A shared transcriptional code orchestrates temporal patterning of the central nervous system

The molecular mechanisms that produce the full array of neuronal subtypes in the vertebrate nervous system are incompletely understood. Here, we provide evidence of a global temporal patterning program comprising sets of transcription factors that stratifies neurons based on the developmental time at which they are generated. This transcriptional code acts throughout the central nervous system, in parallel to spatial patterning, thereby increasing the diversity of neurons generated along the neuraxis. We further demonstrate that this temporal program operates in stem cell−derived neurons and is under the control of the TGFβ signaling pathway. Targeted perturbation of components of the temporal program, Nfia and Nfib, reveals their functional requirement for the generation of late-born neuronal subtypes. Together, our results provide evidence for the existence of a previously unappreciated global temporal transcriptional program of neuronal subtype identity and suggest that the integration of spatial and temporal patterning mechanisms diversifies and organizes neuronal subtypes in the vertebrate nervous system.

Lateral transduction is inherent to the life cycle of the archetypical Salmonella phage P22

Lysogenic induction ends the stable association between a bacteriophage and its host, and the transition to the lytic cycle begins with early prophage excision followed by DNA replication and packaging (ERP). This temporal program is considered universal for P22-like temperate phages, though there is no direct evidence to support the timing and sequence of these events. Here we report that the long-standing ERP program is an observation of the experimentally favored Salmonella phage P22 tsc229 heat-inducible mutant, and that wild-type P22 actually follows the replication-packaging-excision (RPE) program. We find that P22 tsc229 excises early after induction, but P22 delays excision to just before it is detrimental to phage production. This allows P22 to engage in lateral transduction. Thus, at minimal expense to itself, P22 has tuned the timing of excision to balance propagation with lateral transduction, powering the evolution of its host through gene transfer in the interest of self-preservation.

The secret life (cycle) of temperate bacteriophages

Lysogenic induction ends the stable association between a bacteriophage and its host, and the transition to the lytic cycle begins with prophage excision followed by DNA replication and packaging (ERP) – a temporal program that is considered universal for most temperate phages. Here we report that the long-standing ERP program is an artefact of the experimentally favoured Salmonella phage P22 tsc229 heat-inducible mutant, and that wildtype P22 actually follows a replication-packaging-excision (RPE) program. We found that unlike P22 tsc229, P22 delayed excision to just before it was detrimental to phage production. Thus, at minimal expense to itself, P22 has tuned the timing of excision to balance propagation with lateral transduction, powering the evolution of its host through gene transfer in the interest of self-preservation.One Sentence SummaryGenetic analyses propose a new life cycle for temperate bacteriophages.

Temporal patterning of the central nervous system by a shared transcription factor code

The molecular mechanisms that ensure the reproducible generation of neuronal diversity in the vertebrate nervous system are incompletely understood. Here we provide evidence of a temporal patterning program consisting of cohorts of transcription factors expressed in neurons generated at successive developmental timepoints. This program acts in parallel to spatial patterning, diversifying neurons throughout the nervous system and in neurons differentiated in-vitro from stem cells. We demonstrate the TGFβ signalling pathway controls the pace of the temporal program. Furthermore, targeted perturbation of components of the temporal program, Nfia and Nfib, reveals their requirement for the generation of late-born neuronal subtypes. Together, our results provide evidence for the existence of a previously unappreciated global temporal program of neuronal subtype identity and suggest that the integration of spatial and temporal patterning programs diversifies and organises neuronal subtypes in the vertebrate nervous system.

Extrinsic activin signaling cooperates with an intrinsic temporal program to increase mushroom body neuronal diversity

Temporal patterning of neural progenitors leads to the sequential production of diverse neurons. To understand how extrinsic cues influence intrinsic temporal programs, we studied Drosophila mushroom body progenitors (neuroblasts) that sequentially produce only three neuronal types: γ, then α’β’, followed by αβ. Opposing gradients of two RNA-binding proteins Imp and Syp comprise the intrinsic temporal program. Extrinsic activin signaling regulates the production of α’β’ neurons but whether it affects the intrinsic temporal program was not known. We show that the activin ligand Myoglianin from glia regulates the temporal factor Imp in mushroom body neuroblasts. Neuroblasts missing the activin receptor Baboon have a delayed intrinsic program as Imp is higher than normal during the α’β’ temporal window, causing the loss of α’β’ neurons, a decrease in αβ neurons, and a likely increase in γ neurons, without affecting the overall number of neurons produced. Our results illustrate that an extrinsic cue modifies an intrinsic temporal program to increase neuronal diversity.

Extrinsic Activin signaling cooperates with an intrinsic temporal program to increase mushroom body neuronal diversity

SummaryTemporal patterning of neural progenitors leads to the sequential production of diverse neuronal types. To better understand how extrinsic cues interact with intrinsic temporal programs to contribute to temporal patterning, we studied the Drosophila mushroom body neural progenitors (neuroblasts). Each of these four neuroblasts divides ~250 times to sequentially produce only three main neuronal types over the course of ~9 days of development: γ, followed by α’β’, and finally αβ neurons. The intrinsic temporal clock is composed of two RNA-binding proteins, IGF-II mRNA binding protein (Imp) and Syncrip (Syp), that are expressed in opposing temporal gradients. Activin signaling affects the production of α’β’ neurons but whether and how this extrinsic cue interacts with the intrinsic temporal program was not known. We show that the Activin ligand Myoglianin produced from glia regulates the levels of the intrinsic temporal factor Imp in mushroom body neuroblasts. In neuroblasts mutant for the Activin signaling receptor baboon, Imp levels are higher than normal during the α’β’ temporal window, leading to the specific loss of the α’β’ neurons. The intrinsic temporal clock still progresses but with a delay, skipping the α’β’ window without affecting the total number of neurons produced: The number of γ neurons increases, α’β’ disappear and the number of αβ neurons decreases. Our results illustrate that an extrinsic cue modifies an intrinsic temporal program to increase neuronal diversity.

New, easy, quick and efficient DNA replication timing analysis by high-throughput approaches

DNA replication must be faithful and follow a well-defined spatio-temporal program closely linked to transcriptional activity, epigenomic marks, intra-nuclear structures, mutation rate and cell fate determination. Among the readouts of the DNA replication spatio-temporal program, replication timing (RT) analyses require complex, precise and time-consuming experimental procedures, and the study of large-size computer files. We improved the RT protocol to speed it up and increase its quality and reproducibility. Also, we partly automated the RT protocol and developed a user-friendly software: the START-R suite (Simple Tool for the Analysis of the Replication Timing based on R). START-R suite is an open source web application using an R script and an HTML interface to analyze DNA replication timing in a given cell line with microarray or deep-sequencing results. This novel approach can be used by every biologist without requiring specific knowledge in bioinformatics. It also reduces the time required for generating and analyzing simultaneously data from several samples. START-R suite detects constant timing regions (CTR) but also, and this is a novelty, it identifies temporal transition regions (TTR) and detects significant differences between two experimental conditions. The informatic global analysis requires less than 10 minutes.