Two eARCHT3.0 Lines for Optogenetic Silencing of Dopaminergic and Serotonergic Neurons

Dopaminergic and serotonergic neurons modulate and control processes ranging from reward signaling to regulation of motor outputs. Further, dysfunction of these neurons is involved in both degenerative and psychiatric disorders. Elucidating the roles of these neurons has been greatly facilitated by bacterial artificial chromosome (BAC) transgenic mouse lines expressing channelrhodopsin to readily enable cell-type specific activation. However, corresponding lines to silence these monoaminergic neurons have been lacking. We have generated two BAC transgenic mouse lines expressing the outward proton pump, enhanced ArchT3.0 (eArchT3.0), and GFP under control of the regulatory elements of either the dopamine transporter (DAT; Jax# 031663) or the tryptophan hydroxylase 2 (TPH2; Jax# 031662) gene locus. We demonstrate highly faithful and specific expression of these lines in dopaminergic and serotonergic neurons respectively. Additionally we validate effective and sensitive eArchT3.0-mediated silencing of these neurons using slice electrophysiology as well as with a well-established behavioral assay. These new transgenic tools will help expedite the study of dopaminergic and serotonergic system function in normal behavior and disease

Mechanical Stimuli Modulate Lateral Root Organogenesis1[W][OA]

Unlike mammals, whose development is limited to a short temporal window, plants produce organs de novo throughout their lifetime in order to adapt their architecture to the prevailing environmental conditions. The production of lateral roots represents one example of such postembryonic organogenesis. An endogenous developmental program likely imposes an ordered arrangement on the position of new lateral roots. However, environmental stimuli such as nutrient levels also affect the patterning of lateral root production. In addition, we have found that mechanical forces can act as one of the triggers that entrain lateral root production to the environment of the Arabidopsis (Arabidopsis thaliana) plant. We observed that physical bending of the root recruited new lateral root formation to the convex side of the resultant bend. Transient bending of 20 s was sufficient to elicit this developmental program. Such bending triggered a Ca2+ transient within the pericycle, and blocking this change in Ca2+ also blocked recruitment of new lateral root production to the curved region of the root. The initial establishment of the mechanically induced lateral root primordium was independent of an auxin supply from the shoot and was not disrupted by mutants in a suite of auxin transporters and receptor/response elements. These results suggest that Ca2+ may be acting to translate the mechanical forces inherent in growth to a developmental response in roots

Two eARCHT3.0 Lines for Optogenetic Silencing of Dopaminergic and Serotonergic Neurons

Dopaminergic and serotonergic neurons modulate and control processes ranging from reward signaling to regulation of motor outputs. Further, dysfunction of these neurons is involved in both degenerative and psychiatric disorders. Elucidating the roles of these neurons has been greatly facilitated by bacterial artificial chromosome (BAC) transgenic mouse lines expressing channelrhodopsin to readily enable cell-type specific activation. However, corresponding lines to silence these monoaminergic neurons have been lacking. We have generated two BAC transgenic mouse lines expressing the outward proton pump, enhanced ArchT3.0 (eArchT3.0), and GFP under control of the regulatory elements of either the dopamine transporter (DAT; Jax# 031663) or the tryptophan hydroxylase 2 (TPH2; Jax# 031662) gene locus. We demonstrate highly faithful and specific expression of these lines in dopaminergic and serotonergic neurons respectively. Additionally we validate effective and sensitive eArchT3.0-mediated silencing of these neurons using slice electrophysiology as well as with a well-established behavioral assay. These new transgenic tools will help expedite the study of dopaminergic and serotonergic system function in normal behavior and disease.National Institute of Neurological Disorders and Stroke (Grants R01NS098505 and R21NS079992)Simons Foundation (Grant 307913)National Institute of Mental Health (Grant R01MH099647)National Institute on Drug Abuse (Grant P50DA042012)Defense Sciences Office (Contract W911NF-14-2-0013

Geology of the Cenozoic Indus Basin sedimentary rocks : paleoenvironmental interpretation of sedimentation from the western Himalaya during the early phases of India-Eurasia collision.

