Fiber type conversion alters inactivation of voltage-dependent sodium currents in murine C2C12 skeletal muscle cells

Each skeletal muscle of the body contains a unique composition of "fast" and "slow" muscle fibers, each of which is specialized for certain challenges. This composition is not static, and the muscle fibers are capable of adapting their molecular composition by altered gene expression (i.e., fiber type conversion). Whereas changes in the expression of contractile proteins and metabolic enzymes in the course of fiber type conversion are well described, little is known about possible adaptations in the electrophysiological properties of skeletal muscle cells. Such adaptations may involve changes in the expression and/or function of ion channels. In this study, we investigated the effects of fast-to-slow fiber type conversion on currents via voltage-gated Na⁺ channels in the C₂C₁₂ murine skeletal muscle cell line. Prolonged treatment of cells with 25 nM of the Ca²⁺ ionophore A-23187 caused a significant shift in myosin heavy chain isoform expression from the fast toward the slow isoform, indicating fast-to-slow fiber type conversion. Moreover, Na⁺ current inactivation was significantly altered. Slow inactivation less strongly inhibited the Na⁺ currents of fast-to-slow fiber type-converted cells. Compared with control cells, the Na⁺ currents of converted cells were more resistant to block by tetrodotoxin, suggesting enhanced relative expression of the cardiac Na⁺ channel isoform Na_v1.5 compared with the skeletal muscle isoform Na_v1.4. These results imply that fast-to-slow fiber type conversion of skeletal muscle cells involves functional adaptation of their electrophysiological properties. muscle plasticity; myosin heavy chain expression; sodium channel expression     

Intermediate filaments mediate cytoskeletal crosstalk

Intermediate filaments, actin-containing microfilaments and microtubules are the three main cytoskeletal systems of vertebrate and many invertebrate cells. Although these systems are composed of distinctly different proteins, they are in constant and intimate communication with one another. Understanding the molecular basis of this cytoskeletal crosstalk is essential for determining the mechanisms that underlie many cell-biological phenomena. Recent studies have revealed that intermediate filaments and their associated proteins are important components in mediating this crosstalk.     

Solvent specified conformation in poly(alpha-L-glutamic acid) thin films

The ability to control conformational properties of polypeptides in their films is of considerable interest for many possible applications of these materials. By rational choice of the solvent system for film fabrication, control over the conformation of the main chain, the intermolecular hydrogen bonding in the side chain is easily achieved in poly(alpha-L-glutamic acid) (PLGA) thin films. The spectral data from circular dichromism (CD), FT-IR, and solid state (13)C NMR spectroscopies suggest that the beta-sheet conformation is dominant in PLGA films cast from trifluoroacetic acid (TFA) solution, whereas the right-handed alpha-helix is dominant in those cast from pyridine or DMF solution. In comparison with films cast from TFA solutions, the films fabricated from pyridine or DMF solutions exhibit strong intermolecular hydrogen bondings between -COOH groups and have a more ordered arrangement of side chains. Moreover, the extent of alpha-helix conformation of the PLGA backbone in films cast from pyridine or DMF solution is several times higher than that observed in the PLGA powder precipitated from aqueous solution at pH 4. All spectroscopic studies indicate clearly that the solvents (used for casting these films) play a crucial role in directing the organization of PLGA in these thin films.     

Identification and sequence analysis of six new members of the NIMA-related kinase family in Chlamydomonas

The NIMA kinases are an evolutionarily conserved protein family with enigmatic roles in the regulation of mitosis. We report six new members of this family in Chlamydomonas, in addition to the previously identified NIMA-related kinase, Fa2p. Chlamydomonas NIMA-related kinases (CNKs) 1-6 were sequenced from subclones generated by RT-PCR using information from EST libraries and the recently sequenced Chlamydomonas genome. Phylogenetic and bioinformatic approaches were used to determine the relationships of the six new members with known members of the NIMA-related kinase family. Although humans express at least eleven NIMA-related kinases, the eukaryotic microbes that have been studied to date express only one or two members of the family. Thus, the discovery that Chlamydomonas expresses a total of at least seven NIMA-related kinases is intriguing. Our analyses suggest that members of this family may play roles in the assembly and function of cilia.     

