The EF-hand calcium-binding protein calmyrin inhibits the transcriptional and DNA-binding activity of Pax3

ABSTRACTHepatitis C virus (HCV) is a major cause of chronic liver disease and is highly dependent on cellular proteins for virus propagation. To identify the cellular factors involved in HCV propagation, we recently performed protein microarray assays using the HCV nonstructural 5A (NS5A) protein as a probe. Of 90 cellular protein candidates, we selected the soluble resistance-related calcium-binding protein (sorcin) for further characterization. Sorcin is a calcium-binding protein and is highly expressed in certain cancer cells. We verified that NS5A interacted with sorcin through domain I of NS5A, and phosphorylation of the threonine residue 155 of sorcin played a crucial role in protein interaction. Small interfering RNA (siRNA)-mediated knockdown of sorcin impaired HCV propagation. Silencing of sorcin expression resulted in a decrease of HCV assembly without affecting HCV RNA and protein levels. We further demonstrated that polo-like kinase 1 (PLK1)-mediated phosphorylation of sorcin was increased by NS5A. We showed that both phosphorylation and calcium-binding activity of sorcin were required for HCV propagation. These data indicate that HCV modulates sorcin activity via NS5A protein for its own propagation.IMPORTANCESorcin is a calcium-binding protein and regulates intracellular calcium homeostasis. HCV NS5A interacts with sorcin, and phosphorylation of sorcin is required for protein interaction. Gene silencing of sorcin impaired HCV propagation at the assembly step of the HCV life cycle. Sorcin is phosphorylated by PLK1 via protein interaction. We showed that sorcin interacted with both NS5A and PLK1, and PLK1-mediated phosphorylation of sorcin was increased by NS5A. Moreover, calcium-binding activity of sorcin played a crucial role in HCV propagation. These data provide evidence that HCV regulates host calcium metabolism for virus propagation, and thus manipulation of sorcin activity may represent a novel therapeutic target for HCV.

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The Heparin-Binding Activity of Secreted Modular Calcium-Binding Protein 1 (SMOC-1) Modulates Its Cell Adhesion Properties

PLoS ONE ◽

10.1371/journal.pone.0056839 ◽

2013 ◽

Vol 8 (2) ◽

pp. e56839 ◽

Cited By ~ 11

Author(s):

Marina Klemenčič ◽

Marko Novinec ◽

Silke Maier ◽

Ursula Hartmann ◽

Brigita Lenarčič

Keyword(s):

Cell Adhesion ◽

Calcium Binding ◽

Binding Protein ◽

Binding Activity ◽

Calcium Binding Protein ◽

Heparin Binding ◽

Adhesion Properties

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UV stimulation induces nuclear factor κB (NF-κB) DNA-binding activity but not transcriptional activation

Biochemical Society Transactions ◽

10.1042/bst0290688 ◽

2001 ◽

Vol 29 (6) ◽

pp. 688-691 ◽

Cited By ~ 18

Author(s):

K. J. Campbell ◽

N. R. Chapman ◽

N. D. Perkins

Keyword(s):

Dna Binding ◽

Transcriptional Activation ◽

Cellular Response ◽

Uv Light ◽

Binding Protein ◽

Nuclear Factor Κb ◽

Camp Response Element Binding ◽

Binding Activity ◽

Nuclear Factor ◽

Dna Binding Activity

The cellular response to DNA-damaging agents is partly mediated by DNA-binding transcription factors such as p53 and nuclear factor κB (NF-κB). Typically NF-κB activation is associated with resistance to apoptosis. Following stimulation with UV light however, NF-κB activation has been shown to be required for programmed cell death. To study this effect further and to determine the relationship between NF-κB and p53 function, we have examined the effect of UV light on U2OS cells. UV stimulation resulted in the activation of NF-κB DNA-binding and the induction of p53. Surprisingly, and in contrast with tumour necrosis factor α stimulation, this UV-induced NF-κB was transcriptionally inert. These observations suggest a model in which the NF-κB switch from an anti-apoptotic to a pro-apoptotic role within the cell results from modulation of its ability to stimulate gene expression, possibly as a result of the ability of p53 to sequester transcriptional co-activator proteins such as p300/CREB (cAMP-response-element-binding protein)-binding protein.

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A novel 40-kDa membrane-associated EF-hand calcium-binding protein in Plasmodium falciparum.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)50621-4 ◽

1992 ◽

Vol 267 (6) ◽

pp. 3976-3982

Author(s):

D.J. Rawlings ◽

D.C. Kaslow

Keyword(s):

Plasmodium Falciparum ◽

Calcium Binding ◽

Binding Protein ◽

Calcium Binding Protein ◽

Ef Hand

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A novel DNA-binding motif in the nuclear matrix attachment DNA-binding protein SATB1

Molecular and Cellular Biology ◽

10.1128/mcb.14.3.1852-1860.1994 ◽

1994 ◽

Vol 14 (3) ◽

pp. 1852-1860

Author(s):

K Nakagomi ◽

Y Kohwi ◽

L A Dickinson ◽

T Kohwi-Shigematsu

Keyword(s):

Amino Acid ◽

Dna Binding ◽

Direct Contact ◽

Binding Protein ◽

Binding Activity ◽

Binding Domain ◽

Binding Motif ◽

Dna Binding Domain ◽

Sequence Context ◽

Dna Binding Activity

The nuclear matrix attachment DNA (MAR) binding protein SATB1 is a sequence context-specific binding protein that binds in the minor groove, making virtually no contact with the DNA bases. The SATB1 binding sites consist of a special AT-rich sequence context in which one strand is well-mixed A's, T's, and C's, excluding G's (ATC sequences), which is typically found in clusters within different MARs. To determine the extent of conservation of the SATB1 gene among different species, we cloned a mouse homolog of the human STAB1 cDNA from a cDNA expression library of the mouse thymus, the tissue in which this protein is predominantly expressed. This mouse cDNA encodes a 764-amino-acid protein with a 98% homology in amino acid sequence to the human SATB1 originally cloned from testis. To characterize the DNA binding domain of this novel class of protein, we used the mouse SATB1 cDNA and delineated a 150-amino-acid polypeptide as the binding domain. This region confers full DNA binding activity, recognizes the specific sequence context, and makes direct contact with DNA at the same nucleotides as the whole protein. This DNA binding domain contains a novel DNA binding motif: when no more than 21 amino acids at either the N- or C-terminal end of the binding domain are deleted, the majority of the DNA binding activity is lost. The concomitant presence of both terminal sequences is mandatory for binding. These two terminal regions consist of hydrophilic amino acids and share homologous sequences that are different from those of any known DNA binding motifs. We propose that the DNA binding region of SATB1 extends its two terminal regions toward DNA to make direct contact with DNA.

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