This study reassesses the stratigraphy, sedimentology, and provenance of the Indus Basin sedimentary rocks, deposited within the Indus Tsangpo Suture Zone (ITSZ) during the early phases of India‐Eurasia collision. Using field observations, biostratigraphy, and petrographic and isotopic analyses we create a paleodepositional reconstruction within the paleotectonic setting of the early phases of India‐Eurasia collision. We then re‐examine existing constraints to the timing of India‐Eurasia collision previously interpreted from the earliest occurrence of mixed Indian‐ and Eurasian‐derived detritus in the succession. From mid‐Cretaceous to early Paleocene times the Jurutze and Sumda Formations were deposited within an arc‐bounded marine basin between the Dras and Kohistan‐Ladakh Island arcs. The <51 Ma aged deltaic Chogdo Formation then filled the basin until deposition of the 50.8–49.4 Ma aged Nummulitic Limestone during a marine incursion, before continental facies developed in an evolving intermountain basin with the deposition of the Paleogene Indus Group. Within these systems, sediment was sourced from the Eurasian margin to the north and was transported southward into the suture zone. In this section, we see no unequivocal evidence of Indian Plate input to the sedimentary succession (and thus no evidence of mixed Indian‐Eurasian‐derived detritus indicative of India‐Asia collision) until the upper stratigraphic horizons of the Indus Group, when facies are representative of an axial, northwesterly flowing river system. We suggest that the paleo‐Indus River was initiated within the ITSZ during late Oligocene‐early Miocene times. Sedimentation of the Indus Group continued until the late Miocene

Transcriptional and posttranscriptional silencing are mechanistically related

Two distinct gene-silencing phenomena are observed in plants: transcriptional gene silencing (TGS), which involves decreased RNA synthesis because of promoter methylation, and posttranscriptional gene silencing (PTGS), which involves sequence-specific RNA degradation. PTGS is induced by deliberate [1-4] or fortuitous production (R.v.B., unpublished data) of double-stranded RNA (dsRNA). TGS could be the result of DNA pairing [5], but could also be the result of dsRNA, as was shown by the dsRNA-induced inactivation of a transgenic promoter [6]. Here, we show that when targeting flower pigmentation genes in Petunia, transgenes expressing dsRNA can induce PTGS when coding sequences are used and TGS when promoter sequences are taken. For both types of silencing, small RNA species are found, which are thought to be dsRNA decay products [7] and determine the sequence specificity of the silencing process [8, 9]. Furthermore, silencing is accompanied by the methylation of DNA sequences that are homologous to dsRNA. DNA methylation is assumed to be essential for regulating TGS and important for reinforcing PTGS [10]. Therefore, we conclude that TGS and PTGS are mechanistically related. In addition, we show that dsRNA-induced TGS provides an efficient tool to generate gene knockouts, because not only does the TGS of a PTGS-inducing transgene fully revert the PTGS phenotype, but also an endogenous gene can be transcriptionally silenced by dsRNA corresponding to its promoter

T-bet+ B cells are activated by and control endogenous retroviruses through TLR-dependent mechanisms

Abstract Endogenous retroviruses (ERVs) are an integral part of the mammalian genome. The role of immune control of ERVs in general is poorly defined as is their function as anti-cancer immune targets or drivers of autoimmune disease. Here, we generate mouse-strains where Moloney-Murine Leukemia Virus tagged with GFP (ERV-GFP) infected the mouse germline. This enables us to analyze the role of genetic, epigenetic and cell intrinsic restriction factors in ERV activation and control. We identify an autoreactive B cell response against the neo-self/ERV antigen GFP as a key mechanism of ERV control. Hallmarks of this response are spontaneous ERV-GFP+ germinal center formation, elevated serum IFN-γ levels and a dependency on Age-associated B cells (ABCs) a subclass of T-bet+ memory B cells. Impairment of IgM B cell receptor-signal in nucleic-acid sensing TLR-deficient mice contributes to defective ERV control. Although ERVs are a part of the genome they break immune tolerance, induce immune surveillance against ERV-derived self-antigens and shape the host immune response