Dissecting the telomere-independent pathways underlying human cellular senescence

Cellular senescence is an irreversible program of cell cycle arrest triggered in normal somatic cells in response to a variety of intrinsic and extrinsic stimuli including telomere attrition, DNA damage, physiological stress and oncogene activation. Finding that inactivation of the pRB and p53 pathways by SV40-LT antigen cooperates with hTERT to immortalize cells has allowed us to use a thermolabile mutant of SV40-LT to develop human fibroblasts where the cells are immortal if grown at 34oC but undergo an irreversible growth arrest within 5 days at 38oC. When these cells cease dividing, senescence-associated-β-galactosidase (SA-β-Gal) activity is induced and the growth-arrested cells have many features of senescent cells. Since these cells growth-arrest in a synchronous manner, I have used Affymetrix expression profiling to identify the genes differentially expressed upon senescence. This identified 816 up- and 961 down-regulated genes whose expression was reversed when growth arrest was abrogated. I have shown that senescence was associated with activation of the NF-κB pathway and up-regulation of a number of senescence-associated-secretory-proteins including IL6. Perturbation of NF-κB signalling either by direct silencing of NF-κB subunits or by upstream modulation overcame growth-arrest indicating that activation of NF-κB signalling has a causal role in promoting senescence. I also applied a retroviral shRNA screen covering ~10,000 genes to the same cell model. Overlapping with the microarray data revealed particularly interesting targets, such as LTBP3 and Layilin. Finally, I profiled micro-rna expression. 15 of the top micro-rnas down-regulated upon senescence were chosen to express in the HMF3A system. 6 of them were able to bypass the growth-arrest. In conclusion, my work has uncovered novel markers involved in senescence as well as identifying that both activation of p53 and pRb pathway result in activation of NF-κB signalling which promotes senescence. Both results lead to a better understanding of senescence and its pathways

An RNA interference screen for identifying downstream effectors of the p53 and pRB tumour suppressor pathways involved in senescence

Background: Cellular senescence is an irreversible cell cycle arrest that normal cells undergo in response to progressive shortening of telomeres, changes in telomeric structure, oncogene activation or oxidative stress and acts as an important tumour suppressor mechanism.Results: To identify the downstream effectors of the p53-p21 and p16-pRB tumour suppressor pathways crucial for mediating entry into senescence, we have carried out a loss-of-function RNA interference screen in conditionally immortalised human fibroblasts that can be induced to rapidly undergo senescence, whereas in primary cultures senescence is stochastic and occurs asynchronously. These cells are immortal but undergo a rapid irreversible arrest upon activation of the p53-p21 and p16-pRB pathways that can be readily bypassed upon their inactivation. The primary screen identified 112 known genes including p53 and another 29 shRNAmirs targetting as yet unidentified loci. Comparison of these known targets with genes known to be up-regulated upon senescence in these cells, by micro-array expression profiling, identified 4 common genes TMEM9B, ATXN10, LAYN and LTBP2/3. Direct silencing of these common genes, using lentiviral shRNAmirs, bypassed senescence in the conditionally immortalised cells.Conclusion: The senescence bypass screen identified TMEM9B, ATXN10, LAYN and LTBP2/3 as novel downstream effectors of the p53-p21 and p16-pRB tumour suppressor pathways. Although none of them has previously been linked to cellular senescence, TMEM9B has been suggested to be an upstream activator of NF-kappa B signalling which has been found to have a causal role in promoting senescence. Future studies will focus on determining on how many of the other primary hits also have a casual role in senescence and what is the mechanism of action

The 2019 surface acoustic waves roadmap

Today, surface acoustic waves (SAWs) and bulk acoustic waves are already two of the very few phononic technologies of industrial relevance and can been found in a myriad of devices employing these nanoscale earthquakes on a chip. Acoustic radio frequency filters, for instance, are integral parts of wireless devices. SAWs in particular find applications in life sciences and microfluidics for sensing and mixing of tiny amounts of liquids. In addition to this continuously growing number of applications, SAWs are ideally suited to probe and control elementary excitations in condensed matter at the limit of single quantum excitations. Even collective excitations, classical or quantum are nowadays coherently interfaced by SAWs. This wide, highly diverse, interdisciplinary and continuously expanding spectrum literally unites advanced sensing and manipulation applications. Remarkably, SAW technology is inherently multiscale and spans from single atomic or nanoscopic units up even to the millimeter scale. The aim of this Roadmap is to present a snapshot of the present state of surface acoustic wave science and technology in 2019 and provide an opinion on the challenges and opportunities that the future holds from a group of renown experts, covering the interdisciplinary key areas, ranging from fundamental quantum effects to practical applications of acoustic devices in life science

