Divergent roles of lysyl oxidase family members in ornithine decarboxylase-and RAS-transformed mouse fibroblasts and human melanoma cells

We have previously shown that proto-oncoprotein c-Jun is activated in ornithine decarboxylase (ODC)- and RAS-transformed mouse fibroblasts, and that the transformed morphology of these cells can be reversed by expressing the transactivation domain deletion mutant of c-Jun (TAM67). Here, we found that lysyl oxidase (Lox), encoding an extracellular matrix-modifying enzyme, is downregulated in a c-Jun-dependent manner in ODC-transformed fibroblasts (Odc cells). In addition to Lox, the Lox family members Lox-like 1 and 3 (Loxl1 and Loxl3) were found to be downregulated in Odc as well as in RAS-transformed fibroblasts (E4), whereas Lox-like 4 (Loxl4) was upregulated in Odc and downregulated in E4 cells compared to normal N1 fibroblasts. Tetracycline-regulatable LOX re-expression in Odc cells led to inhibition of cell growth and invasion in three-dimensional Matrigel in an activity-independent manner. On the contrary, LOX and especially LOXL2, LOXL3, and LOXL4 were found to be upregulated in several human melanoma cell lines, and LOX inhibitor B-aminopropionitrile inhibited the invasive growth of these cells particularly when co-cultured with fibroblasts in Matrigel. Knocking down the expression of LOX and especially LOXL2 in melanoma cells almost completely abrogated the invasive growth capability. Further, LOXL2 was significantly upregulated in clinical human primary melanomas compared to benign nevi, and high expression of LOXL2 in primary melanomas was associated with formation of metastases and shorter survival of patients. Thus, our studies reveal that inactive pro-LOX (together with Lox propeptide) functions as a tumor suppressor in ODC- and RAS-transformed murine fibroblasts by inhibiting cell growth and invasion, and active LOX and LOXL2 as tumor promoters in human melanoma cells by promoting their invasive growth. Copyright: Kielosto et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0 (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Peer reviewe

Measuring turbulent CO₂ fluxes with a closed-path gas analyzer in a marine environment

In this study, we introduce new observations of sea–air fluxes of carbon dioxide using the eddy covariance method. The measurements took place at the Utö Atmospheric and Marine Research Station on the island of Utö in the Baltic Sea in July–October 2017. The flux measurement system is based on a closed-path infrared gas analyzer (LI-7000, LI-COR) requiring only occasional maintenance, making the station capable of continuous monitoring. However, such infrared gas analyzers are prone to significant water vapor interference in a marine environment, where CO2 fluxes are small.Two LI-7000 analyzers were run in parallel to test the effect of a sample air drier which dampens water vapor fluctuations and a virtual impactor, included to remove liquid sea spray, both of which were attached to the sample air tubing of one of the analyzers. The systems showed closely similar (R2 = 0.99) sea–air CO2 fluxes when the latent heat flux was low, which proved that neither the drier nor the virtual impactor perturbed the CO2 flux measurement. However, the undried measurement had a positive bias that increased with increasing latent heat flux, suggesting water vapor interference.For both systems, cospectral densities between vertical wind speed and CO2 molar fraction were distributed within the expected frequency range, with a moderate attenuation of high-frequency fluctuations. While the setup equipped with a drier and a virtual impactor generated a slightly higher flux loss, we opt for this alternative for its reduced water vapor cross-sensitivity and better protection against sea spray. The integral turbulence characteristics were found to agree with the universal stability dependence observed over land. Nonstationary conditions caused unphysical results, which resulted in a high percentage (65&thinsp;%) of discarded measurements. After removing the nonstationary cases, the direction of the sea–air CO2 fluxes was in good accordance with independently measured CO2 partial pressure difference between the sea and the atmosphere. Atmospheric CO2 concentration changes larger than 2&thinsp;ppm during a 30&thinsp;min averaging period were found to be associated with the nonstationarity of CO2 fluxes.</p

The mucin MUC4 and its membrane partner ErbB2 regulate biological properties of human CAPAN-2 pancreatic cancer cells via different signalling pathways.

The mucin MUC4 and its membrane partner the ErbB2 oncogenic receptor are potential interacting partners in human pancreatic tumour development. However, the way they function is still largely unknown. In this work, we aimed to identify the cellular mechanisms and the intracellular signalling pathways under the control of both ErbB2 and MUC4 in a human pancreatic adenocarcinomatous cell line. Using co-immunoprecipitation and GST pull-down, we show that MUC4 and ErbB2 interact in the human pancreatic adenocarcinomatous cell line CAPAN-2 via the EGF domains of MUC4. Stable cell clones were generated in which either MUC4 or ErbB2 were knocked down (KD) by a shRNA approach. Biological properties of these cells were then studied in vitro and in vivo. Our results show that ErbB2-KD cells are more apoptotic and less proliferative (decreased cyclin D1 and increased p27kip1 expression) while migration and invasive properties were not altered. MUC4-KD clones were less proliferative with decreased cyclin D1 expression, G1 cell cycle arrest and altered ErbB2/ErbB3 expression. Their migration properties were reduced whereas invasive properties were increased. Importantly, inhibition of ErbB2 and MUC4 expression did not impair the same signalling pathways (inhibition of MUC4 expression affected the JNK pathway whereas that of ErbB2 altered the MAPK pathway). Finally, ErbB2-KD and MUC4-KD cells showed impaired tumour growth in vivo. Our results show that ErbB2 and MUC4, which interact physically, activate different intracellular signalling pathways to regulate biological properties of CAPAN-2 pancreatic cancer cells