Purification of Cortactin Using Anion Exchange Chromatography

Cortactin is an actin-binding protein that has been shown to be involved in cellular migration and metastases in cancer. Bacterially expressed and purified cortactin protein is often used in in vitro assays to examine cortactin’s role in promoting cell migration via actin remodeling. Cortactin has a theoretical molecular weight of 60 kDa; however, using SDS-PAGE analysis, the protein runs as two bands of molecular weights 80 kDa and 85 kDa, suggesting that cortactin has an unusual protein folding pattern. Our current lack of understanding of cortactin structure limits our ability to determine the role of cortactin in facilitating motility phenotypes. To elucidate the forms of cortactin produced from bacterially expressed and purified cortactin protein, we used a two-step purification system including affinity purification and anion exchange chromatography. After analysis with non-denaturing polyacrylamide gel electrophoresis, we found cortactin protein from different anion chromatography elution fractions did not separate to similar locations on the gel across all fractions. We hypothesize that the variations in the bands are a result of different folding patterns of cortactin protein in what was once thought of as a homogenous protein pool. When testing the role of cortactin in mediating cell migration, these folding differences may have significant effects on the results of functional assays such as actin polymerization or sedimentation

Not a Simple Relationship between Neptune’s Migration Speed and Kuiper Belt Inclination Excitation

We present numerical simulations of giant planet migration in our solar system and examine how the speed of planetary migration affects inclinations in the resulting population of small bodies (test particles) scattered outward and subsequently captured into Neptune's 3:2 mean motion resonance (the Plutinos), as well as the hot classical Kuiper Belt population. We do not find a consistent relationship between the degree of test particle inclination excitation and e-folding planet migration timescales in the range 5-50 Myr. Our results present a counterexample to Nesvorny's finding that the Plutino and hot classical inclinations showed a marked increase with increasing e-folding timescales for Neptune's migration. We argue that these differing results are likely due to differing secular architectures of the giant planets during and after migration. Small changes in the planets' initial conditions and differences in the numerical implementation of planet migration can result in different amplitudes of the planets' inclination secular modes, and this can lead to different final inclination distributions for test particles in the simulations. We conclude that the observed large inclination dispersion of Kuiper Belt objects does not require Neptune's migration to be slow; planetary migration with e-folding timescales of 5, 10, 30, and 50 Myr can all yield inclination dispersions similar to the observed Plutino and hot classical populations, with no correlation between the degree of inclination excitation and migration speed.NASA [NNX14AG93G, 80NSSC19K0785]; NSF [AST-1312498, AST-1824869]This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Molecular mechanisms of neural crest cell attachment and migration on types I and IV collagen

We have examined the mechanisms involved in the interaction of avian neural crest cells with collagen types I and IV (Col I and IV) during their adhesion and migration in vitro. For this purpose native Col IV was purified from chicken tissues, characterized biochemically and ultrastructurally. Purified chicken Col I and Col IV, and various proteolytic fragments of the collagens, were used in quantitative cell attachment and migration assays in conjunction with domain-specific collagen antibodies and antibodies to avian integrin subunits. Neural crest cells do not distinguish between different macromolecular arrangements of Col I during their initial attachment, but do so during their migration, showing a clear preference for polymeric Col I. Interaction with Col I is mediated by the α1β1 integrin, through binding to a segment of the α1(I) chain composed of fragment CNBr3. Neural crest cell attachment and migration on Col IV involves recognition of conformation-dependent sites within the triple-helical region and the noncollagenous, carboxyl-terminal NC1 domain. This recognition requires integrity of inter- and intrachain disulfide linkages and correct folding of the molecule. Moreover, there also is evidence that interaction sites within the NC1 domain may be cryptic, being exposed during migration of the cells in the intact collagen as a result of the prolonged cell-substratum contact. In contrast to Col I, neural crest cell interaction with Col IV is mediated by β1-class integrins other than α1β1

Cap rock efficiency of geothermal systems in fold-and-thrust belts: Evidence from paleo-thermal and structural analyses in Rosario de La Frontera geothermal area (NW Argentina)

