Optical control of mammalian endogenous transcription and epigenetic states

A theoretical underpinning of the standard model of fundamental particles and interactions is CPT invariance, which requires that the laws of physics be invariant under the combined discrete operations of charge conjugation, parity and time reversal. Antimatter, the existence of which was predicted by Dirac, can be used to test the CPT theorem—experimental investigations involving comparisons of particles with antiparticles are numerous. Cold atoms and anti-atoms, such as hydrogen and antihydrogen, could form the basis of a new precise test, as CPT invariance implies that they must have the same spectrum. Observations of antihydrogen in small quantities and at high energies have been reported at the European Organization for Nuclear Research (CERN) and at Fermilab, but these experiments were not suited to precision comparison measurements. Here we demonstrate the production of antihydrogen atoms at very low energy by mixing trapped antiprotons and positrons in a cryogenic environment. The neutral anti-atoms have been detected directly when they escape the trap and annihilate, producing a characteristic signature in an imaging particle detector

Optical Control of Mammalian Endogenous Transcription and Epigenetic States

The dynamic nature of gene expression enables cellular programming, homeostasis, and environmental adaptation in living systems. Dissection of causal gene functions in cellular and organismal processes therefore necessitates approaches that enable spatially and temporally precise modulation of gene expression. Recently, a variety of microbial and plant-derived light-sensitive proteins have been engineered as optogenetic actuators, enabling high precision spatiotemporal control of many cellular functions1-11. However, versatile and robust technologies that enable optical modulation of transcription in the mammalian endogenous genome remain elusive. Here, we describe the development of Light-Inducible Transcriptional Effectors (LITEs), an optogenetic two-hybrid system integrating the customizable TALE DNA-binding domain12-14 with the light-sensitive cryptochrome 2 protein and its interacting partner CIB1 from Arabidopsis thaliana. LITEs do not require additional exogenous chemical co-factors, are easily customized to target many endogenous genomic loci, and can be activated within minutes with reversibility3,4,6,7,15. LITEs can be packaged into viral vectors and genetically targeted to probe specific cell populations. We have applied this system in primary mouse neurons, as well as in the brain of awake mice in vivo to mediate reversible modulation of mammalian endogenous gene expression as well as targeted epigenetic chromatin modifications. The LITE system establishes a novel mode of optogenetic control of endogenous cellular processes and enables direct testing of the causal roles of genetic and epigenetic regulation in normal biological processes and disease states

Expression of survivin detected by immunohistochemistry in the cytoplasm and in the nucleus is associated with prognosis of leiomyosarcoma and synovial sarcoma patients

<p>Abstract</p> <p>Background</p> <p>Survivin, a member of the inhibitor of apoptosis-protein family suppresses apoptosis and regulates cell division. It is strongly overexpressed in the vast majority of cancers. We were interested if survivin detected by immunohistochemistry has prognostic relevance especially for patients of the two soft tissue sarcoma entities leiomyosarcoma and synovial sarcoma.</p> <p>Methods</p> <p>Tumors of leiomyosarcoma (n = 24) and synovial sarcoma patients (n = 26) were investigated for their expression of survivin by immunohistochemistry. Survivin expression was assessed in the cytoplasm and the nucleus of tumor cells using an immunoreactive scoring system (IRS).</p> <p>Results</p> <p>We detected a survivin expression (IRS > 2) in the cytoplasm of 20 leiomyosarcomas and 22 synovial sarcomas and in the nucleus of 12 leiomyosarcomas and 9 synovial sarcomas, respectively. There was no significant difference between leiomyosarcoma and synovial sarcoma samples in their cytoplasmic or nuclear expression of survivin. Next, all sarcoma patients were separated in four groups according to their survivin expression in the cytoplasm and in the nucleus: group 1: negative (IRS 0 to 2); group 2: weak (IRS 3 to 4); group 3: moderate (IRS 6 to 8); group 4: strong (IRS 9 to 12). In a multivariate Cox's regression hazard analysis survivin expression detected in the cytoplasm or in the nucleus was significantly associated with overall survival of patients in group 3 (RR = 5.7; P = 0.004 and RR = 5.7; P = 0.022, respectively) compared to group 2 (reference). Patients whose tumors showed both a moderate/strong expression of survivin in the cytoplasm and a moderate expression of survivin in the nucleus (in both compartments IRS ≥ 6) possessed a 24.8-fold increased risk of tumor-related death (P = 0.003) compared to patients with a weak expression of survivin both in the cytoplasm and in the nucleus.</p> <p>Conclusion</p> <p>Survivin protein expression in the cytoplasma and in the nucleus detected by immunohistochemistry is significantly associated with prognosis of leiomyosarcoma and synovial sarcoma patients.</p

