Actin Capping Protein Research Articles

In vivo proximity proteomics of nascent synapses reveals a novel regulator of cytoskeleton-mediated synaptic maturation

Excitatory synapse formation during development involves the complex orchestration of both structural and functional alterations at the postsynapse. However, the molecular mechanisms that underlie excitatory synaptogenesis are only partially resolved, in part because the internal machinery of developing synapses is largely unknown. To address this, we apply a chemicogenetic approach, in vivo biotin identification (iBioID), to discover aspects of the proteome of nascent synapses. This approach uncovered sixty proteins, including a previously uncharacterized protein, CARMIL3, which interacts in vivo with the synaptic cytoskeletal regulator proteins SrGAP3 (or WRP) and actin capping protein. Using new CRISPR-based approaches, we validate that endogenous CARMIL3 is localized to developing synapses where it facilitates the recruitment of capping protein and is required for spine structural maturation and AMPAR recruitment associated with synapse unsilencing. Together these proteomic and functional studies reveal a previously unknown mechanism important for excitatory synapse development in the developing perinatal brain.

Cancer Stem-Cell Marker CD44v9-Positive Cells Arise From Helicobacter pylori-Infected CAPZA1-Overexpressing Cells.

Background & AimsCD44 variant 9 (CD44v9)-positive cancer stem-like cells strongly contribute to the development and recurrence of gastric cancer. However, the origin of CD44v9-positive cells is uncertain.MethodsCD44v9, β-catenin, and epithelial splicing regulatory protein 1 signals were assessed by real-time reverse-transcription polymerase chain reaction, immunoblot analysis, or immunofluorescence microscopy. Capping actin protein of muscle Z-line α subunit 1 (CAPZA1) expression was assessed by immunoblot analysis or immunohistochemical analysis of Mongolian gerbils' gastric mucosa or human biopsy specimens. Levels of oxidative stress were assessed by measuring malondialdehyde and protein carbonylation. Histone H3 acetylation levels in the CAPZA1 proximal promoter region were measured by using chromatin immunoprecipitation analysis with an antibody against the acetylated histone H3 in human gastric carcinoma cell line (AGS) cells.ResultsCD44v9 is expressed in CAPZA1-overexpressing cells in human gastric cancer tissues. CAPZA1 overexpression enhanced expression of β-catenin, which is a transcription factor for CD44, and epithelial splicing regulatory protein 1, which increases alternative splicing of CD44 to generate CD44v9. CAPZA1-overexpressing cells after cytotoxin-associated gene A accumulation showed CD44v9 expression by inducing nuclear accumulation of β-catenin, concomitant with the enhancement of expression of Sal-like protein 4 and Krüppel-like factor 5, which encode reprogramming factors. Oxidative stress increased the CAPZA1 expression in AGS cells through the enhancement of histone H3 acetylation of CAPZA1 promoter. CAPZA1 expression was increased depending on oxidative stress in H pylori–infected gastric mucosa.ConclusionsCD44v9 expression is evoked from CAPZA1-overexpressing cells after accumulation of cytotoxin-associated gene A. Our findings provide important insights into the mechanisms underlying the development of CD44v9-positive cells.

Gelsolin-like actin-capping protein has prognostic value and promotes tumorigenesis and epithelial-mesenchymal transition via the Hippo signaling pathway in human bladder cancer.

Background:Transitional cell carcinoma (TCC) of the bladder, the major histologic subtype of bladder cancer, is increasing in incidence and mortality, which requires the identification of effective biomarkers. Actin-regulating proteins have recently been proposed as important antitumor druggable targets. As a gelsolin-family actin-modulating protein, CAPG (gelsolin-like actin-capping protein) generated great interest due to its crucial effects in various biological and physiological processes; however, the role and mechanism of CAPG in TCCs remain unknown.Materials and methods:Bioinformatic analysis and immunohistochemistry of clinical specimens were performed to detect the expression level of CAPG. Both in vitro and in vivo assays were used to determine the oncogenic effect of CAPG in TCCs. Male 4–5-week-old BALB/c nude mice were used for in vivo tumorigenesis assays, while SCID mice were used for in vivo metastatic assays. Affymetrix microarray was used to identify the underlying molecular mechanism. Western blot and immunofluorescence were used to validate the expression and localization of proteins.Results:CAPG was frequently upregulated in TCCs and associated with clinical aggressiveness and worse prognosis. Functional assays demonstrated that CAPG could contribute to the tumorigenesis, metastasis and epithelial-mesenchymal transition (EMT) of TCCs both in vitro and in vivo. A novel mechanism that CAPG promoted TCC development via inactivating the Hippo pathway, leading to a nucleus translocation of Yes-associated protein was suggested.Conclusions:The current study identified CAPG as a novel and critical oncogene in TCCs, supporting the pursuit of CAPG as a potential target for TCC intervention.

