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VILIP-1, a member of the neuronal Ca2+ sensor protein family, is able to act as a tumor suppressor in carcinoma cells by inhibiting cell proliferation and migration. In order to study the role of VILIP-1 in skin carcinogenesis we generated transgenic mice overexpressing VILIP-1 in epidermis under the control of the bovine keratin K5 promoter (K5-VILIP-1). We studied the susceptibility of FVB wild type and VILIP-1 transgenic mice to chemically mediated carcinogenesis. After 30 weeks of treatment with a two-stage carcinogenesis protocol, all animals showed numerous skin tumors. Nevertheless, K5- VILIP-1 mice showed decreased squamous cell carcinoma (SCC) multiplicity of ~49% (p<0.02) with respect to the corresponding SCC multiplicity observed in wild type (WT) mice. In addition, the relative percentage of low-grade cutaneous SCCs grade I (defined by the differentiation pattern according to the Broders grading scale) increased approximately 50% in the K5-VILIP1 mice when compared with SCCs in WT mice. Similar tendency was observed using a complete carcinogenesis protocol for skin carcinogenesis using benzo(a)pyrene (B(a)P). Further studies of tumors and primary epidermal keratinocyte cultures showed that matrix metalloproteinase 9 (MMP-9) levels and cell proliferation decreased in K5-VILIP-1 mice when compared with their wild counterparts. In addition tissue inhibitor of metalloproteinase 1 (TIMP-1) expression was higher in K5-VILIP-1 keratinocytes. These results show that VILIP-1 overexpression decreases the susceptibility to skin carcinogenesis in experimental mouse cancer models, thus supporting its role as a tumor suppressor gene. [ABSTRACT FROM AUTHOR]

The visinin-like protein (VSNL) subfamily, including VILIP-1 (the founder protein), VILIP-2, VILIP-3, hippocalcin, and neurocalcin δ, constitute a highly homologous subfamily of neuronal calcium sensor (NCS) proteins. Comparative studies have shown that VSNLs are expressed predominantly in the brain with restricted expression patterns in various subsets of neurons but are also found in peripheral organs. In addition, the proteins display differences in their calcium affinities, in their membrane-binding kinetics, and in the intracellular targets to which they associate after calcium binding. Even though the proteins use a similar calcium-myristoyl switch mechanism to translocate to cellular membranes, they show calcium-dependent localization to various subcellular compartments when expressed in the same neuron. These distinct calcium-myristoyl switch properties might be explained by specificity for defined phospholipids and membrane-bound targets; this enables VSNLs to modulate various cellular signal transduction pathways, including cyclic nucleotide and MAPK signaling. An emerging theme is the direct or indirect effect of VSNLs on gene expression and their interaction with components of membrane trafficking complexes, with a possible role in membrane trafficking of different receptors and ion channels, such as glutamate receptors of the kainate and AMPA subtype, nicotinic acetylcholine receptors, and Ca2+-channels. One hypothesis is that the highly homologous VSNLs have evolved to fulfil specialized functions in membrane trafficking and thereby affect neuronal signaling and differentiation in defined subsets of neurons. VSNLs are involved in differentiation processes showing a tumor-invasion-suppressor function in peripheral organs. Finally, VSNLs play neuroprotective and neurotoxic roles and have been implicated in neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

VILIP-1, a member of the neuronal Ca++ sensor protein family, acts as a tumor suppressor gene in an experimental animal model by inhibiting cell proliferation, adhesion and invasiveness of squamous cell carcinoma cells. Western Blot analysis of human tumor cells showed that VILIP-1 expression was undetectable in several types of human tumor cells, including 11 out of 12 non-small cell lung carcinoma (NSCLC) cell lines. The down-regulation of VILIP-1 was due to loss of VILIP-1 mRNA transcripts. Rearrangements, large gene deletions or mutations were not found. Hypermethylation of the VILIP-1 promoter played an important role in gene silencing. In most VILIP-1-silent cells the VILIP-1 promoter was methylated. In vitro methylation of the VILIP-1 promoter reduced its activity in a promoter-reporter assay. Transcriptional activity of endogenous VILIP-1 promoter was recovered by treatment with 5′-aza-2′-deoxycytidine (5′-Aza-dC). Trichostatin A (TSA), a histone deacetylase inhibitor, potently induced VILIP-1 expression, indicating that histone deacetylation is an additional mechanism of VILIP-1 silencing. TSA increased histone H3 and H4 acetylation in the region of the VILIP-1 promoter. Furthermore, statistical analysis of expression and promoter methylation (n = 150 primary NSCLC samples) showed a significant relationship between promoter methylation and protein expression downregulation as well as between survival and decreased or absent VILIP-1 expression in lung cancer tissues (p<0.0001). VILIP-1 expression is silenced by promoter hypermethylation and histone deacetylation in aggressive NSCLC cell lines and primary tumors and its clinical evaluation could have a role as a predictor of short-term survival in lung cancer patients. [ABSTRACT FROM AUTHOR]

The family of neuronal Ca2+ sensor (NCS) proteins is known to influence a variety of physiological and pathological processes by affecting signalling of different receptors and ion channels. Recently, it has been shown that the NCS protein VILIP-1 influences the activity of the receptor guanylyl cyclase GC-B. In transfected cell lines, VILIP-1 performs a Ca2+-dependent membrane association, the reversible Ca2+-myristoyl switch of VILIP-1, which leads to an increase in natriuretic peptide-stimulated cGMP levels. In this study, we have investigated the effect of VILIP-1 on cGMP signalling in C6 cells and in primary hippocampal neurons, where VILIP-1 and GC-B are co-expressed in many but not all neurons and partially co-localize in the soma and in dendrites. Our data indicate that VILIP-1 modulates GC-B activity by influencing clathrin-dependent receptor recycling. These data support a general physiological role for VILIP-1 in membrane trafficking in the intact hippocampus, where the NCS protein may affect processes, such as neuronal differentiation and synaptic plasticity e.g. by influencing cGMP-signalling. [ABSTRACT FROM AUTHOR]

Visinin-like protein-1 (VILIP-1) is a member of the neuronal EF-hand Ca(2+)-sensor protein family. VILIP-1 is expressed in the central nervous system where it plays a crucial role in regulating cAMP levels, cell signaling, and differentiation. Screening of mouse skin tumor cell lines for differentially expressed genes showed high-level VILIP-1 expression in less aggressive squamous cell carcinoma (SCC) and papilloma cell lines. Conversely, expression was markedly decreased or lost in invasive SCC and spindle cell carcinoma cell lines. In addition, immunohistochemistry of normal skin and primary tumors showed that VILIP-1 is expressed in basal cells of the normal intrafollicular epidermis as well as in basal cells of papillomas. The expression was decreased in low-grade SCCs and disappeared in most high-grade SCCs. When two high-grade carcinoma cell lines were transfected with VILIP1-cDNA, the VILIP-1 transfectants had significantly higher cAMP levels than the respective vector alone-transfected lines. VILIP-1-transfected cells were less invasive (both in vivo and in vitro) than the control transfectants. Reduced invasiveness and elevation of cAMP levels were accompanied by decreased MMP-9, as well as decreased RhoA activity. These results indicate that VILIP-1 plays an important role in regulating tumor cell invasiveness and that its loss could aid in enhancing the advanced malignant phenotype.