Neurite extension is regulated by multiple signaling cascades that ultimately converge on the actin and microtubule networks [1]. Rho GTPases, molecular switches that oscillate between an inactive, GDP-bound state and an active, GTP-bound state, play a pivotal role in controlling actin cytoskeleton dynamics in the growth cone, whereas the dynamic behavior and interactions of microtubules are largely regulated by proteins called plus-end-tracking proteins (+TIPs), which associate with the ends of growing microtubules. Here, we show that the +TIP Navigator 1 (NAV1) is important for neurite outgrowth and interacts and colocalizes with TRIO, a Rho guanine nucleotide exchange factor that enables neurite outgrowth by activating the Rho GTPases Rac1 and RhoG. We find that binding of NAV1 enhances the affinity of TRIO for Rac1 and RhoG, and that NAV1 regulates TRIO-mediated Rac1 activation and neurite outgrowth. TRIO is also a +TIP, as it interacts with the core +TIP EB1 and tracks microtubule plus ends via EB1 and NAV1. Strikingly, the EB1-mediated recruitment of TRIO to microtubule ends is required for proper neurite outgrowth, and stabilization of the microtubule network by paclitaxel affects both the TRIO-NAV1 interaction and the accumulation of these proteins in neurite extensions. We propose that EB1-labeled ends of dynamic microtubules facilitate the formation and localization of functional NAV1-TRIO complexes, which in turn regulate neurite outgrowth by selectively activating Rac1. Our data reveal a novel link between dynamic microtubules, actin cytoskeleton remodeling, and neurite extension.

Osteoarthritis is a degenerative joint disease that ranks among the leading causes of adult disability. Mechanisms underlying osteoarthritis pathogenesis are not yet fully elucidated, putting limits to current disease management and treatment. Based on the phenomenological evidence for dysregulation within the glycome of chondrocytes and the network of a family of adhesion/growth-regulatory lectins, that is, galectins, we tested the hypothesis that Galectin-1 is relevant for causing degeneration. Immunohistochemical analysis substantiated that Galectin-1 upregulation is associated with osteoarthritic cartilage and subchondral bone histopathology and severity of degeneration (p < 0.0001, n = 29 patients). In vitro, the lectin was secreted and it bound to osteoarthritic chondrocytes inhibitable by cognate sugar. Glycan-dependent Galectin-1 binding induced a set of disease markers, including matrix metalloproteinases and activated NF-κB, hereby switching on an inflammatory gene signature (p < 10−16). Inhibition of distinct components of the NF-κB pathway using dedicated inhibitors led to dose-dependent impairment of Galectin-1–mediated transcriptional activation. Enhanced secretion of effectors of degeneration such as three matrix metalloproteinases underscores the data’s pathophysiological relevance. This study thus identifies Galectin-1 as a master regulator of clinically relevant inflammatory-response genes, working via NF-κB. Because inflammation is critical to cartilage degeneration in osteoarthritis, this report reveals an intimate relation of glycobiology to osteoarthritic cartilage degeneration.

Chronic lung hypoxia causes vascular remodeling with pulmonary artery smooth muscle cell (SMCPA) hyperplasia, resulting in pulmonary hypertension and cor pulmonale. We investigated SMCPA and pulmonary artery adventitial fibroblasts (FBPA) for their proliferative response to hypoxia. Strong SMCPA growth occurred under hypoxic conditions in SMCPA/FBPA co-cultures, but not in SMCPA monocultures. SMCPA growth was fully reproduced by transferring serum-free supernatant from hypoxic cultured FBPA to normoxic SMCPA. Hypoxia-inducible-transcription-factor subtypes (HIF-1alpha, HIF-2alpha, HIF-3alpha) and its dependent target genes, carrying the hypoxia-responsive-element as regulatory component, were strongly activated in both hypoxic FBPA and SMCPA. HIF-transcription-factor decoy technique, employed to FBPA during hypoxic culturing, blocked the mitogenic activity of FBPA conditioned medium on SMCPA. The data suggest that hypoxia-driven gene regulation in pulmonary artery fibroblasts results in a mitogenic stimulus on adjacent pulmonary artery smooth muscle cells, and HIF-transcription-decoy may offer a new therapeutic approach to suppress these events.

The mechanical strength of individual Si-C bonds was determined as a function of the applied force-loading rate by dynamic single-molecule force spectroscopy, using an atomic force microscope. The applied force-loading rates ranged from 0.5 to 267 nN/s, spanning 3 orders of magnitude. As predicted by Arrhenius kinetics models, a logarithmic increase of the bond rupture force with increasing force-loading rate was observed, with average rupture forces ranging from 1.1 nN for 0.5 nN/s to 1.8 nN for 267 nN/s. Three different theoretical models, all based on Arrhenius kinetics and analytic forms of the binding potential, were used to analyze the experimental data and to extract the parameters fmax and D(e) of the binding potential, together with the Arrhenius A-factor. All three models well reproduced the experimental data, including statistical scattering; nevertheless, the three free parameters allow so much flexibility that they cannot be extracted unambiguously from the experimental data. Successful fits with a Morse potential were achieved with fmax = 2.0-4.8 nN and D(e) = 76-87 kJ/mol, with the Arrhenius A-factor covering 2.45 x 10(-10)-3 x 10(-5) s(-1), respectively. The Morse potential parameters and A-factor taken from gas-phase density functional calculations, on the other hand, did not reproduce the experimental forces and force-loading rate dependence.