No abstract

The impairment of retinoic acid (RA)-dependent signaling is a frequent event during carcinogenesis. Cellular retinoic acid-binding proteins (CRABP1 and CRABP2) are important modulators of RA activity. Up to date, the role of these proteins in cancer progression remains poorly investigated. Here, we studied for the first time the simultaneous messenger RNA (mRNA) and protein expression of CRABPs in non-small cell lung cancer (NSCLC) samples. CRABP1 and CRABP2 mRNA levels were elevated in 42 and 56 % of NSCLC samples, respectively. Decrease of CRABP2 mRNA expression was significantly associated with the presence of lymph node metastases. Protein expression of CRABP1 and CRABP2 was detected in 50 and 56 % of tumor samples, respectively. We also found a positive correlation between CRABP1 and CRABP2 expression. Taken together, we demonstrated significant changes in CRABP expression in NSCLC samples. Importantly, the presented data provide the first evidence of potential involvement of CRABP2 in lung cancer metastasis.

RAS oncogenes play a critical role in oncogenic transformation and metastases formation. Here we show that Ha-ras greatly stimulates spontaneous metastatic activity of transformed cells through the Ras/RalGDS/ RalA intracellular signaling pathway. Introduction of RalA alone leads to a drastic increase of metastatic activity of transformed cells. We demonstrate that metastatic ability of cells could be dramatically enhanced by RalA stimulation or, conversely, hampered by RalA suppression. Furthermore, we found that during in vivo selection cells acquire high metastatic properties as a result of endogenous RalA activation. The ability of RalA to induce metastasis was demonstrated in spontaneously transformed as well as in virus transformed fibroblasts.

CRABP1 (cellular retinoic acid binding protein 1) belongs to the family of fatty acid binding proteins. Retinoic acid binding is the only known functional activity of this protein. The role of CRABP1 in human carcinogenesis remains poorly understood. Here, for the first time we demonstrated pro-metastatic and pro-tumorigenic activity of CRABP1 in mesenchymal tumors. Further functional analysis revealed that the pro-tumorigenic effect of CRABP1 does not depend on retinoic acid binding activity. These results suggest that CRABP1 could have an alternative intracellular functional activity that contributes to the high malignancy of transformed mesenchymal cells. Microarray analysis detected CRABP1-mediated alterations in the expression of about 100 genes, including those encoding key regulatory proteins. CRABP1 is ubiquitously expressed in monophasic synovial sarcomas, while in biphasic synovial sarcomas it is expressed uniquely by the spindle cells of the aggressive mesenchymal component. High level of CRABP1 expression is associated with lymph node metastasis and poor differentiation/high grade of pancreatic neuroendocrine tumors (pNETs). Presented data suggest CRABP1 as a promising biomarker of pNETs' clinical behavior. Our results give the first evidence of pro-tumorigenic and pro-metastatic activity of CRABP1 in mesenchymal and neuroendocrine tumors.

Proteins involved in the organizing of lipid rafts can be found in exosomes, as shown for caveolin‐1, and they could contribute to exosomal cargo sorting, as shown for flotillins. Stomatin belongs to the same stomatin/prohibitin/flotillin/HflK/C family of lipid rafts proteins, but it has never been studied in exosomes except for extracellular vesicles (EVs) originating from blood cells. Here we first show the presence of stomatin in exosomes produced by epithelial cancer cells (non–small cell lung cancer, breast, and ovarian cancer cells) as well as in EVs from biological fluids, including blood plasma, ascitic fluids, and uterine flushings. A high abundance of stomatin in EVs of various origins and its enrichment in exosomes make stomatin a promising exosomal marker. Comparison with other lipid raft proteins and exosomal markers showed that the level of stomatin protein in exosomes from different sources corresponds well to that of CD9, while it differs essentially from flotillin‐1 and flotillin‐2 homologs, which in turn are present in exosomes in nearly equal proportions. In contrast, the level of vesicular caveolin‐1 as well as its EV‐to‐cellular ratio vary drastically depending on cell type.

No abstract

Protein Phosphorylation as a Key Mechanism of mTORC1/2 Signaling Pathways 5 TRRAP (PIKK family members)) domain and FRB (FKPB12-rapamycin binding domain), which serves as a docking site for the rapamycin -FKBP12 complex formation. Downstream lies a catalytic kinase domain and a FATC (FAT Carboxyterminal) domain, located at the Cterminus of the protein ( Figure 1A). The FAT and FATC domains are always found in combination, so it has been hypothesized that the interactions between FAT and FATC might contribute to the catalytic kinase activity of mTOR via unknown mechanisms [26,27].

The small G-protein ADP-ribosylation factor 6 (Arf6) belongs to the Ras GTPases superfamily and is mostly known for its actin remodeling functions and involvement in the processes of plasma membrane reorganization and vesicular transport. The majority of data indicates that Arf6 contributes to cancer progression through activation of cell motility and invasion. Alternatively, we found that the expression of a wild-type or a constitutively active Arf6 does not influence tumor cell motility and invasion but instead significantly stimulates cell proliferation and activates phospholipase D (PLD). Conversely the expression of a mutant Arf6 (Arf6N48I), that is, unable to interact with PLD has no effect on proliferation but promotes motility, invasion, and matrix degradation by uPA extracellular proteinase. Studying the mechanisms of Arf6-dependent stimulation of cell proliferation, we found some signaling pathways contributing to Arf6 promitogenic activity. Namely, we showed that Arf6 in a PLD-mTORC1-dependent manner activates S6K1 kinase, a well-known regulator of mitogen-stimulated translation initiation. Furthermore, we demonstrated an Arf6-dependent phosphorylation of mTORC1 downstream targets, 4E-BP1 and ribosomal S6 protein, confirming an existence of Arf6-PLD-mTORC1-S6K1/4E-BP1 signaling pathway and also demonstrated its impact on proliferation stimulation. Next, we found that Arf6 activation potentiates Erk1/2 and p38MAP kinases phosphorylation. Surprisingly, p38 opposite to Erk1/2 significantly contributes to Arf6-dependent proliferation increase promoting S6 ribosomal protein phosphorylation at Ser235/236 residues. Therefore, we demonstrated Arf6 proliferation stimulating activity and revealed PLD-mTORC1 and p38MAP kinase as Arf6 partners mediating promitogenic activity. These results highlight a new aspect of Arf6 functioning in cancer cell biology.