Function of caveolae in Ca2+ entry and Ca2+-dependent signal transduction

The correct spatial and temporal control of Ca²⁺ signaling is essential for such cellular activities as fertilization, secretion, motility, and cell division. There has been a long-standing interest in the role of caveolae in regulating intracellular Ca²⁺ concentration. In this review we provide an updated view of how caveolae may regulate both Ca²⁺ entry into cells and Ca²⁺-dependent signal transduction     

Activation of focal adhesion kinase in human lung cancer cells involves multiple and potentially parallel signaling events

Integrins are adhesion receptors that transmit signals bidirectionally across the plasma membrane. In our previous report we have shown that the squamous lung cancer cell line, Calu-1, binds to collagen type IV (Coll IV) through beta1-integrin and results in phosphorylation of focal adhesion kinase (FAK) (Ann Thorac Surg 2004; 78:450-457). Considering the critical role of FAK in cell migration, proliferation, and survival, here we investigated potential mechanisms of its activation and regulation in Calu-1 cells. We observed the phosphorylation of Tyr397 of FAK (the autophosphorylation site of FAK) and paxillin, the immediate downstream substrate of FAK following the adhesion of Calu-1 cells to Coll IV. FAK remains phosphorylated during proliferation either on Coll IV or on uncoated plates for 72 h, as determined by peroxivanadate treatment. Exposure of Calu-1 cells with 60 microM genistein, reduces FAK phosphorylation (7.6 fold) and cell proliferation. Extracellular signal regulated kinases (ERKs) were also phosphorylated after Coll IV attachment. Disruption of Calu-1 cell cytoskeleton integrity by 1-5 muM Cytochalasin D resulted in the inhibition of cell adhesion (50% to 75%, p<0.19 - 6.6 x 10(7)) and ERKs phosphorylation (2 fold) without any effect on FAK phosphorylation. Protein Kinase C inhibitor, Calphostin C at 100 and 250 nM concentrations did not block Coll IV induced FAK phosphorylation but activated the ERKs in a dose dependent manner. beta1-integrin is essential for Coll IV induced FAK activation, but it is not physically associated with FAK as determined by immunodetection assay. Collectively, this report defines the existence of multiple and potentially parallel Coll IV/beta1-integrin mediated signaling events in Calu-1 cells, which involve FAK, ERKs, and PKC.     

Comparative bioacoustics: a roadmap for quantifying and comparing animal sounds across diverse taxa

Animals produce a wide array of sounds with highly variable acoustic structures. It is possible to understand the causes and consequences of this variation across taxa with phylogenetic comparative analyses. Acoustic and evolutionary analyses are rapidly increasing in sophistication such that choosing appropriate acoustic and evolutionary approaches is increasingly difficult. However, the correct choice of analysis can have profound effects on output and evolutionary inferences. Here, we identify and address some of the challenges for this growing field by providing a roadmap for quantifying and comparing sound in a phylogenetic context for researchers with a broad range of scientific backgrounds. Sound, as a continuous, multidimensional trait can be particularly challenging to measure because it can be hard to identify variables that can be compared across taxa and it is also no small feat to process and analyse the resulting high-dimensional acoustic data using approaches that are appropriate for subsequent evolutionary analysis. Additionally, terminological inconsistencies and the role of learning in the development of acoustic traits need to be considered. Phylogenetic comparative analyses also have their own sets of caveats to consider. We provide a set of recommendations for delimiting acoustic signals into discrete, comparable acoustic units. We also present a three-stage workflow for extracting relevant acoustic data, including options for multivariate analyses and dimensionality reduction that is compatible with phylogenetic comparative analysis. We then summarize available phylogenetic comparative approaches and how they have been used in comparative bioacoustics, and address the limitations of comparative analyses with behavioural data. Lastly, we recommend how to apply these methods to acoustic data across a range of study systems. In this way, we provide an integrated framework to aid in quantitative analysis of cross-taxa variation in animal sounds for comparative phylogenetic analysis. In addition, we advocate the standardization of acoustic terminology across disciplines and taxa, adoption of automated methods for acoustic feature extraction, and establishment of strong data archival practices for acoustic recordings and data analyses. Combining such practices with our proposed workflow will greatly advance the reproducibility, biological interpretation, and longevity of comparative bioacoustic studies.     

