Is Lamprey Metamorphosis Regulated by Thyroid Hormones?

Lampreys are one of just a few fishes which have a true (firstor first type) of metamorphosis in their life cycle. In thesea lamprey (Petromyzon marinus), spontaneous metamorphosisis initiated when the size (length and weight), condition factor,and lipid stores reach appropriate levels and coincide withthe postwinter rise in water temperature. The serum levels ofthe thyroid hormones, thyroxine (T₄) and triiodothyronine (T₃),drop dramatically at the onset of metamorphosis and metamorphosiscan be induced with treatment of animals with the goitrogen,KCIO₄, which also results in a decline in serum levels of thyroidhormones. The fact that thyroid hormone treatment can blockspontaneous and induced metamorphosis is support for the viewthat thyroid hormones, particularly T₃, operates like a juvenilehormone in lamprey metamorphosis; this view is counter to therole of thyroid hormones in metamorphosis of other vertebrates.The monodeiodinase pathways, whereby T₄ is converted to T₃ orto the biologically inactive reverse T₃, and even further degradationof T₃, may be a significant mechanism directing metamorphicchange. Lamprey metamorphosis is facultative in that it is initiatedor inhibited depending upon the coordination of a complex integrationof environmental, metabolic and hormonal cues. Thyroid hormonesdo not regulate lamprey metamorphosis in the sense observedin other vertebrate metamorphoses but they are important tothe developmental process. Some of the features of the involvementof thyroid hormones in lamprey metamorphosis may be relatedto the presence of the endostyle in larvae which in turn reflectsthe ancient origins of this vertebrate and perhaps the conservationof an ancient method of induction of metamorphosis. Some cluefor other factors which initiate lamprey metamorphosis may comethrough the examination of inducers of metamorphosis in lowerchordates     

Is Peripheral Immunity Regulated by Blood-Brain Barrier Permeability Changes?

S100B is a reporter of blood-brain barrier (BBB) integrity which appears in blood when the BBB is breached. Circulating S100B derives from either extracranial sources or release into circulation by normal fluctuations in BBB integrity or pathologic BBB disruption (BBBD). Elevated S100B matches the clinical presence of indices of BBBD (gadolinium enhancement or albumin coefficient). After repeated sub-concussive episodes, serum S100B triggers an antigen-driven production of anti-S100B autoantibodies. We tested the hypothesis that the presence of S100B in extracranial tissue is due to peripheral cellular uptake of serum S100B by antigen presenting cells, which may induce the production of auto antibodies against S100B. To test this hypothesis, we used animal models of seizures, enrolled patients undergoing repeated BBBD, and collected serum samples from epileptic patients. We employed a broad array of techniques, including immunohistochemistry, RNA analysis, tracer injection and serum analysis. mRNA for S100B was segregated to barrier organs (testis, kidney and brain) but S100B protein was detected in immunocompetent cells in spleen, thymus and lymph nodes, in resident immune cells (Langerhans, satellite cells in heart muscle, etc.) and BBB endothelium. Uptake of labeled S100B by rat spleen CD4+ or CD8+ and CD86+ dendritic cells was exacerbated by pilocarpine-induced status epilepticus which is accompanied by BBBD. Clinical seizures were preceded by a surge of serum S100B. In patients undergoing repeated therapeutic BBBD, an autoimmune response against S100B was measured. In addition to its role in the central nervous system and its diagnostic value as a BBBD reporter, S100B may integrate blood-brain barrier disruption to the control of systemic immunity by a mechanism involving the activation of immune cells. We propose a scenario where extravasated S100B may trigger a pathologic autoimmune reaction linking systemic and CNS immune responses.     

How Does Auxin Enhance Cell Elongation? Roles of Auxin‐Binding Proteins and Potassium Channels in Growth Control

Elongation growth and a several other phenomena in plant development are controlled by the plant hormone auxin. A number of recent discoveries shed light on one of the classical problems of plant physiology: the perception of the auxin signal. Two types of auxin receptors are currently known: the AFB/TIR family of F box proteins and ABP1. ABP1 appears to control membrane transport processes (H+ secretion, osmotic adjustment) while the TIR/AFBs have a role in auxin-induced gene expression. Models are proposed to explain how membrane transport (e.g., K+ and H+ fluxes) can act as a cross-linker for the control of more complex auxin responses such as the classical stimulation of cell elongation.     

Does Auxin Binding Protein 1 Control Both Cell Division and Cell Expansion?