Chirality of Matter Shows Up via Spin Excitations

Right- and left-handed circularly polarized light interact differently with electronic charges in chiral materials. This asymmetry generates the natural circular dichroism and gyrotropy, also known as the optical activity. Here we demonstrate that optical activity is not a privilege of the electronic charge excitations but it can also emerge for the spin excitations in magnetic matter. The square-lattice antiferromagnet Ba$_2$CoGe$_2$O$_7$ offers an ideal arena to test this idea, since it can be transformed to a chiral form by application of external magnetic fields. As a direct proof of the field-induced chiral state, we observed large optical activity when the light is in resonance with spin excitations at sub-terahertz frequencies. In addition, we found that the magnetochiral effect, the absorption difference for the light beams propagating parallel and anti-parallel to the applied magnetic field, has an exceptionally large amplitude close to 100%. All these features are ascribed to the magnetoelectric nature of spin excitations as they interact both with the electric and magnetic components of light

Cell-based screen for altered nuclear phenotypes reveals senescence progression in polyploid cells after Aurora kinase B inhibition.

Cellular senescence is a widespread stress response and is widely considered to be an alternative cancer therapeutic goal. Unlike apoptosis, senescence is composed of a diverse set of subphenotypes, depending on which of its associated effector programs are engaged. Here we establish a simple and sensitive cell-based prosenescence screen with detailed validation assays. We characterize the screen using a focused tool compound kinase inhibitor library. We identify a series of compounds that induce different types of senescence, including a unique phenotype associated with irregularly shaped nuclei and the progressive accumulation of G1 tetraploidy in human diploid fibroblasts. Downstream analyses show that all of the compounds that induce tetraploid senescence inhibit Aurora kinase B (AURKB). AURKB is the catalytic component of the chromosome passenger complex, which is involved in correct chromosome alignment and segregation, the spindle assembly checkpoint, and cytokinesis. Although aberrant mitosis and senescence have been linked, a specific characterization of AURKB in the context of senescence is still required. This proof-of-principle study suggests that our protocol is capable of amplifying tetraploid senescence, which can be observed in only a small population of oncogenic RAS-induced senescence, and provides additional justification for AURKB as a cancer therapeutic target.This work was supported by the University of Cambridge, Cancer Research UK, Hutchison Whampoa; Cancer Research UK grants A6691 and A9892 (M.N., N.K., C.J.T., D.C.B., C.J.C., L.S.G, and M.S.); a fellowship from the Uehara Memorial Foundation (M.S.).This is the author accepted manuscript. The final version is available from the American Society for Cell Biology via http://dx.doi.org/10.1091/mbc.E15-01-000

The 2019 surface acoustic waves roadmap

Abstract Today, surface acoustic waves (SAWs) and bulk acoustic waves are already two of the very few phononic technologies of industrial relevance and can been found in a myriad of devices employing these nanoscale earthquakes on a chip. Acoustic radio frequency filters, for instance, are integral parts of wireless devices. SAWs in particular find applications in life sciences and microfluidics for sensing and mixing of tiny amounts of liquids. In addition to this continuously growing number of applications, SAWs are ideally suited to probe and control elementary excitations in condensed matter at the limit of single quantum excitations. Even collective excitations, classical or quantum are nowadays coherently interfaced by SAWs. This wide, highly diverse, interdisciplinary and continuously expanding spectrum literally unites advanced sensing and manipulation applications. Remarkably, SAW technology is inherently multiscale and spans from single atomic or nanoscopic units up even to the millimeter scale. The aim of this Roadmap is to present a snapshot of the present state of surface acoustic wave science and technology in 2019 and provide an opinion on the challenges and opportunities that the future holds from a group of renown experts, covering the interdisciplinary key areas, ranging from fundamental quantum effects to practical applications of acoustic devices in life science.EU Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie Grant Agreement No. 642688 (SAWtrain)

Spin phonon and magnetoelectric coupling in oxygen isotope substituted TbMnO3 investigated by Raman scattering

In order to investigate the effects leading to the strong magnetoelectric coupling in the type II multiferroic TbMnO3 we have studied the thermal properties and temperature dependence of the lattice vibrations of TbMn16O3 and its isotopically substituted counterpart TbMn18O3. Heat capacity measurements on powder samples revealed no significant change in the Mn3 and Tb3 magnetic phase transition temperatures, as well as the multiferroic ordering temperature upon isotope substitution, indicating that a change in the dynamical modulation of the MnO6 octahedral distortions and rotations altering the Mn O Mn bond angles has no influence on the magnetic properties of TbMnO3. Raman light scattering experiments have been performed on isotopically substituted single crystals to determine the temperature induced changes in phonon energies and linewidths at the sinusoidal and multiferroic phase transitions. A detailed modeling indicates that the spin phonon coupling can be accounted for the pronounced anomalies in the temperature dependence of the phonon behaviors at the transition to the sinusoidal spin phase at TMnN 41 K and to the multiferroic spin spiral phase at TFE 28 K. No further effects such as the appearance of the electric polarization or the electromagnon were required to explain the data, especially below the multiferroic phase transitio