Does Intensity Modulated Radiation Therapy (IMRT) prevent additional toxicity of treating the pelvic lymph nodes compared to treatment of the prostate only?

<p>Abstract</p> <p>Background</p> <p>To evaluate the risk of rectal, bladder and small bowel toxicity in intensity modulated radiation therapy (IMRT) of the prostate only compared to additional irradiation of the pelvic lymphatic region.</p> <p>Methods</p> <p>For ten patients with localized prostate cancer, IMRT plans with a simultaneous integrated boost (SIB) were generated for treatment of the prostate only (plan-PO) and for additional treatment of the pelvic lymph nodes (plan-WP). In plan-PO, doses of 60 Gy and 74 Gy (33 fractions) were prescribed to the seminal vesicles and to the prostate, respectively. Three plans-WP were generated with prescription doses of 46 Gy, 50.4 Gy and 54 Gy to the pelvic target volume; doses to the prostate and seminal vesicles were identical to plan-PO. The risk of rectal, bladder and small bowel toxicity was estimated based on NTCP calculations.</p> <p>Results</p> <p>Doses to the prostate were not significantly different between plan-PO and plan-WP and doses to the pelvic lymph nodes were as planned. Plan-WP resulted in increased doses to the rectum in the low-dose region ≤ 30 Gy, only, no difference was observed in the mid and high-dose region. Normal tissue complication probability (NTCP) for late rectal toxicity ranged between 5% and 8% with no significant difference between plan-PO and plan-WP. NTCP for late bladder toxicity was less than 1% for both plan-PO and plan-WP. The risk of small bowel toxicity was moderately increased for plan-WP.</p> <p>Discussion</p> <p>This retrospective planning study predicted similar risks of rectal, bladder and small bowel toxicity for IMRT treatment of the prostate only and for additional treatment of the pelvic lymph nodes.</p

CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling

CRISPR-Cas9 is a versatile genome editing technology for studying the functions of genetic elements. To broadly enable the application of Cas9 in vivo, we established a Cre-dependent Cas9 knockin mouse. We demonstrated in vivo as well as ex vivo genome editing using adeno-associated virus (AAV)-, lentivirus-, or particle-mediated delivery of guide RNA in neurons, immune cells, and endothelial cells. Using these mice, we simultaneously modeled the dynamics of KRAS, p53, and LKB1, the top three significantly mutated genes in lung adenocarcinoma. Delivery of a single AAV vector in the lung generated loss-of-function mutations in p53 and Lkb1, as well as homology-directed repair-mediated Kras[superscript G12D] mutations, leading to macroscopic tumors of adenocarcinoma pathology. Together, these results suggest that Cas9 mice empower a wide range of biological and disease modeling applications.National Science Foundation (U.S.). Graduate Research Fellowship (Grant 1122374)Damon Runyon Cancer Research Foundation (Fellowship DRG-2117-12)Massachusetts Institute of Technology. Simons Center for the Social Brain (Postdoctoral Fellowship)European Molecular Biology Organization (Fellowship)Foundation for Polish Science (Fellowship)American Society for Engineering Education. National Defense Science and Engineering Graduate FellowshipNational Science Foundation (U.S.). Graduate Research FellowshipMassachusetts Institute of Technology (Presidential Graduate Fellowship)Human Frontier Science Program (Strasbourg, France) (Postdoctoral Fellowship)National Human Genome Research Institute (U.S.) (CEGS P50 HG006193)Howard Hughes Medical InstituteKlarman Cell ObservatoryNational Cancer Institute (U.S.) (Center of Cancer Nanotechnology Excellence Grant U54CA151884)National Institutes of Health (U.S.) (Controlled Release Grant EB000244)National Heart, Lung, and Blood Institute (Program of Excellence in Nanotechnology (PEN) Award Contract HHSN268201000045C)Massachusetts Institute of Technology (Poitras Gift 1631119)Stanley CenterSimons Foundation (6927482)Nancy Lurie Marks Family Foundation (6928117)United States. Public Health Service (National Institutes of Health (U.S.) R01-CA133404)David H. Koch Institute for Integrative Cancer Research at MIT (Marie D. and Pierre Casimir-Lambert Fund)MIT Skoltech InitiativeNational Cancer Institute (U.S.) (Koch Institute Support (Core) Grant P30-CA14051)National Institute of Mental Health (U.S.) (Director’s Pioneer Award DP1-MH100706)National Institute of Neurological Disorders and Stroke (U.S.) (Transformative R01 Grant R01-NS 07312401)National Science Foundation (U.S.) (Waterman Award)W. M. Keck FoundationKinship Foundation. Searle Scholars ProgramKlingenstein FoundationVallee FoundationMerkin Foundatio

Optical Control of Mammalian Endogenous Transcription and Epigenetic States

The dynamic nature of gene expression enables cellular programming, homeostasis, and environmental adaptation in living systems. Dissection of causal gene functions in cellular and organismal processes therefore necessitates approaches that enable spatially and temporally precise modulation of gene expression. Recently, a variety of microbial and plant-derived light-sensitive proteins have been engineered as optogenetic actuators, enabling high precision spatiotemporal control of many cellular functions1-11. However, versatile and robust technologies that enable optical modulation of transcription in the mammalian endogenous genome remain elusive. Here, we describe the development of Light-Inducible Transcriptional Effectors (LITEs), an optogenetic two-hybrid system integrating the customizable TALE DNA-binding domain12-14 with the light-sensitive cryptochrome 2 protein and its interacting partner CIB1 from Arabidopsis thaliana. LITEs do not require additional exogenous chemical co-factors, are easily customized to target many endogenous genomic loci, and can be activated within minutes with reversibility3,4,6,7,15. LITEs can be packaged into viral vectors and genetically targeted to probe specific cell populations. We have applied this system in primary mouse neurons, as well as in the brain of awake mice in vivo to mediate reversible modulation of mammalian endogenous gene expression as well as targeted epigenetic chromatin modifications. The LITE system establishes a novel mode of optogenetic control of endogenous cellular processes and enables direct testing of the causal roles of genetic and epigenetic regulation in normal biological processes and disease states

Convergence Without Hard Criteria: Does EU Soft Law Affect Domestic Unemployment Protection Schemes?

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Applications of CRISPR–Cas systems in neuroscience

Genome-editing tools, and in particular those based on CRISPR-Cas (clustered regularly interspaced short palindromic repeat (CRISPR)-CRISPR-associated protein) systems, are accelerating the pace of biological research and enabling targeted genetic interrogation in almost any organism and cell type. These tools have opened the door to the development of new model systems for studying the complexity of the nervous system, including animal models and stem cell-derived in vitro models. Precise and efficient gene editing using CRISPR-Cas systems has the potential to advance both basic and translational neuroscience research.National Institute of Mental Health (U.S.) (Grant 5DP1-MH100706)National Institute of Mental Health (U.S.) (Grant 1R01-MH110049)National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (Grant 5R01DK097768-03