Cap rock characterization of geothermal systems is often neglected despite fracturing may reduce its efficiency and favours fluid migration. We investigated the siliciclastic cap rock of Rosario de La Frontera geothermal system (NW Argentina) in order to assess its quality as a function of fracture patterns and related thermal alteration. Paleothermal investigations (XRD on fine-grained fraction of sediments, organic matter optical analysis and fluid inclusions on veins) and 1D thermal modelling allowed us to distinguish the thermal fingerprint associated to sedimentary burial from that related to fluid migration. The geothermal system is hosted in a Neogene N-S anticline dissected by high angle NNW- and ENE-striking faults. Its cap rock can be grouped into two quality categories: • rocks acting as good insulators, deformed by NNW–SSE and E–W shear fractures, NNE-SSW gypsum- and N-S-striking calcite-filled veins that developed during the initial stage of anticline growth. Maximum paleo-temperatures (< 60 °C) were experienced during deposition to folding phases.• rocks acting as bad insulators, deformed by NNW-SSE fault planes and NNW- and WNW-striking sets of fractures associated to late transpressive kinematics. Maximum paleo-temperatures higher than about 115 °C are linked to fluid migration from the reservoir to surface (with a reservoir top at maximum depths of 2.5 km) along fault damage zones.This multi-method approach turned out to be particularly useful to trace the main pathways of hot fluids and can be applied in blind geothermal systems where either subsurface data are scarce or surface thermal anomalies are lacking.Fil: Maffucci, R.. Universita Degli Studi Della Tuscia; Italia. Universita Degli Studi Roma Tre; ItaliaFil: Corrado, Sveva. Universita Degli Studi Roma Tre; ItaliaFil: Aldega, L.. Instituto de Investigaciones Universitarias Roma la Sapienza; ItaliaFil: Bigi, S.. Instituto de Investigaciones Universitarias Roma la Sapienza; ItaliaFil: Chiodi, Agostina Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Investigaciones en Energía no Convencional. Universidad Nacional de Salta. Facultad de Ciencias Exactas. Departamento de Física. Instituto de Investigaciones en Energía no Convencional; ArgentinaFil: Di Paolo, L.. Eni E&P Division; ItaliaFil: Giordano, G.. Universita Degli Studi Roma Tre; ItaliaFil: Invernizzi, C.. Universita Degli Di Camerino; Itali

Myelin pathology: Involvement of molecular chaperones and the promise of chaperonotherapy

The process of axon myelination involves various proteins including molecular chaperones. Myelin alteration is a common feature in neurological diseases due to structural and functional abnormalities of one or more myelin proteins. Genetic proteinopathies may occur either in the presence of a normal chaperoning system, which is unable to assist the defective myelin protein in its folding and migration, or due to mutations in chaperone genes, leading to functional defects in assisting myelin maturation/migration. The latter are a subgroup of genetic chaperonopathies causing demyelination. In this brief review, we describe some paradigmatic examples pertaining to the chaperonins Hsp60 (HSPD1, or HSP60, or Cpn60) and CCT (chaperonin-containing TCP-1). Our aim is to make scientists and physicians aware of the possibility and advantages of classifying patients depending on the presence or absence of a chaperonopathy. In turn, this subclassification will allow the development of novel therapeutic strategies (chaperonotherapy) by using molecular chaperones as agents or targets for treatment

On the evolution of the resonant planetary system HD128311

A significant number of the known multiple exoplanetary systems are containing a pair of giant planets engaged in a low order mean motion resonance. Such a resonant condition protects the dynamics of these planets resulting in very stable orbits. According to recent studies the capture into a resonance is the result of a planetary migration process induced by the interaction of the planets with a protoplanetary disk. If the migration is slow enough (adiabatic) next to a mean motion resonance, the two planets will also be in apsidal corotation. The recently refined orbital parameters of the system HD 128311 suggest that the two giant planets are in a 2:1 mean motion resonance, however without exhibiting apsidal corotation. Thus the evolution of this system can not be described by an adiabatic migration process alone. We present possible evolution scenarios of this system combining migration processes and sudden perturbations. We model migration scenarios through numerical integration of the gravitational N-body problem with additional non-conservative forces. Planet-planet scattering has been investigated by N-body simulations. We show that the present dynamical state of the system HD128311 may be explained by such evolutionary processes.Comment: 4 Pages, 7 Figures, accepted for AA Letter