1 (Foxo1) was identified as a novel target gene of miR-425, which was able to inhibit Th17 cell differentiation, and it was observed to be markedly decreased in PBMC and mucosa of patients with IBD. Notably, in vivo inhibition of miR-425 significantly alleviated the disease severity of TNBS-induced colitis in mice, with down-regulated levels of IL-17A. Our data reveal a novel mechanism in which the elevated miR-425 in IBD mediates pathogenic Th17 cell generation through down-regulation of Foxo1. In vivo blockade of miR-425 may serve as a novel therapeutic approach in the treatment of IBD. Highlights • MiR-425 expression is significantly increased in patients with IBD. • MiR-425 promotes pathogenic Th17 cell differentiation. • Foxo1 is a novel target of miR-425. • In vivo inhibition of miR-425 alleviates TNBS-induced colitis in mice. [ABSTRACT FROM AUTHOR]

SM22α is a kind of 22-kDa protein which is exclusively expressed in smooth muscle containing tissues of the vertebrates. Here we report molecular cloning of a novel zebrafish SM22α gene. The full length of zebrafish SM22α cDNA is 1296 bp and it encodes a polypeptide of 201 amino acids which shares 69.2%, 69.7%, 69.2%, 67.2%, and 61.2% overall identity with human, mouse, rat, chicken, and bovine SM22α, respectively. Characterization of zebrafish SM22α genomic sequence reveals that it spans 7.7 kb and contains five exons and four introns. The expression pattern of SM22α in zebrafish embryonic development is studied by whole-mount in situ hybridization. Strong expression is observed in vascular, gut, swim bladder, branchial arches, and fin epidermis. Furthermore, we carry out gene knock-down by antisense morpholino oligonucleotide, which results in disappearance of yolk extension, caudal fin aberrance, and deficiency of circulation system in zebrafish embryo. Cross-section of SM22α-deficient embryo suggests that SM22α may play roles in smooth muscle cell morphology transform. [Copyright &y& Elsevier]

Cancer stem cells (CSCs) play essential role in the progression of many tumors. Wnt/β-catenin pathway is crucial in maintaining the stemness of CSCs. (−)-Epigallocatechin-3-gallate (EGCG), the major bioactive component in green tea, has been shown to possess anti-cancer activity. To date, the interventional effect of EGCG on lung CSCs has not been elucidated yet. In the present study, tumorsphere formation assay was used to enrich lung CSCs from A549 and H1299 cells. We revealed that Wnt/β-catenin pathway was activated in lung CSCs, and downregulation of β-catenin, abolished lung CSCs traits. Our study further illustrated that EGCG effectively diminished lung CSCs activity by inhibiting tumorsphere formation, decreasing lung CSCs markers, suppressing proliferation and inducing apoptosis. Moreover, We showed that EGCG downregulated Wnt/β-catenin activation, while upregulation of Wnt/β-catenin dampened the inhibitory effects of EGCG on lung CSCs. Taken together, these results demonstrated the role of Wnt/β-catenin pathway in regulating lung CSCs traits and EGCG intervention of lung CSCs. Findings from this study could provide new insights into the molecular mechanisms of lung CSCs intervention. [ABSTRACT FROM AUTHOR]