Rab11-family interacting protein 2 and myosin Vb are required for CXCR2 recycling and receptor-mediated chemotaxis

Agonist-stimulated internalization followed by recycling to the cell membrane play an important role in fine-tuning the activity of chemokine receptors. Because the recycling of chemokine receptors is critical for the reestablishment of the cellular responsiveness to ligand, it is crucial to understand the mechanisms underlying the receptor recycling and resensitization. In the present study, we have demonstrated that the chemokine receptor CXCR2 associated with myosin Vb and Rab11-family interacting protein 2 (FIP2) in a ligand-dependent manner. Truncation of the C-terminal domain of the receptor did not affect the association, suggesting that the interactions occur upstream of the C terminus of CXCR2. After ligand stimulation, the internalized CXCR2 colocalized with myosin Vb and Rab11-FIP2 in Rab11a-positive vesicles. The colocalization lasted for approximately 2 h, and little colocalization was observed after 4 h of ligand stimulation. CXCR2 also colocalized with myosin Vb tail or Rab11-FIP2 (129-512), the N-terminal-truncated mutants of myosin Vb and Rab11-FIP2, respectively, but in a highly condensed manner. Expression of the enhanced green fluorescent protein-tagged myosin Vb tail significantly retarded the recycling and resensitization of CXCR2. CXCR2 recycling was also reduced by the expression Rab11-FIP2 (129-512). Moreover, expression of the myosin Vb tail reduced CXCR2- and CXCR4-mediated chemotaxis. These data indicate that Rab11-FIP2 and myosin Vb regulate CXCR2 recycling and receptor-mediated chemotaxis and that passage of internalized CXCR2 through Rab11a-positive recycling system is critical for physiological response to a chemokine.     

Evolution and biogeography of marine angelfishes (Pisces: Pomacanthidae)

Phylogenetic relationships among angelfishes (Pomacanthidae) and their putative sister taxon, the butterflyfishes (Chaetodontidae), were examined using 12S and 16S mitochondrial DNA sequences. ML and MP trees were highly congruent with good basal resolution. Monophyly of the two families was supported, although a clade comprising the Chaetodontidae and one of the outgroups, the Scatophagidae, formed the sister clade to the Pomacanthidae. All genera and subgenera within the Pomacanthidae were examined. The relationships among the 24 representative species were consistent with traditional generic boundaries, with the exception of the genus Centropyge, but differed from previous phylogenies. Estimated ages of divergence based on trans-isthmian pairs were compared with independent fossil evidence. Trans-isthmian estimates were highly conservative, while fossil-calibrated estimates were most consistent with available evidence. Fossil calibrated estimates suggest that the family has been impacted by both the Terminal Tethyan Event and the closure of the Isthmus of Panama. Within the family, ecological diversity and species-level diversification are restricted primarily to a single pygmy angelfish clade with an origin near the Oligocene-Miocene boundary.     

Alpha 1-adrenergic receptor responses in alpha 1AB-AR knockout mouse hearts suggest the presence of alpha 1D-AR

Two functional alpha(1)-adrenergic receptor (AR) subtypes (alpha(1A) and alpha(1B)) have been identified in the mouse heart. However, it is unclear whether the third known subtype, alpha(1D)-AR, is also present. To investigate this, we determined whether there were alpha(1)-AR responses in hearts from a novel mouse model lacking alpha(1A)- and alpha(1B)-ARs (double knockout) (ABKO). In Langendorff-perfused hearts, alpha(1)-ARs were stimulated with phenylephrine. For ABKO hearts, phenylephrine reduced left ventricular pressure and coronary flow (to 87 +/- 2% and 86 +/- 4% of initial, respectively, n = 11, P < 0.01). These effects were blocked by prazosin and 8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]-8-azaspirol[4,5]decane-7,9-dione] dihydrochloride, suggesting that alpha(1D)-AR-mediated responses were present. In contrast, right ventricular trabeculae from ABKO hearts did not respond to phenylephrine, suggesting that in ABKO perfused hearts, the effects of phenylephrine were not mediated by direct actions on cardiomyocytes. A novel finding was that alpha(1)-AR stimulation caused positive inotropy in the wild-type mouse heart, in contrast to negative inotropy observed in mouse cardiac muscle strips. We conclude that mouse hearts lacking alpha(1A)- and alpha(1B)-ARs retain functional alpha(1)-AR responses involving decreases of coronary flow and ventricular pressure that reflect alpha(1D)-AR-mediated vasoconstriction. Furthermore, alpha(1)-AR inotropic responses depend critically on the experimental conditions.