PI3K\u03b1-regulated gelsolin activity is a critical determinant of cardiac cytoskeletal remodeling and heart disease

Biomechanical stress and cytoskeletal remodeling are key determinants of cellular homeostasis and tissue responses to mechanical stimuli and injury. Here we document the increased activity of gelsolin, an actin filament severing and capping protein, in failing human hearts. Deletion of gelsolin prevents biomechanical stress-induced adverse cytoskeletal remodeling and heart failure in mice. We show that phosphatidylinositol (3,4,5)-triphosphate (PIP3) lipid suppresses gelsolin actin-severing and capping activities. Accordingly, loss of PI3Kα, the key PIP3-producing enzyme in the heart, increases gelsolin-mediated actin-severing activities in the myocardium in vivo, resulting in dilated cardiomyopathy in response to pressure-overload. Mechanical stretching of adult PI3Kα-deficient cardiomyocytes disrupts the actin cytoskeleton, which is prevented by reconstituting cells with PIP3. The actin severing and capping activities of recombinant gelsolin are effectively suppressed by PIP3. Our data identify the role of gelsolin-driven cytoskeletal remodeling in heart failure in which PI3Kα/PIP3 act as negative regulators of gelsolin activity.

CAPZA1 determines the risk of gastric carcinogenesis by inhibiting Helicobacter pylori CagA-degraded autophagy

ABSTRACTHelicobacter pylori-derived CagA, a type IV secretion system effector, plays a role as an oncogenic driver in gastric epithelial cells. However, upon delivery into gastric epithelial cells, CagA is usually degraded by macroautophagy/autophagy. Hence, the induction of autophagy in H. pylori-infected epithelial cells is an important host-protective ability against gastric carcinogenesis. However, the mechanisms by which autophagosome-lysosome fusion is regulated, are unknown. Here, we report that enhancement of LAMP1 (lysosomal associated membrane protein 1) expression is necessary for autolysosome formation. LAMP1 expression is induced by nuclear translocated LRP1 (LDL receptor related protein 1) intracellular domain (LRP1-ICD) binding to the proximal LAMP1 promoter region. Nuclear translocation of LRP1-ICD is enhanced by H. pylori infection. In contrast, CAPZA1 (capping actin protein of muscle Z-line alpha subunit 1) inhibits LAMP1 expression via binding to LRP1-ICD in the nuclei. The binding of CAPZA1 to LRP1-ICD prevents LRP1-ICD binding to the LAMP1 proximal promoter. Thus, in CAPZA1-overexpressing gastric epithelial cells infected with H. pylori, autolysosome formation is inhibited and CagA escapes autophagic degradation. These findings identify CAPZA1 as a novel negative regulator of autolysosome formation and suggest that deregulation of CAPZA1 expression leads to increased risk of gastric carcinogenesis.Abbreviations: CagA: cytotoxin-associated gene A; CAPZA1: capping actin protein of muscle Z-line alpha subunit 1; ChIP: chromatin immunoprecipitation; GTF2I: general transcription factor IIi; HDAC: histone deacetylase; LAMP1: lysosomal associated membrane protein 1; LRP1: LDL receptor related protein 1; LRP1-ICD: CagA intracellular domain; qPCR: quantitative polymerase chain reaction; VacA: vacuolating cytotoxin.

Lack of the Actin Capping Protein, Eps8, Affects NMDA-Type Glutamate Receptor Function and Composition.