The Auxin-Binding Protein 1 (ABP1) was identified over 30 years ago thanks to it''s high affinity for active auxins. ABP1 plays an essential role in plant life yet to this day, its function remains ‘enigmatic.’ A recent study by our laboratory shows that ABP1 is critical for regulation of the cell cycle, acting both in G₁ and at the G₂/M transition. We showed that ABP1 is likely to mediate the permissive auxin signal for entry into the cell cycle. These data were obtained by studying a conditional functional knock-out of ABP1 generated by cellular immunization in the model tobacco cell line, Bright Yellow 2.Key Words: auxin responses, auxin-binding protein 1, immunomodulation, cellular immunisation     

Matrix-dependent Tiam1/Rac Signaling in Epithelial Cells Promotes Either Cell–Cell Adhesion or Cell Migration and Is Regulated by Phosphatidylinositol 3-Kinase

We previously demonstrated that both Tiam1, an activator of Rac, and constitutively active V12Rac promote E-cadherin–mediated cell–cell adhesion in epithelial Madin Darby canine kidney (MDCK) cells. Moreover, Tiam1 and V12Rac inhibit invasion of Ras-transformed, fibroblastoid MDCK-f3 cells by restoring E-cadherin–mediated cell–cell adhesion. Here we show that the Tiam1/Rac-induced cellular response is dependent on the cell substrate. On fibronectin and laminin 1, Tiam1/Rac signaling inhibits migration of MDCK-f3 cells by restoring E-cadherin–mediated cell– cell adhesion. On different collagens, however, expression of Tiam1 and V12Rac promotes motile behavior, under conditions that prevent formation of E-cadherin adhesions. In nonmotile cells, Tiam1 is present in adherens junctions, whereas Tiam1 localizes to lamellae of migrating cells. The level of Rac activation by Tiam1, as determined by binding to a glutathione-S-transferase– PAK protein, is similar on fibronectin or collagen I, suggesting that rather the localization of the Tiam1/Rac signaling complex determines the substrate-dependent cellular responses. Rac activation by Tiam1 requires PI3-kinase activity. Moreover, Tiam1- but not V12Rac-induced migration as well as E-cadherin–mediated cell– cell adhesion are dependent on PI3-kinase, indicating that PI3-kinase acts upstream of Tiam1 and Rac.     

Is Cell Size Important?

Cell size plays an indirect role in cell proliferation, as cells must double in size before dividing. Cell size is largely determined by the activity of RNA polymerase I that controls ribosomal RNA synthesis and ribosome biogenesis. The type 1 insulin-like growth factor receptor (IGF-IR) and its docking protein, insulin receptor substrate-1 (IRS-1) control, in a non-redundant way, about 50% of cell and body size. This is certainly true in mice, flies and cells in culture, but also probably in higher mammals. Interestingly, the insulin receptor (InR) cannot substitute for the IGF-IR in controlling cell size. This is probably due to the fact that the IGF-IR is more effective than the InR in translocationg to the nuclei IRS-1, which then binds UBF1, one of the proteins that regulate RNA pol I activity.     

Is Cell Rheology Governed by Nonequilibrium-to-Equilibrium Transition of Noncovalent Bonds?

A living cell deforms or flows in response to mechanical stresses. A recent report shows that dynamic mechanics of living cells depends on the timescale of mechanical loading, in contrast to the prevailing view of some authors that cell rheology is timescale-free. Yet the molecular basis that governs this timescale-dependent behavior is elusive. Using molecular dynamics simulations of protein-protein noncovalent interactions, we show that multipower laws originate from a nonequilibrium-to-equilibrium transition: when the loading rate is faster than the transition rate, the power-law exponent α₁ is weak; when the loading rate is slower than the transition rate, the exponent α₂ is strong. The model predictions are confirmed in both embryonic stem cells and differentiated cells. Embryonic stem cells are less stiff, more fluidlike, and exhibit greater α₁ than their differentiated counterparts. By introducing a near-equilibrium frequency f_eq, we show that all data collapse into two power laws separated by f/f_eq, which is unity. These findings suggest that the timescale-dependent rheology in living cells originates from the nonequilibrium-to-equilibrium transition of the dynamic response of distinct, force-driven molecular processes.     