KDM2A is a histone demethylase, which primarily catalyzes the demethylation of H3K36me2. Abnormal expression of KDM2A is observed in many types of cancers; however, the molecular events connected to KDM2A expression remain unclear. We report that KDM2A performs an oncogenic function in esophageal squamous cell carcinoma (ESCC) and is robustly expressed in ESCC cells. ShRNA-mediated knockdown of KDM2A resulted in a significant inhibition of the malignant phenotype of ESCC cell lines, whereas ectopic expression of KDM2A showed the opposite effect. We also analyzed the function of KDM2A using a CRISPR-CAS9 depletion system and subsequent rescue experiment, which also indicated a cancerous role of KDM2A. Interestingly, analysis of the gene expression network controlled by KDM2A using RNA-seq revealed an unexpected feature: KDM2A could induce expression of a set of well-documented oncogenic genes, including IL6 and LAT2, while simultaneously suppressing another set of oncogenes, including MAT2A and HMGCS1. Targeted inhibition of the upregulated oncogene in the KDM2A-depleted cells led to a synergistic suppressive effect on the malignant phenotype of ESCC cells. Our results revealed the dual role of KDM2A in ESCC cells, which may have therapeutic implications. • Ablation of KDM2A attenuates the malignant phenotype of ESCC cell lines. • Augmented expression of KDM2A promotes the malignant phenotype of ESCC cells. • KDM2A activates one set of oncogenic genes and represses another set of cancerous genes simultaneously. • Inhibition of activated oncogenes generates a synergistic antitumor effect in KDM2A-deficient ESCC cells. • The dual role of KDM2A may be used to develop a combination treatment for ESCC. [ABSTRACT FROM AUTHOR]

Abstract: δ-sarcoglycan, one member of the sarcoglycan complex, is a very conservative muscle-specific protein exclusively expressed in the skeletal and cardiac muscles of vertebrates. Mutations in sarcoglycans are known to be involved in limb-girdle muscular dystrophy (LGMD) and dilated cardiomyopathy (DCM) in humans. To address the role of δ-sarcoglycan gene in zebrafish development, we have studied expression pattern of δ-sarcoglycan in zebrafish embryos and examined the role of δ-sarcoglycan in zebrafish embryonic development by morpholino. Strong expression of δ-sarcoglycan was observed in various muscles including those of the segment, heart, eye, jaw, pectoral fin, branchial arches, and swim bladder in zebrafish embryo. δ-sarcoglycan was also expressed in midbrain and retina. Knockdown of δ-sarcoglycan resulted in severe abnormality in both the cardiac and skeletal muscles. Some severe ones displayed serious morphological abnormality such as hypoplastic head, linear heart, very weak heartbeats, and runtish trunk, all dead within 5dpf. Whole-mount in situ hybridization analysis showed that adaxial cells and muscle pioneers were affected in δ-sarcoglycan knockdown embryos. In addition, absence of δ-sarcoglycan protein severely delayed the cardiac development and influenced the differentiation of cardiac muscle, and the cardiac left-right asymmetry was dramatically changed in morpholino-treated embryos. These data together suggest that δ-sarcoglycan plays an important role in early heart and muscle development. [Copyright &y& Elsevier]

Human ATP-binding cassette transporter 8 of subfamily B (hABCB8) is an ABC transporter that located in the inner membrane of mitochondria. The ABCB8 is involved in the maturation of Fe–S and protects the heart from oxidative stress. Here, we present the cryo-EM structure of human ABCB8 binding with AMPPNP in inward-facing conformation with resolution of 4.1 Å. hABCB8 shows an open-inward conformation when ATP is bound. Unexpectedly, cholesterol molecules were identified in the transmembrane domain of hABCB8. Our results provide structural basis for the transport mechanism of the ABC transporter in mitochondria. • The human ABCB8 transporter was expressed using Bac-to-Bac system. • The cryo-EM structure of human ABCB8 binding with AMPPNP in inward-facing conformation with resolution of 4.1 Å. • The human ABCB8 shows an open-inward conformation when AMPPNP/ATP is bound. • Cholesterol molecules were identified in the transmembrane domain of human ABCB8. [ABSTRACT FROM AUTHOR]