Actin-based remodeling underlines spine morphogenesis and plasticity and is crucially involved in the processes that constantly reshape the circuitry of the adult brain in response to external stimuli, leading to learning and memory formation and supporting cognitive functions. Hence spine morphology and synaptic strength are tightly linked and indeed abnormalities in spine number and morphology have been described in a number of neurological disorders such as autism spectrum disorders (ASDs), schizophrenia and intellectual disabilities. We have recently demonstrated that the actin regulating protein, Epidermal growth factor receptor pathway substrate 8 (Eps8), is essential for spine growth and long term potentiation. Indeed, mice lacking Eps8 display immature filopodia-like spines, which are unable to undergo potentiation, and are impaired in cognitive functions. Furthermore, reduced levels of Eps8 have been found in the brain of a cohort of patients affected by ASD compared to controls. Here we investigated whether the lack of Eps8, which is also part of the N-methyl-d-aspartate (NMDA) receptor complex, affects the functional maturation of the postsynaptic compartment. Our results demonstrate that Eps8 knock out mice (Eps8 KO) neurons display altered synaptic expression and subunit composition of NMDA receptors (i.e., increased GluN2B-, decreased GluN2A-containing receptors) and impaired GluN2B to GluN2A subunit shift. Indeed Eps8 KO neurons display increased content of GluN2B containing NMDA receptors both at the synaptic and extrasynaptic level. Furthermore, Eps8 KO neurons display an increased content of extra-synaptic GluN2B-containing receptors, suggesting that also the synaptic targeting of NMDA receptors is affected by the lack of Eps8. These data demonstrate that, besides regulation of spine morphogenesis, Eps8 also regulates the synaptic balance of NMDA receptors subunits GluN2A and GluN2B.

High Throughput, Absolute Determination of the Content of a Selected Protein at Tissue Levels Using Quantitative Dot Blot Analysis (QDB).

Lacking a convenient, quantitative, high throughput immunoblot method for absolute determination of the content of a specific protein at cellular and tissue level significantly hampers the progress in proteomic research. Results derived from currently available immunoblot techniques are also relative, preventing any efforts to combine independent studies with a large-scale analysis of protein samples. In this study, we demonstrate the process of quantitative dot blot analysis (QDB) to achieve absolute quantification in a high throughput format. Using a commercially available protein standard, we are able to determine the absolute content of capping actin protein, gelsolin-like (CAPG) in protein samples prepared from three different mouse tissues (kidney, spleen, and prostate) together with a detailed explanation of the experimental details. We propose the QDB analysis as a convenient, quantitative, high throughput immunoblot method of absolute quantification of individual proteins at the cellular and tissue level. This method will substantially aid biomarker validation and pathway verification in various areas of biological and biomedical research.

High Throughput, Absolute Determination of the Content of a Selected Protein at Tissue Levels Using Quantitative Dot Blot Analysis (QDB)

Lacking a convenient, quantitative, high throughput immunoblot method for absolute determination of the content of a specific protein at cellular and tissue level significantly hampers the progress in proteomic research. Results derived from currently available immunoblot techniques are also relative, preventing any efforts to combine independent studies with a large-scale analysis of protein samples. In this study, we demonstrate the process of quantitative dot blot analysis (QDB) to achieve absolute quantification in a high throughput format. Using a commercially available protein standard, we are able to determine the absolute content of capping actin protein, gelsolin-like (CAPG) in protein samples prepared from three different mouse tissues (kidney, spleen, and prostate) together with a detailed explanation of the experimental details. We propose the QDB analysis as a convenient, quantitative, high throughput immunoblot method of absolute quantification of individual proteins at the cellular and tissue level. This method will substantially aid biomarker validation and pathway verification in various areas of biological and biomedical research.

PKC epsilon signaling effect on actin assembly is diminished in cardiomyocytes when challenged to additional work in a stiff microenvironment.