Inhibition of Cell Expansion by Rapid ABP1-Mediated Auxin Effect on Microtubules? A Critical Comment

Critical analysis of a recent article raises questions regarding the inhibition of cell expansion by rapid ABP1-mediated auxin effect on microtubules.How do cortical microtubules shape plant cells? This has been an important question ever since the microtubular cytoskeleton was found to orientate the deposition of cellulose microfibrils in the primary cell wall and control long-term anisotropic cell expansion under isotropic turgor pressure (Green, 1962). In axial plant organs, longitudinal microtubule/microfibril arrays hamper expansion in length and favor expansion in girth, while transverse microtubule/microfibril arrays have the opposite effect (Baskin, 2001). By generating mechanical anisotropy in the cell wall, microtubule orientation controls the ratio of longitudinal versus circumferential cell expansion (the allometric ratio). A recent study by Chen et al. (2014) concludes that auxin inhibits cell growth by causing a rapid reorientation of microtubules from a transverse to a longitudinal orientation in cells of the submeristematic root zone and the elongation zone of the hypocotyl of Arabidopsis (Arabidopsis thaliana) seedlings. This conclusion warrants attention because the cell expansion that drives auxin-mediated organ elongation is generally thought to be controlled by the regulated breaking of bonds between existing cell wall polymers by chemical means (wall loosening; Cosgrove, 2005).It has been clear for more than 25 years that auxin does induce rapid changes in the orientation of microtubules in growing cells, either during straight growth (Bergfeld et al., 1988) or tropic bending (Nick et al., 1990). These studies form part of a group of investigations that show that numerous growth-affecting endogenous, environmental, and even artificial physical factors have very similar effects on microtubule orientation: during active cell expansion or related mechanical strains, microtubules are aligned against the direction of expansion, and they are aligned with it during the inhibition of expansion (Fischer and Schopfer, 1997). An important insight that emerges from this extensive evidence is that the type of reorientation elicited by a particular factor depends on its physiological context, thereby allowing auxin to induce either transverse or longitudinal microtubule orientations depending on whether elongation growth is promoted (as in shoot organs) or inhibited (as in roots). Clearly, ordered microtubule reorientations require the input of directional information (Williamson, 1990). Auxin signaling as such cannot provide this information, but the directional growth responses produced by auxin can. This brings us to the crucial question: is the orientation of microtubules determined by the effector signal directly or by changes in growth? The answer given by Chen et al. (2014) comes as a surprise: the inhibition of cell expansion is mediated by the rapid AUXIN-BINDING PROTEIN1 (ABP1)-dependent action of auxin on microtubules. The authors imply that it is microtubular reorientation per se that is responsible for the sudden growth inhibition caused by auxin in roots and hypocotyls rather than any changes in cell wall structure. Related microtubule reorientations at the concave side of gravitropically curving roots are interpreted in a similar way.Coming as an even bigger surprise, Chen and collaborators do not provide any evidence to support the claim made in their title, nor do they touch the obvious question of how microtubule reorientation from transverse to longitudinal might produce the growth inhibition elicited by auxin in the Arabidopsis root so quickly (Evans et al., 1994). Modification of the allometric ratio by changing the orientation of newly deposited cellulose microfibrils happens over hours and, therefore, is much too slow to account directly for the inhibition of cell expansion by auxin. Accompanying data on the growth changes produced by auxin in their experiments are not included in the report, nor are specifications of the investigated cell layers, despite the fact that the different cell layers show different responses to hormones (Ubeda-Tomás et al., 2012). Hence, critical questions regarding the quantitative relationship between microtubule reorientation and cell elongation remain unanswered. The gap between the experimental data presented and the far-reaching conclusions derived from them has been pointed out by Baskin (2015).There is, to our knowledge, no evidence for any fast growth-controlling mechanisms involving microtubules. There is, on the other hand, ample evidence for a causal relationship between wall-relaxing processes (such as the secretion or activation of wall-loosening enzymes or the generation of reactive oxygen species) and the rapid regulation of cell growth by auxin (Cosgrove, 2005; Perrot-Rechenmann, 2010). Modification of the allometric ratio by changing the orientation of newly deposited cellulose microfibrils appears much too slow (happening over hours) to account for the inhibition of cell expansion within minutes (Evans et al., 1994). Gravitropic bending of maize (Zea mays) roots has been shown to proceed normally even after the disassembly or immobilization of microtubules, and the inhibition of bending prohibits unilateral microtubule reorientation (Baluška et al., 1996). Chen et al. (2014) do not consider any of this evidence.So, can the data presented by Chen and collaborators be explained without coming into conflict with previously published results? The answer to this question is straightforward and has long been accessible in the pertinent literature: the observed changes in microtubule pattern are trivial consequences of either growth inhibition or the auxin insensitivity of growth in abp1 (Tromas et al., 2009). Consider the following points.First, mechanical forces can reorientate microtubules. There is a large body of experimental evidence that indicates that microtubule reorientations in single cells or tissues can be induced by oriented mechanical forces causing oriented stresses and strains in the affected cell walls (Landrein and Hamant, 2013). For example, Fisher and Cyr (2000) subjected protoplasts, embedded in an elastic agarose matrix, to mechanically induced stretching. The originally randomly oriented microtubules responded to this treatment by aligning at right angles to the major tensive force vector. Similarly, growing coleoptile segments respond to mechanical bending by reorientating the microtubules of epidermal cells at right angles to the direction of tension (extended side) and parallel to the direction of compression (compressed side; Fischer and Schopfer, 1997).Second, anisotropic cell growth mirrors patterns of stress and strain within the cell wall (Hamant and Traas, 2010). In biophysical terms, turgid plant cells can be regarded as pressurized vessels surrounded by an elastically stretched wall. Auxin-driven cell enlargement is brought about by changing the yielding properties of the wall and the resulting expansion in the direction of growth (Cosgrove, 2005).Third, experiments with maize coleoptiles have shown that auxin, in addition to effecting growth-related microtubule reorientations, strongly promotes their responsivity to mechanical forces. The epidermal microtubules of auxin-deprived coleoptile segments barely respond to bending stresses but reorientate rapidly after a 1-h treatment with auxin (Fischer and Schopfer, 1997). Hence, cell wall strains generated by growth or applied stresses interact in orientating microtubules in a synergistic manner, pointing to a common signaling mechanism activated by changes in strain rate.Summing up, there is well-founded evidence for the conclusion that the microtubule reorientations observed by Chen and collaborators occur as a result of changes in the physical strain pattern that underlies the auxin-induced changes in cell expansion. In agreement with established knowledge, the primary effect of auxin may be a rapid inhibition of cell wall loosening, mediated by the production of hydrogen peroxide (Ivanchenko et al., 2013). Based on these arguments and the weight of published evidence, we conclude that Chen and collaborators have inversed cause and effect.Their observation that the inactivation of ABP1 (and downstream components of the ABP1 pathway) causes microtubules to become unresponsive to auxin and lose their transverse pattern in roots may be explained as trivial consequences of growth inhibition (Tromas et al., 2009). Serious questions now hang over the roles for ABP1 in auxin signaling and auxin-controlled development (Gao et al., 2015; Liu, 2015). However, this does not come as a complete surprise. Hayashi et al. (2008) previously showed that the auxin-induced growth inhibition of root and hypocotyl in Arabidopsis can be suppressed by α[2,4-dimethylphenly-ethyl-2-oxo]-IAA (auxinole). This antagonist specifically competes with auxin at the TIR1-AUX/IAA-type receptor complexes and does not bind to ABP1, thereby suggesting that ABP1 does not act as a receptor in these pathways.There are lessons here, not the least that literature from the pre-Arabidopsis era remains a relevant and valuable source of information.     

