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Layer 5 extratelencephalic (ET) neurons are present across neocortical areas and send axons to multiple subcortical targets1–6. Two cardinal subtypes exist7,8: (1) Slco2a1-expressing neurons (ETdist), which predominate in the motor cortex and project distally to the pons, medulla and spinal cord; and (2) Nprs1- or Hpgd-expressing neurons (ETprox), which predominate in the visual cortex and project more proximally to the pons and thalamus. An understanding of how area-specific ETdist and ETprox emerge during development is important because they are critical for fine motor skills and are susceptible to spinal cord injury and amyotrophic lateral sclerosis9–12. Here, using cross-areal mapping of axonal projections in the mouse neocortex, we identify the subtype-specific developmental dynamics of ET neurons. Whereas subsets of ETprox emerge by pruning of ETdist axons, others emerge de novo. We outline corresponding subtype-specific developmental transcriptional programs using single-nucleus sequencing. Leveraging these findings, we use postnatal in vivo knockdown of subtype-specific transcription factors to reprogram ET neuron connectivity towards more proximal targets. Together, these results show the functional transcriptional programs driving ET neuron diversity and uncover cell subtype-specific gene regulatory networks that can be manipulated to direct target specificity in motor corticofugal pathways.Using cross-areal mapping of axonal projections in the mouse neocortex, we identify the subtype-specific developmental dynamics of extratelencephalic neurons and show the functional transcriptional programs driving extratelencephalic neuron diversity. [ABSTRACT FROM AUTHOR]

It is estimated that only 0.02% of disseminated tumour cells are able to seed overt metastases1. While this suggests the presence of environmental constraints to metastatic seeding, the landscape of host factors controlling this process remains largely unclear. Here, combining transposon technology2 and fluorescence niche labelling3, we developed an in vivo CRISPR activation screen to systematically investigate the interactions between hepatocytes and metastatic cells. We identify plexin B2 as a critical host-derived regulator of liver colonization in colorectal and pancreatic cancer and melanoma syngeneic mouse models. We dissect a mechanism through which plexin B2 interacts with class IV semaphorins on tumour cells, leading to KLF4 upregulation and thereby promoting the acquisition of epithelial traits. Our results highlight the essential role of signals from the liver parenchyma for the seeding of disseminated tumour cells before the establishment of a growth-promoting niche. Our findings further suggest that epithelialization is required for the adaptation of CRC metastases to their new tissue environment. Blocking the plexin-B2–semaphorin axis abolishes metastatic colonization of the liver and therefore represents a therapeutic strategy for the prevention of hepatic metastases. Finally, our screening approach, which evaluates host-derived extrinsic signals rather than tumour-intrinsic factors for their ability to promote metastatic seeding, is broadly applicable and lays a framework for the screening of environmental constraints to metastasis in other organs and cancer types.Interactions between plexin B2 on hepatocytes and sempahorins on disseminated tumour cells regulate metastatic seeding in the liver. [ABSTRACT FROM AUTHOR]

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 functions. 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 domain with the light-sensitive cryptochrome 2 protein and its interacting partner CIB1 from Arabidopsis thaliana. LITEs do not require additional exogenous chemical cofactors, are easily customized to target many endogenous genomic loci, and can be activated within minutes with reversibility. 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 freely behaving 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.

The ability to record transcriptional events within a cell over time would help to elucidate how molecular events give rise to complex cellular behaviours and states. However, current molecular recording technologies capture only a small set of defined stimuli. Here we use CRISPR spacer acquisition to capture and convert intracellular RNAs into DNA, enabling DNA-based storage of transcriptional information. In Escherichia coli, we show that defined stimuli, such as an RNA virus or arbitrary sequences, as well as complex stimuli, such as oxidative stress, result in quantifiable transcriptional records that are stored within a population of cells. We demonstrate that the transcriptional records enable us to classify and describe complex cellular behaviours and to identify the precise genes that orchestrate differential cellular responses. In the future, CRISPR spacer acquisition-mediated recording of RNA followed by deep sequencing (Record-seq) could be used to reconstruct transcriptional histories that describe complex cell behaviours or pathological states.An RNA-adapting CRISPR-Cas system is coupled with amplification and sequencing steps to record, retrieve and analyse changes in the transcriptome of a bacterial cell over time., Author(s): Florian Schmidt [sup.1] , Mariia Y. Cherepkova [sup.1] , Randall J. Platt [sup.1] [sup.2] Author Affiliations:(1) Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland(2) Department of Chemistry, [...]

The ultimate goal of genome editing is to be able to make any specific change to the blueprint of life. A 'search-and-replace' method for genome editing takes us a giant leap closer to this ambitious goal. A 'search-and-replace' method for editing the genome., Author(s): Randall J. Platt Author Affiliations: CRISPR tool modifies genes precisely by copying RNA into the genome Variation in the DNA sequences that constitute the blueprint of life is essential [...]