The stiffness of the microenvironment surrounding a cell can result in cytoskeletal remodeling, leading to altered cell function and tissue macrostructure. In this study, we tuned the stiffness of the underlying substratum on which neonatal rat cardiomyocytes were grown in culture to mimic normal (10 kPa), pathological stiffness of fibrotic myocardium (100 kPa), and a nonphysiological extreme (glass). Cardiomyocytes were then challenged by beta adrenergic stimulation through isoproterenol treatment to investigate the response to acute work demand for cells grown on surfaces of varying stiffness. In particular, the PKCɛ signaling pathway and its role in actin assembly dynamics were examined. Significant changes in contractile metrics were seen on cardiomyocytes grown on different surfaces, but all cells responded to isoproterenol treatment, eventually reaching similar time to peak tension. In contrast, the assembly rate of actin was significantly higher on stiff surfaces, so that only cells grown on soft surfaces were able to respond to acute isoproterenol treatment. Förster Resonance Energy Transfer of immunofluorescence on the cytoskeletal fraction of cardiomyocytes confirmed that the molecular interaction of PKCɛ with the actin capping protein, CapZ, was very low on soft substrata but significantly increased with isoproterenol treatment, or on stiff substrata. Therefore, the stiffness of the culture surface chosen for in vitro experiments might mask the normal signaling and affect the ability to translate basic science more effectively into human therapy.

Capping Actin Protein Overexpression in Human Colorectal Carcinoma and Its Contributed Tumor Migration.

Objective Human colorectal cancer (CRC) is the third most common cancer; patients with metastatic colorectal cancer (mCRC) show poor prognosis than those with CRC cases. There are no reliable molecular biomarkers for the diagnosis of CRC prognosis except with pathological features. Therefore, it is urgent to develop a biomarker for diagnosis and/or prediction of human CRC. In addition, capping actin protein (CapG) belongs to the gelsolin family and has been reported to contribute on tumor invasion/metastasis in multiple human cancers. Here, we are the first to evaluate the expression of CapG in human CRCs. Study Design To investigate the expression levels of CapG in human tissue array by immunohistochemistry (IHC) staining. Moreover, the mRNA and protein levels were also confirmed in four CRC cell lines and determined using real-time RT-PCR and Western blotting. Finally, a Matrigel transwell invasion assay was used to evaluate the invasion ability in CapG high or low expression cells. Results We demonstrated that CapG could be determined in the normal colon tissue and human CRC specimens. However, CapG was significantly overexpressed in the mCRC specimens compared with that in CRC specimens and normal cases. It was also detectable in the four CRC cell lines including mRNA and protein levels. We also found that knockdown of the expression of CapG reduced tumor migration. Conclusions In this study, we suggested that CapG could be used as a biomarker for metastatic CRC in the clinical specimens. Moreover, our in vitro study demonstrated that CapG might contribute on tumor metastasis in human CRCs.

The actin capping protein in Aspergillus nidulans enhances dynein function without significantly affecting Arp1 filament assembly

The minus-end-directed microtubule motor cytoplasmic dynein requires the dynactin complex for in vivo functions. The backbone of the vertebrate dynactin complex is the Arp1 (actin-related protein 1) mini-filament whose barbed end binds to the heterodimeric actin capping protein. However, it is unclear whether the capping protein is a dynactin component in lower eukaryotic organisms, especially because it does not appear to be a component of the budding yeast dynactin complex. Here our biochemical data show that the capping protein is a component of the dynactin complex in the filamentous fungus Aspergillus nidulans. Moreover, deletion of the gene encoding capping protein alpha (capA) results in a defect in both nuclear distribution and early-endosome transport, two dynein-mediated processes. However, the defect in either process is less severe than that exhibited by a dynein heavy chain mutant or the ∆p25 mutant of dynactin. In addition, loss of capping protein does not significantly affect the assembly of the dynactin Arp1 filament or the formation of the dynein-dynactin-∆C-HookA (Hook in A. nidulans) complex. These results suggest that fungal capping protein is not important for Arp1 filament assembly but its presence is required for enhancing dynein function in vivo.

Gelsolin-like Actin-capping Protein (CapG) Overexpression in the Cytoplasm of Human Hepatocellular Carcinoma, Associated with Cellular Invasion, Migration and Tumor Prognosis.