Association Between the Rat Homologue of CO-029, a Metastasis-associated Tetraspanin Molecule and Consumption Coagulopathy

Recently, we have described a panel of metastasis-associated antigens in the rat, i.e., of molecules expressed on metastasizing, but not on nonmetastasizing tumor lines. One of these molecules, recognized by the monoclonal antibody D6.1 and named accordingly D6.1A, was found to be abundantly expressed predominantly on mesenchyme-derived cells. The DNA of the antigen has been isolated and cloned. Surprisingly, the gene product proved to interfere strongly with coagulation.

The 1.182-kb cDNA codes for a 235–amino acid long molecule with a 74.2% homology in the nucleotide and a 70% homology in the amino acid sequence to CO-029, a human tumor-associated molecule. According to the distribution of hydrophobic and hydrophilic amino acids, D6.1A belongs to the tetraspanin superfamily. Western blotting of D6.1A-positive metastasizing tumor lines revealed that the D6.1A, like many tetraspanin molecules, is linked to further membrane molecules, one of which could be identified as α6β1 integrin. Transfection of a low-metastasizing tumor cell line with D6.1A cDNA resulted in increased metastatic potential and provided a clue as to the functional role of D6.1A. We noted massive bleeding around the metastases and, possibly as a consequence, local infarctions predominantly in the mesenteric region and all signs of a consumption coagulopathy. By application of the D6.1 antibody the coagulopathy was counterregulated, though not prevented.

It has been known for many years that tumor growth and progression is frequently accompanied by thrombotic disorders. Our data suggest that the phenomenon could well be associated with the expression of tetraspanin molecules.

     

Water in the orchestration of the cell machinery. Some misunderstandings: a short review

Nowadays, biologists can explore the cell at the nanometre level. They discover an unsuspected world, amazingly overcrowded, complex and heterogeneous, in which water, also, is complex and heterogeneous. In the cell, statistical phenomena, such as diffusion, long considered as the main transport for water soluble substances, must be henceforth considered as inoperative to orchestrate cell activity. Results at this level are not yet numerous enough to give an exact representation of the cell machinery; however, they are sufficient to cease reasoning in terms of statistics (diffusion, law of mass action, pH, etc.) and encourage cytologists and biochemists to prospect thoroughly the huge panoply of the biophysical properties of macromolecule-water associations at the nanometre level. Our main purpose, here, is to discuss some of the more common misinterpretations due to the ignorance of these properties, and expose briefly the bases for a better approach of the cell machinery. Giorgio Careri, who demonstrated the correlation between proton currents at the surface of lysozyme and activity of this enzyme was one of the pioneers of this approach.     

Comparative aspects of polyglutamine binding domain in PQBP-1 among Vertebrata

We investigated the evolutionary conservation of polyglutamine binding protein-1 (PQBP-1) among Vertebrata. PQBP-1s were highly conserved and shared the same domain features including a WW domain, a polar amino acid rich domain (PRD), a nuclear localization signal (NLS), and a C-terminal domain (CTD) among Eutheria, but not always among Vertebrata. PQBP-1s of Vertebrata contained a variable region in the middle portion corresponding to the position of PRD. The full form of PRD including both 7aa and DR/ER repeats was specific to Eutheria. PRD of non-eutherian Amniota was minimal. Amphibia had no PRD. The DR/ER repeat was solo in fishes. Agnatha PRD was also rich in polar amino acids, but contained no repetitive sequence. We investigated 3 polyQ-containing proteins known to interact with PQBP-1: BRN-2, Huntingtin, and ATAXIN-1, and showed a diverse nature of protein-protein interaction in Vertebrata. There appears to be no interaction between PQBP-1 and BRN-2, Huntingtin, or ATAXIN-1 in Amphibia, while the interaction between PQBP-1 and BRN-2 is expected to be conserved among Mammalia, and the interaction between PQBP-1 and Huntingtin or ATAXIN-1 depends on the lineage in Eutheria.