Robust evidence suggested that gut-derived lipopolysaccharide (LPS) plays a significant role in various liver injury diseases; however, the role of gut-derived LPS in acetaminophen (APAP) overdose-induced acute liver injury remains unclear. The present study aimed to investigate the effect of gut-derived LPS on APAP-induced liver injury. Our results revealed that reduction of gut-derived LPS using multiple antibiotics could significantly exacerbate APAP-induced liver injury and increase mortality in mice. By contrast, pretreatment with exogenous LPS could reverse APAP-induced liver hepatotoxicity in mice and rats. We observed that TNF-α secretion in the liver was significantly increased after LPS pretreatment. In addition, depletion of TNF-α or TNFR1 inhibited the protective effects of LPS against APAP-induced hepatotoxicity, which indicated that the TNF-α/TNFR1 pathway was required to protect against APAP-induced liver injury. Mechanistically, LPS reduces oxidative stress by upregulating the expression of hepatic GSH, reducing MDA levels in liver tissues, and upregulating the expression of several antioxidant genes after APAP injection. However, the production of hepatic GSH was not enhanced in the liver tissues of rats lacking TNF-α or TNFR1 and MDA levels were not reduced after LPS and APAP co-treatment. The above results suggested LPS alleviated APAP-induced oxidative stress via the TNF-α/TNFR1 pathway. • Reduction of gut-derived LPS by antibiotics aggravated APAP-induced liver injury. • LPS pretreatment alleviated APAP-induced liver injury by suppressing oxidative stress. • LPS alleviated APAP-induced oxidative stress via the TNF-α/TNFR1 pathway. [ABSTRACT FROM AUTHOR]

Comprehensive knowledge on the murine CT26 colon carcinoma line is a classic model used in the pharmacodynamic experiments involving IDO-1 inhibitors, immune-related checkpoint antibodies and immune related mechanisms. In this study, we determined the impact of different subcutaneous inoculation locations on tumor growth and immune factor expression. CT26 cells were treated with the IDO-1 inhibitor, INCB024360, following INF-γ stimulation and analyzed for kynurenine concentration. Female Balb/c mice were inoculated with CT26 cells in either or both the right upper flank or the right lower flank. Isolated tumors were evaluated for changes in tumor volume following treatment with anti-PD-1, anti-CTLA-4, or no treatment. Isolated tumors were also evaluated for changes in immune cell subpopulations and expression of key immune factors using FACS. Treatment of two CT26 cell lines with INCB024360 produced similar results. IC 50 values were 222.5 and 276.0, and the peak inhibitory rates were 97.99% and 91.85% respectively. Analysis of tumor growth revealed that tumor volumes were larger (1925 mm3 vs. 767 mm3), and the anti-tumor effects of both anti-PD-1 and anti-CTLA-4 were different in mice inoculated in the right lower flank than in those inoculated in the upper flank. FACS analysis revealed that the CD8+T subpopulation in the right upper flank was higher than that in the lower flank (*P < 0.05). By contrast, the myeloid cell populations was lower in the right upper flank than it was in the right lower flank (*P < 0.05). The INF-γ populations in CD8+T (*P < 0.05) and regulatory T (Treg) cell subpopulation (*P < 0.05) were also more abundant in the right upper flank than in the right lower flank. In contrast to the uniform of the results from the in vitro experiment, the anti-CTLA-4 and anti-PD-1 antibodies had different efficacies depending on the location of the subcutaneous inoculation of CT26 in mice. The differences in the percentages of CD8+T, myeloid cells, INF-γ in CD8+T, and Treg subpopulations indicated that the tumor microenvironment was affected by inoculation position. Taken together, these results suggest that tumors isolated from same cell line with different inoculation positions are different enough to be considered different models. • CT26 cells were inoculated in Balb/c mice produced difference tumor growth curve. • Anti-CTLA-4 and anti-PD-1 antibodies treatment with the difference inoculated site produced significant difference in anti-tumor effect. • Tumor microenvironment (the expression level of immune cell subpopulations) depends on inoculated positions in mice. [ABSTRACT FROM AUTHOR]