Induction of TNF-α by LPS in Schwann Cell is Regulated by MAPK Activation Signals

Mitogen-activated protein kinases (MAPKs) are important mediators of cytokine expression and are critically involved in the immune response. The lipopolysaccharide (LPS) of gram-negative bacteria induces the expression of cytokines and proinflammatory genes via the toll-like receptor 4 (TLR4) signaling pathway in diverse cell types. In vivo, Schwann cells (SCs) at the site of injury may also produce tumor necrosis factor-- α (TNF-α). However, the precise mechanisms of TNF-α synthesis are still not clear. The purpose of the present study was to elucidate the underlying molecular mechanisms in the cultured SCs for its ability to activate the MAPKs and TNF-α gene, in response to LPS. Using enzyme-linked immunosorbent assay (ELISA), it was confirmed that treatment with LPS stimulated the synthesis of TNF-α in a concentration- and time-dependent manner. Intracellular location of TNF-α was detected under confocal microscope. Moreover, LPS activated extracellular signal-regulated kinase (ERK1/2), P38 and stress activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) and induced their phosphorylation. LPS-elicited SCs TNF-α production was also drastically suppressed by PD98059 (ERK inhibitor), SB202190 (P38 inhibitor), or SP600125 (SAPK/JNK inhibitor). Additionally, the expression of CD14 and TLR4 was examined by RT–PCR. It was demonstrated that the expression of CD14, TLR4 was crucial for the SCs responses to LPS. In conclusion, the results provide novel mechanisms for the response of SCs to LPS stimulation, through MAPKs signaling pathways. Chun Cheng and Yongwei Qin contributed equally to this work.     

