Functional role of inositol‐1,4,5‐trisphosphate‐3‐kinase‐A for motility of malignant transformed cells

Abstract

Cell migration is one of the hallmarks of metastatic disease and thus identification of migration promoting proteins is crucial for the understanding of metastasis formation. Here we show that the neuron-specific, F-actin bundling inositol-1,4,5-trisphosphate-3-kinase-A (ITPKA) is ectopically expressed in tumor cells and critically involved in migration. Down-regulation of ITPKA expression in transformed cell-lines with ectopic expression of ITPKA significantly decreased migration and the number of linear and branched cell protrusion. Conversely, up-regulation of ITPKA in tumor cell lines with low endogenous ITPKA expression increased migration and formation of cell processes. In vitro, ITPKA alone induced the formation of linear actin filaments, whereas ITPKA mediated formation of branched protrusions seems to result from interaction between ITPKA and the F-actin cross-linking protein filamin C. Based on these actin-modulating and migration-promoting effects of ITPKA we examined its expression in clinical samples of different tumor entities, starting with the analysis of multiple tumor tissue arrays. As in lung adenocarcinoma specimens, the highest ITPKA expression rate was found, this tumor entity was examined in more detail. ITPKA was expressed early in adenocarcinoma progression (pN0) and was largely maintained in invasive and metastatic tumor cell populations (pN1/2, lymph node metastases). Together with our result that high expression of ITPKA increases motility of tumor cells we conclude that the observed expression of ITPKA early in tumor development increases the metastatic potential of lung adenocarcinoma cells. Therefore, we suggest that ITPKA may be a promising therapeutic molecular target for anti metastatic therapy of lung cancer.Under physiological conditions, inositol-1,4,5-trisphosphate-3-kinase-A (ITPKA, GeneAtlas, V133A gcma, BioGPS, http:// biogps.gnf.org/#goto¼genereport&id¼3706) is only expressed in hippocampal, cortical and cerebellar neurons.1 In these cells expression of ITPKA increases during brain development 1 and is highest in mature neurons, where the protein accumulates in dendritic spines.2 Through conversion of inositol-1,4,5-trisphosphate (InsP 3 ) to inositol-1,3,4,5-tetrakisphosphate (InsP 4 ) ITPKA is an important regulator of InsP 3 induced calcium signaling in dendritic spines. 2,3 In addition to this catalytic activity, it has been shown that ITPKA has Factin bundling activity. It bundles actin filaments by formation of homodimers and binding to F-actin, making ITPKA to an important regulator of spine morphology. 4 Beside this physiological role, we recently have found a pathological function of ITPKA in tumor cells. We revealed that particular tumor cell lines express ITPKA and demonstrated that this ectopic expression of ITPKA increased the metastatic potential of tumor cells. Because of its F-actin bundling activity, ITPKA induces the formation of filopodia-and lamellipodia-like protrusions, 5,6 which are critical for cells to migrate. 7 This nonenzymatic migration-prom...

Expression Regulation of the Metastasis-Promoting Protein InsP3-Kinase-A in Tumor Cells

“…Antibodies were purchased from Santa Cruz: GAPDH (Sc69778), cyclin A (Sc-596), and ITPKA (Sc69778). Specifity of the ITPKA antibody was evaluated in a former study (7). In addition, brain lysates of ITPKA wild-type and knockout mice were analyzed by Western blotting (Supplementary Fig.…”

“…We found ectopic expression of the neuron-specific isoform A of ITPK in particular tumor cell lines and revealed that this upregulated expression increased their metastatic potential (6)(7)(8). Therefore, we examined the mechanism underlying regulation of the ITPKA level.…”

“…Because of this association and its characteristic to form homodimmers, ITPKA bundles actin filaments, resulting in increased size and/or number of dendritic spines (4,5). In tumor cells that have gained the ability to express ITPKA, the F-actin bundling activity of ITPKA, and its interaction with the Factin cross-linking protein filamin C, induces the formation of new filopodia and lamellipodia (6)(7)(8). As formation of these cellular protrusions is a prerequisite for cells to migrate, the actin-modulating activity of ITPKA increases the migratory and the metastatic potential of tumor cells (6).…”

“…In addition, ITPKA promotes migration by its ITPK activity mediating the induction of store-operated calcium entry and with this the stimulation of calcium-dependent actin-modulating proteins (6). Current studies on lung cancer patients show that adenocarcinoma cells express high levels of ITPKA in the primary tumors and in metastasis, whereas ITPKA expression was mainly found in metastasis in small cell and squamous carcinoma (7). Furthermore, it has been shown that only particular tumor cell lines could express ITPKA (6,7).…”

“…Current studies on lung cancer patients show that adenocarcinoma cells express high levels of ITPKA in the primary tumors and in metastasis, whereas ITPKA expression was mainly found in metastasis in small cell and squamous carcinoma (7). Furthermore, it has been shown that only particular tumor cell lines could express ITPKA (6,7). However, the mechanism underlying this tumor cell and stage-specific regulation of ITPKA expression has not been elucidated yet.…”