Abstract Background/Aims The expression of transcription factor Zbtb1 is essential for the maintenance and development of various blood cells in the hematopoietic system. In the current study, we found that the total number of thymocytes in PLZF deficient mice was reduced compared with thymocytes in wild-type mice, and the number of early T-cell progenitors decreased. However, the decrease of thymocytes in PLZF deficient mice was not cell intrinsic. This study adds new information regarding the regulation of the PLZF gene in the development and self-renewal of T cells. Methods The thymus was isolated from newborn mice, and the two lobes of each thymus were physically separated. Each host received a thymus, two lobes , each placed at one end of the kidney, as described in the literature. Tail vein blood was periodically collected from some of the recipients and analyzed for the presence of peripheral blood T cells. Results In PLZF-EGFP reporter mice and neonatal thymus transplantation to the kidney, we found that PLZF was highly expressed in DN1 (Lineage−CD44+CD25−) cells of thymic grafts of Rag2/ γ c−/- recipient mice. We found that the proportion of PLZF wild-type and mutant-derived cells in the thymocytes of recipient mice after bone marrow transplantation is approximately equal to the competitive bone marrow chimeric mouse model, and all mice contain a normal thymus. Conclusion The development of T cells suggests that the effect of the PLZF gene on T cell differentiation and development is not cell intrinsic. However, in the neonatal mouse thymic transplant model in the Rag2/γc−/− recipient mouse, deletion of the PLZF gene results in a significant decrease in the proportion of DN1 cells from the donor in the thymic graft. Highlights • The number of total thymocytes and early T cell progenitor cells (ETP cells) is reduced in PLZF-deficient mice. • The decrease in thymocytes in PLZF-deficient mice is not intrinsic to cells. • PLZF is highly expressed in graft DN1 cells of the neonatal mouse thymic transplantation model. • PLZF deletion may affect donor DN1 cell self-renewal in neonatal mouse thymic transplantation model. [ABSTRACT FROM AUTHOR]

Abstract Inflammatory bowel disease (IBD) has been well-documented as a chronic gastrointestinal autoimmune disease, but its etiology remains to be elusive. Ascl2 (achaete-scute complex homologue 2), identified as a homologue of the Drosophila achaete-scute gene, has been shown to play an essential for the pathogenesis of autoimmune diseases and cancers. However, whether it is associated with the pathogenesis of IBD remains unclear. Here, we demonstrated that Ascl2 was greatly down-regulated in human IBD and experimental colitis. Interestingly, CD4+ T cell expression of Ascl2 was regulated by intestinal microbiota. Moreover, we revealed that Ascl2 inhibited the differentiation of Th17 cells and restrained their pathogenicity through facilitating IL-10 production. We further showed that Blimp-1 might be involved in the Ascl2-inducing IL-10 expression in CD4+ T cells under Th17 differentiating condition. Notably, lentivirus-mediated overexpression of Ascl2 remarkably alleviated the severity of 2,4,6-trinitrobenzenesulfonic acid solution (TNBS)-induced colitis in mice, with decreased level of colonic IL-17A. Our findings demonstrated an unappreciated mechanism whereby Ascl2 negatively modulates pathogenic Th17 cell differentiation via promoting IL-10 production, and alleviates intestinal inflammation. Thus, Ascl2 may serve as a novel therapeutic target of IBD. Highlights • Ascl2 expression is significantly down-regulated in patients with IBD. • Ascl2 expression is regulated by intestinal microbiota. • Ascl2 suppresses Th17 differentiation and facilitates IL-10 production. • In vivo over expression of Ascl2 alleviates experimental colitis in mice. [ABSTRACT FROM AUTHOR]

Abstract RNA helicases are almost ubiquitous important enzymes that take part in multiple aspects of RNA metabolism. Prokaryotes encode fewer RNA helicases than eukaryotes, suggesting that individual prokaryotic RNA helicases may take on multiple roles. The specific functions and molecular mechanisms of bacterial DEAH/RHA helicases are poorly understood, and no structures are available of these bacterial enzymes. Here, we report the first crystal structure of the DEAH/RHA helicase HrpB of Escherichia coli in a complex with ADP•AlF 4. It showed an atypical globular structure, consisting of two RecA domains, an HA2 domain and an OB domain, similar to eukaryotic DEAH/RHA helicases. Notably, it showed a unique C-terminal extension that has never been reported before. Activity assays indicated that EcHrpB binds RNA but not DNA, and does not exhibit unwinding activity in vitro. Thus, within cells, the EcHrpB may function in helicase activity-independent RNA metabolic processes. Highlights • Structure of Escherichia coli HrpB in a complex with ADP•AlF 4 is reported. • The CTE domain of EcHrpB is unique and harbors eight helices and nine β-sheets. • EcHrpB may function in helicase activity-independent RNA metabolic processes. [ABSTRACT FROM AUTHOR]