The Nuclear Localization of γ-Tubulin Is Regulated by SadB-mediated Phosphorylation

γ-Tubulin is an important cell division regulator that arranges microtubule assembly and mitotic spindle formation. Cytosolic γ-tubulin nucleates α- and β-tubulin in a growing microtubule by forming the ring-shaped protein complex γTuRC. Nuclear γ-tubulin also regulates S-phase progression by moderating the activities of E2 promoter-binding factors. The mechanism that regulates localization of γ-tubulin is currently unknown. Here, we demonstrate that the human Ser/Thr kinase SadB short localizes to chromatin and centrosomes. We found that SadB-mediated phosphorylation of γ-tubulin on Ser³⁸⁵ formed chromatin-associated γ-tubulin complexes that moderate gene expression. In this way, the C-terminal region of γ-tubulin regulates S-phase progression. In addition, chromatin levels of γ-tubulin were decreased by the reduction of SadB levels or expression of a non-phosphorylatable Ala³⁸⁵-γ-tubulin but were enhanced by expression of SadB, wild-type γ-tubulin, or a phosphomimetic Asp³⁸⁵-γ-tubulin mutant. Our results demonstrate that SadB kinases regulate the cellular localization of γ-tubulin and thereby control S-phase progression.     

Cell Surface Localization of α3β4 Nicotinic Acetylcholine Receptors Is Regulated by N-Cadherin Homotypic Binding and Actomyosin Contractility

Neuronal nicotinic acetylcholine receptors (nAChRs) are widely expressed throughout the central and peripheral nervous system and are localized at synaptic and extrasynaptic sites of the cell membrane. However, the mechanisms regulating the localization of nicotinic receptors in distinct domains of the cell membrane are not well understood. N-cadherin is a cell adhesion molecule that mediates homotypic binding between apposed cell membranes and regulates the actin cytoskeleton through protein interactions with the cytoplasmic domain. At synaptic contacts, N-cadherin is commonly localized adjacent to the active zone and the postsynaptic density, suggesting that N-cadherin contributes to the assembly of the synaptic complex. To examine whether N-cadherin homotypic binding regulates the cell surface localization of nicotinic receptors, this study used heterologous expression of N-cadherin and α3β4 nAChR subunits C-terminally fused to a myc-tag epitope in Chinese hamster ovary cells. Expression levels of α3β4 nAChRs at cell-cell contacts and at contact-free cell membrane were analyzed by confocal microscopy. α3β4 nAChRs were found distributed over the entire surface of contacting cells lacking N-cadherin. In contrast, N-cadherin-mediated cell-cell contacts were devoid of α3β4 nAChRs. Cell-cell contacts mediated by N-cadherin-deleted proteins lacking the β-catenin binding region or the entire cytoplasmic domain showed control levels of α3β4 nAChRs expression. Inhibition of actin polymerization with latrunculin A and cytochalasin D did not affect α3β4 nAChRs localization within N-cadherin-mediated cell-cell contacts. However, treatment with the Rho associated kinase inhibitor Y27632 resulted in a significant increase in α3β4 nAChR levels within N-cadherin-mediated cell-cell contacts. Analysis of α3β4 nAChRs localization in polarized Caco-2 cells showed specific expression on the apical cell membrane and colocalization with apical F-actin and the actin nucleator Arp3. These results indicate that actomyosin contractility downstream of N-cadherin homotypic binding regulates the cell surface localization of α3β4 nAChRs presumably through interactions with a particular pool of F-actin.     

Cellular Auxin Homeostasis under High Temperature Is Regulated through a SORTING NEXIN1–Dependent Endosomal Trafficking Pathway

High-temperature-mediated adaptation in plant architecture is linked to the increased synthesis of the phytohormone auxin, which alters cellular auxin homeostasis. The auxin gradient, modulated by cellular auxin homeostasis, plays an important role in regulating the developmental fate of plant organs. Although the signaling mechanism that integrates auxin and high temperature is relatively well understood, the cellular auxin homeostasis mechanism under high temperature is largely unknown. Using the Arabidopsis thaliana root as a model, we demonstrate that under high temperature, roots counterbalance the elevated level of intracellular auxin by promoting shootward auxin efflux in a PIN-FORMED2 (PIN2)-dependent manner. Further analyses revealed that high temperature selectively promotes the retrieval of PIN2 from late endosomes and sorts them to the plasma membrane through an endosomal trafficking pathway dependent on SORTING NEXIN1. Our results demonstrate that recycling endosomal pathway plays an important role in facilitating plants adaptation to increased temperature.