Human hepatocellular carcinoma (HCC) is one of the most common cancers worldwide. Patients with metastatic HCC (mHCC) show poor prognosis and high mortality. In previous reports, gelsolin-like actin-capping protein (CapG) has been demonstrated to regulate cancer invasion and metastasis in various human cancers. In this study, the expression of CapG was verified in normal and/or HCCs' specimens and HCC cell lines. Moreover, the bio-activity of CapG was also investigated. The expression of CapG was examined in HCC's tissue-array by immunohistochemical (IHC) staining. The mRNA and protein of CapG in three HCC cell lines were determined using real-time RT-PCR and western blot. Moreover, a trans-well migration model and a matrigel-trans-well invasion assay were used to address the bio-activity of CapG in HCC cell lines. CapG was detected in the cytoplasm of normal liver tissue and HCC specimens. Importantly, CapG expression was elevated in the HCC specimens compared to normal cases and was significantly overexpressed in mHCC cases compared to normal cases. Moreover, patients with highly expressed CapG showed greater mortality in HCC cases. In addition, the RNA and protein levels of CapG among three HCC cell lines showed a positive association with cellular migration and invasive ability. CapG knockdown with shRNA in HCC cells also verified this finding. In the present study, it is demonstrated that CapG is expressed in the cytoplasm and could be used as a prognostic or diagnostic biomarker for mHCC in clinical specimens. Moreover, CapG might contribute to tumor motility and cancer-associated mortality.

Expression pattern of prohibitin, capping actin protein of muscle Z-line beta subunit and tektin-2 gene in Murrah buffalo sperm and its relationship with sperm motility.

ObjectiveThe aim of the current study is to investigate the relationship between prohibitin (PHB), capping actin protein of muscle Z-line beta subunit (CAPZB), and tektin-2 (TEKT2) and sperm motility in Murrah buffalo.MethodsWe collected the high-motility and low-motility semen samples, testis, ovary, muscle, kidney, liver, brain and pituitary from Murrah buffalo, and analysed the expression of PHB, CAPZB, and TEKT2 in mRNA (message RNA) and protein level.ResultsQuantitative reverse transcription-polymerase chain reaction (qRT-PCR) result showed that the expression of PHB was higher and CAPZB, TEKT2 were specifically expressed in testis as compared to the other 6 tissues, and that in testis, the expression of TEKT2 was higher than that of CAPZB and PHB. Immunohistochemistry test revealed that all three genes were located on the convoluted seminiferous tubule and enriched in spermatogenic cells. Both qRT-PCR and Western Blot results showed that the expression levels of PHB, CAPZB, and TEKT2 were significantly lower in the low-motility semen group compared to the high-motility semen group (p<0.05).ConclusionThe expression levels of PHB, CAPZB, and TEKT2 in Murrah buffalo sperm have a high positive correlation with sperm motility. And the three genes may be potential molecular markers for the decline of buffalo sperm motility.

The role and mechanism of capping actin protein of muscle Z-line alpha subunit 1 in invasion and metastasis of human pancreatic cancer

Objective To observe the expression of capping actin protein of muscle Z-line alpha subunit 1 (CAPZA1) in pancreatic cancer and its relationship with prognosis, and explore the role and mechanism of CAPZA1 in regulating the invasion and metastasis of pancreatic cancer cells. Methods Immunohistochemistry was used to detect the expression of CAPZA1 and epithelial-mesenchymal transition (EMT) markers in pancreatic cancer tissues. The statistical analysis was made on the prognosis of patients with pancreatic cancer. Western blotting and real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) were used to detect the expression of CAPZA1 and EMT markers in pancreatic cancer cells. Transwell, and cell counting kit-8 (CCK-8) were used to detect migration, invasion and proliferation of pancreatic cancer cells. Results The expression of CAPZA1 was negatively correlated with the biological characteristics of pancreatic cancer cells and the prognosis of patients. The expression of CAPZA1 was negatively correlated with the invasion and metastasis potentials of pancreatic cancer cells. Inhibition of CAPZA1 expression could promote the invasion and metastasis of pancreatic cancer cells (192.0±13.8 vs. 153.0±17.1, P=0.004), while over-expression of CAPZA1 could inhibit the invasion and metastasis of pancreatic cancer cells (107.0±18.5 vs. 153.0±17.1, P=0.004). The expression of CAPZA1 was correlated with the expression of EMT markers E-cadherin (0.76±0.09 vs. 0.57±0.14, P=0.034), N-cadherin (0.53±0.12 vs. 0.74±0.10, P=0.017) and Vimentin (0.56±0.20 vs. 0.93±0.19, P=0.017). Conclusion CAPZA1 can inhibit the EMT of pancreatic cancer cells and reduce the invasion ability of them0 by regulating the assembly of cell skeleton. Key words: Pancreatic cancer; Capping actin protein of muscle Z-line alpha subunit 1; Cell skeleton; Invasion; Epithelial-mesenchymal transition

