Mig1 and Hxk2 are two major mediators of glucose repression in Saccharomyces cerevisiae. However, the mechanism by which Hxk2 participates in the glucose repression signaling pathway is not completely understood. Recently, it has been demonstrated that Hxk2 interacts with Mig1 to generate a repressor complex located in the nucleus of S. cerevisiae. However, the mechanism by which Mig1 favors the presence of Hxk2 in the nucleus is not clear, and the function of Hxk2 at the nuclear repressor complex level is still unknown. Here, we report that serine 311 of Mig1 is a critical residue for interaction with Hxk2 and that this interaction is regulated by glucose. Our findings suggest that Snf1 interacts constitutively with the Hxk2 component of the repressor complex at high and low glucose conditions. Furthermore, we show that Snf1 binds to Mig1 under low glucose conditions and that binding is largely abolished after a shift to high glucose medium. We found that phosphorylation of serine 311 of Mig1 by Snf1 kinase is essential for Mig1 protein nuclear export and derepression of the SUC2 gene in glucose-limited cells. These results allow postulating that the Hxk2 operates by interacting both with Mig1 and Snf1 to inhibit the Mig1 phosphorylation at serine 311 during high glucose grown.

Microfluidic devices are becoming mainstream tools to recapitulate in vitro the behavior of cells and tissues. In this study, we use microfluidic devices filled with hydrogels of mixed collagen-Matrigel composition to study the migration of lung cancer cells under different cancer invasion microenvironments. We present the design of the microfluidic device, characterize the hydrogels morphologically and mechanically and use quantitative image analysis to measure the migration of H1299 lung adenocarcinoma cancer cells in different experimental conditions. Our results show the plasticity of lung cancer cell migration, which turns from mesenchymal in collagen only matrices, to lobopodial in collagen-Matrigel matrices that approximate the interface between a disrupted basement membrane and the underlying connective tissue. Our quantification of migration speed confirms a biphasic role of Matrigel. At low concentration, Matrigel facilitates migration, most probably by providing a supportive and growth factor retaining environment. At high concentration, Matrigel slows down migration, possibly due excessive attachment. Finally, we show that antibody-based integrin blockade promotes a change in migration phenotype from mesenchymal or lobopodial to amoeboid and analyze the effect of this change in migration dynamics, in regards to the structure of the matrix. In summary, we describe and characterize a robust microfluidic platform and a set of software tools that can be used to study lung cancer cell migration under different microenvironments and experimental conditions. This platform could be used in future studies, thus benefitting from the advantages introduced by microfluidic devices: precise control of the environment, excellent optical properties, parallelization for high throughput studies and efficient use of therapeutic drugs.

Nucleocytoplasmic shuttling of Hxk2 induced by glucose levels has been reported recently. Here we present evidence that indicates that Hxk2 nucleocytoplasmic traffic is regulated by phosphorylation and dephosphorylation at serine 14. Moreover, we identified the protein kinase Snf1 and the protein phosphatase Glc7-Reg1 as novel regulatory partners for the nucleocytoplasmic shuttling of Hxk2. Functional studies revealed that, in contrast to the wild-type protein, the dephosphorylation-mimicking mutant of Hxk2 retains its nuclear localization in low glucose conditions, and the phosphomimetic mutant of Hxk2 retains its cytoplasmic localization in high glucose conditions. Interaction experiments of Hxk2 with Kap60 and Xpo1 indicated that nuclear import of the S14D mutant of Hxk2 is severely decreased but that the export is significantly enhanced. Conversely, nuclear import of the S14A mutant of Hxk2 was significantly enhanced, although the export was severely decreased. The interaction of Hxk2 with Kap60 and Xpo1 was found to occur in the dephosphorylated and phosphorylated states of the protein, respectively. In addition, we found that Hxk2 is a substrate for Snf1. Mutational analysis indicated that serine 14 is a major in vitro and in vivo phosphorylation site for Snf1. We also provide evidence that dephosphorylation of Hxk2 at serine 14 is a protein phosphatase Glc7-Reg1-dependent process. Taken together, this study establishes a functional link between Hxk2, Reg1, and Snf1 signaling, which involves the regulation of Hxk2 nucleocytoplasmic shuttling by phosphorylation-dephosphorylation of serine 14.

In many tissues, stearoyl-CoA desaturase 1 (SCD1) catalyzes the biosynthesis of monounsaturated fatty acids (MUFAS), (i.e., palmitoleate and oleate) from their saturated fatty acid (SFA) precursors (i.e., palmitate and stearate), influencing cellular membrane physiology and signaling, leading to broad effects on human physiology. In addition to its predominant role in lipid metabolism and body weight control, SCD1 has emerged recently as a potential new target for the treatment for various diseases, such as nonalcoholic steatohepatitis, Alzheimer’s disease, cancer, and skin disorders. Sterculic acid (SA) is a cyclopropene fatty acid originally found in the seeds of the plant Sterculia foetida with numerous biological activities. On the one hand, its ability to inhibit stearoyl-CoA desaturase (SCD) allows its use as a coadjuvant of several pathologies where this enzyme has been associated. On the other hand, additional effects independently of its SCD inhibitory properties, involve anti-inflammatory and protective roles in retinal diseases such as age-related macular degeneration (AMD). This review aims to summarize the mechanisms by which SA exerts its actions and to highlight the emerging areas where this natural compound may be of help for the development of new therapies for human diseases.

Hkx2 (hexokinase 2) from Saccharomyces cerevisiae was one of the first metabolic enzymes described as a multifunctional protein. Hxk2 has a double subcellular localization: it functions as a glycolytic enzyme in the cytoplasm and as a regulator of gene transcription of several Mig1-regulated genes in the nucleus. To get more insights into the structure–function relationships of the Hxk2 protein, we followed two different approaches. In the first, we deleted the last eight amino acids of Hxk2 and replaced Ser304 with phenylalanine to generate Hxk2wca. Analysis of this mutant demonstrated that these domains play an essential role in the catalytic activity of yeast Hxk2, but has no effect on the regulatory function of this protein. In the second, we analysed whether amino acids from Lys6 to Met15 of Hxk2 (Hxk2wrf) are essential for the regulatory role of Hxk2 and whether there is an effect on the hexose kinase activity of this protein. In the present paper, we report that the Hxk2wca mutant protein interacts with the Mig1 transcriptional repressor and the Snf1 protein kinase in the nucleus at the level of the SUC2–Mig1 repressor complex. We have demonstrated that Hxk2wca maintained full regulatory function because the glucose-repression signalling of the wild-type machinery is maintained. We also report that the Hxk2wrf mutant allele is incapable of glucose repression signalling because it does not interact with Mig1 at the level of the SUC2–Mig1 repressor complex. The two mutants, Hxk2wca and Hxk2wrf retain single functions, as a transcriptional factor or as an enzyme with hexose-phosphorylating activity, but have lost the original bifunctionality of Hxk2.