Mitochondrial calcium uniporter (MCU) is the pore-forming subunit of the entire uniporter complex and plays an important role in mitochondrial calcium uptake. However, the single channel recording of MCU remains controversial. Here, we expressed and purified different MCU proteins and then reconstituted them into planar lipid bilayers for single channel recording. We showed that MCU alone from Pyronema omphalodes (pMCU) is active with prominent single channel Ca 2+ currents. In sharp contrast, MCU alone from Homo sapiens (hMCU) is inactive. The essential MCU regulator (EMRE) activates hMCU, and therefore, the complex (hMCU-hEMRE) shows prominent single channel Ca 2+ currents. These single channel currents are sensitive to the specific MCU inhibitor Ruthenium Red. Our results clearly demonstrate that active MCU can conduct large amounts of calcium into the mitochondria. [ABSTRACT FROM AUTHOR]

Mitochondria Ca 2+ overload has long been recognized as a cell death trigger. Unexpectedly, we demonstrated a signaling complex composed of Calmodulin (CaM), Arabidopsis thaliana Bcl-2-associated athanogene 5 (AtBAG5) and Heat-shock cognate 70 protein (Hsc70) within Arabidopsis thaliana mitochondria which transduces mitochondria Ca 2+ elevations to suppress leaf senescence. Gain- and loss-of-function AtBAG5 mutant plants revealed that, mitochondria Ca 2+ elevation significantly increase chlorophyll retention and decrease H 2 O 2 level in dark-induced leaf senescence assay. Based on our findings, we proposed a molecular mechanism in which chronic mitochondria Ca 2+ elevation reduced ROS levels and thus inhibits leaf senescence. [ABSTRACT FROM AUTHOR]

The overexpression of PDZ-binding kinase/T-LAK cell-originated protein kinase (PBK/TOPK) has been associated with hematologic tumors, breast cancer and various other cancers. However, the three-dimensional structure of PBK has not been solved. In this study, we determined the crystal structure of human PBK, which has two phospho-mimicking mutations T9E and T198E. The structural data indicated that PBK may assemble into an inactive dimer in alkaline conditions. Analytical size-exclusion chromatography and analytical ultracentrifugation confirmed that PBK exists in a conformational transition between dimers and monomers at different pH conditions. Co-IP and kinase assays suggested that the active state of PBK is a monomer and does not form a dimer even under alkaline conditions. These results showed that the conformational transition of PBK is important for its kinase activity regulation. Collectively, our observations may provide a novel starting point for structure-based functional studies. [ABSTRACT FROM AUTHOR]

Abstract: It has been well recognized that human epidermal growth factor receptor 2 (HER2) level in breast cancer (BC) is closely related to the malignant biologic behaviors of the tumor, including invasion and metastasis. Yet, there has been a lack of directly observable evidence to support such notion. Here we report a quantum dots (QDs)-based double-color imaging technique to simultaneously show the HER2 level on BC cells and the type IV collagen in the tumor matrix. In benign breast tumor, the type IV collagen was intact. With the increasing of HER2 expression level, there has been a progressive decrease in type IV collagen around the cancer nest. At HER2 (3+) expression level, there has virtually been a total destruction of type IV collagen. Moreover, HER2 (3+) BC cells also show direct invasion into the blood vessels. This novel imaging method provides direct observable evidence to support the theory that the HER2 expression level is directly related to BC invasion. [Copyright &y& Elsevier]