Capping protein regulates actin dynamics during cytokinetic midbody maturation

During cytokinesis, a cleavage furrow generated by actomyosin ring contraction is restructured into the midbody, a platform for the assembly of the abscission machinery that controls the final separation of daughter cells. The polymerization state of F-actin is important during assembly, ingression, disassembly, and closure of the contractile ring and for the cytoskeletal remodeling that accompanies midbody formation and progression to abscission. Actin filaments must be cleared from the abscission sites before the final cut can take place. Although many conserved proteins interact with and influence the polymerization state of actin filaments, it is poorly understood how they regulate cytokinesis in higher eukaryotes. We report here that the actin capping protein (CP), a barbed end actin binding protein, participates in the control of actin polymerization during later stages of cytokinesis in human cells. Cells depleted of CP furrow and form early midbodies, but they fail cytokinesis. Appropriate recruitment of the ESCRT-III abscission machinery to the midbody is impaired, preventing the cell from progressing to the abscission stage. To generate actin filaments of optimal length, different actin nucleators, such as formins, balance CP's activity. Loss of actin capping activity leads to excessive accumulation of formin-based linear actin filaments. Depletion of the formin FHOD1 results in partial rescue of CP-induced cytokinesis failure, suggesting that it can antagonize CP activity during midbody maturation. Our work suggests that the actin cytoskeleton is remodeled in a stepwise manner during cytokinesis, with different regulators at different stages required for successful progression to abscission.

Regulation of IL-1 signaling through control of focal adhesion assembly.

IL-1 signaling is adhesion-restricted in many cell types, but the mechanism that drives it is not defined. We screened for proteins recruited to nascent adhesions in IL-1-treated human fibroblasts with tandem mass tag-mass spectrometry. We used fibronectin bead preparations to enrich 10 actin-associated proteins. There was a 1.2 times log 2-fold enrichment of actin capping protein (ACP) at 30 min after IL-1 stimulation. Knockdown (KD) of ACP by siRNA reduced IL-1-induced ERK activation(by 56%, matrix metalloproteinase-3 (MMP-3) expression by 48%, and MMP-9 expression by 62% (in all reductions, P < 0.01). Confocal or structured illumination microscopy showed that ACP was diffused throughout the cytosol but strongly accumulated at the ruffled border of spreading cells. ACP colocalized with nascent paxillin- and vinculin-containing adhesions at the ruffled border, but not with mature adhesions in the center. ACP KD promoted the formation of large, stable adhesions. Immunoprecipitation and proximity ligation analysis showed that ACP was associated with the IL-1 signal transduction proteins myeloid differentiation factor 88 (MyD88) and IL-1 receptor-associated kinase (IRAK) at the ruffled border of the leading edge. IL-1-induced phospho-ERK and MyD88 or IRAK colocalized at the leading edge. We concluded that ACP is required for recruitment and function of IL-1 signaling complexes in nascent adhesions at the leading edge of the cell.-Wang, Q., Delcorde, J., Tang, T., Downey, G. P., McCulloch, C. A. Regulation of IL-1 signaling through control of focal adhesion assembly.

Actin-capping protein CapG is associated with prognosis, proliferation and metastasis in human glioma.

Glioma is the most aggressive and malignant primary brain tumor in adults. In the present study, we identified a vital oncoprotein, capping actin protein, gelsolin-like (CapG), and investigated its roles in the prognosis, proliferation and metastasis in glioma. The mRNA and protein levels of CapG were significantly increased in human glioma, and higher CapG expression was an independent prognostic factor for predicting unfavorable prognosis. The expression level of CapG was found to be associated with several common molecular features of glioblastoma (GBM; WHO grade IV glioma) in The Cancer Genome Atlas (TCGA) cohort. When analyzing the prognosis of GBM patients according to these molecular features, we observed that the prognostic value of CapG was affected by amplification of CDK6 or EGFR. However, overexpression of CapG markedly promoted cell growth in vitro, while depletion of CapG significantly inhibited cell proliferation by blocking the cell cycle in G1/S transition. Moreover, CapG manipulation in glioma cell lines U87 and U251 showed CapG-dependent cellular migration and invasiveness. These data suggest that CapG may serve as a prognostic biomarker with potentially important therapeutic implications for glioma.

Structural Basis for S100B Interaction with its Target Proteins.

The S100B protein is an intra- and extracellular signaling protein that plays a role in a multitude of cellular processes and abnormal S100B is associated with various neurological diseases and cancers. S100B recognizes and binds effector proteins in a calcium-dependent manner. S100B has been shown to interact with the actin capping protein CapZ, protein kinase C, Hdm2 and 4, RAGE receptor, and p53, among others. These protein partners interact with a common area on the S100B protein surface, validating the method of using the consensus sequence for S100B target search. In addition, each S100B target protein distinguishes itself by additional contacts with S100B. This perspective suggests that the combination of sequence homology search and structural analysis promises to identify newer S100B-binding partners beyond the use of the consensus sequence alone as the given example in the XPB subunit of the TFIIH general transcription factor. XPB is a helicase required for both transcription and DNA repair. Inherited xpb mutations are associated with human disease Xeroderma Pigmentasum, Cockayne syndrome, and trichothiodystrophy. S100B protein is likely associated with much more biological pathways and processes. We believe that S100B will attract more and more attentions in the scientific community and S100B related studies will have important implications in human health and medicine.

Molecular mechanisms of breast cancer metastasis by gene expression profile analysis.

Metastasis is the main cause of breast cancer-related mortalities. The present study aimed to uncover the relevant molecular mechanisms of breast cancer metastasis and to explore potential biomarkers that may be used for prognosis. Expression profile microarray data GSE8977, which contained 22 stroma samples (15 were from normal breast and 7 were from invasive ductal carcinoma tumor samples), were obtained from the Gene Expression Omnibus database. Following data preprocessing, differentially expressed genes (DEGs) were selected based on analyses conducted using the linear models for microarray analysis package from R and Bioconductor software. The resulting data were used in subsequent function and pathway enrichment analyses, as well as protein-protein interaction (PPI) network and subnetwork analyses. Transcription factors (TFs) and tumor-associated genes were also identified among the DEGs. A total of 234 DEGs were identified, which were enriched in immune response, cell differentiation and cell adhesion-related functions and pathways. Downregulated DEGs included TFs, such as the proto-oncogene SPI1, pre-B-cell leukemia homeobox 3 (PBX3) and lymphoid enhancer-binding factor 1 (LEF1), as well as tumor suppressors (TSs), such as capping actin protein, gelsolin like (CAPG) and tumor protein p53-inducible nuclear protein 1 (TP53INP1). Upregulated DEGs also included TFs and tumor suppressors, consisting of transcription factor 7-like 2 (TCF7L2) and pleiomorphic adenoma gene-like 1 (PLAGL1). DEGs that were identified at the hub nodes in the PPI network and the subnetwork were epidermal growth factor receptor (EGFR) and spleen-associated tyrosine kinase (SYK), respectively. Several genes crucial in the metastasis of breast cancer were identified, which may serve as potential biomarkers, many of which were associated with cell adhesion, proliferation or immune response, and may influence breast cancer metastasis by regulating these function or pathways.

An RNAi Screen in a Novel Model of Oriented Divisions Identifies the Actin-Capping Protein Z β as an Essential Regulator of Spindle Orientation

Oriented cell divisions are controlled by a conserved molecular cascade involving Gαi, LGN, and NuMA. We developed a new cellular model of oriented cell divisions combining micropatterning and localized recruitment of Gαi and performed an RNAi screen for regulators acting downstream of Gαi. Remarkably, this screen revealed a unique subset of dynein regulators as being essential for spindle orientation, shedding light on a core regulatory aspect of oriented divisions. We further analyze the involvement of one novel regulator, the actin-capping protein CAPZB. Mechanistically, we show that CAPZB controls spindle orientation independently of its classical role in the actin cytoskeleton by regulating the assembly, stability, and motor activity of the dynein/dynactin complex at the cell cortex, as well as the dynamics of mitotic microtubules. Finally, we show that CAPZB controls planar divisions invivo in the developing neuroepithelium. This demonstrates the power of this in cellulo model of oriented cell divisions to uncover newgenes required in spindle orientation in vertebrates.