Biology subjects

Smith, K. A.

Publications and source records attributed to Smith, K. A..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Assessing a Role for Sarcospan in Immune Function

Sarcospan (SSPN) is a tetraspanin-like member of the dystrophin-glycoprotein complex (DGC) with expression reported at the transcript level in B cells and several other immune cell populations. SSPN is best known for its expression in skeletal, cardiac, and vascular smooth muscle, however it is expressed in other cell types and its roles in non-muscle cells have not been well described. Analysis of functional interactions of SSPN with effectors of hematopoietic cell lineages uncovered strong associations with several proteins with known roles in hematopoiesis and immune response. In this study, we utilized flow cytometry to uncover SSPN+ immune cell populations expressing the SSPN protein. We performed immunophenotyping of global SSPN-deficient (SSPN-/-) C57BL/6J male and female mice to assess whether SSPN plays an essential role in generation of specific immune cells. By flow cytometry we found that B cells, CD11b+ and CD11c+ (monocytes, tissue macrophages, and a subset of dendritic cells) express the highest levels of SSPN at their cell membranes. To pinpoint potential roles for SSPN in B cell development, SSPN-/- bone marrow cells were isolated and differentiated using phorbol myristate acetate (PMA). To test B cell effector functions, antibody isotype analysis was performed to assess whether SSPN influences antibody class switching. Blood serum of WT and SSPN-/- mice were tested for presence of important developmental signals for B cell maturation, development and isotype switching. Our study is the first to document SSPN protein expression on distinct classes of immune cells including CD11b+, CD11c+ and B cells (CD19+). Immunophenotyping revealed several differences in immune cell populations in the murine spleens: 1) a significant increase in % T cells in SSPN-/- males, 2) slight reduction of % B cells in SSPN-/- females, 3) a significant increase % of dendritic cells in SSPN-/- females, and 4) no significant alterations in NK cells. Under unstimulated conditions, SSPN-/- female mice had a significantly lower IgG2b titers compared to WT. Overall, the observation of SSPN expression in distinct immune cells, potential role(s) in hematopoiesis and B cell effector function is intriguing and warrants further investigation. HighlightsThe sarcospan protein has an established role in stabilization of striated muscle membranes and is an essential member of the dystrophin-glycoprotein complex. This study investigates the reported presence of Sspn in B cells and examines potential roles in immune function including: O_LIDetermining a connection with hematopoiesis and establishment of niches C_LIO_LIImpact on immune content in spleens of sarcospan-deficient mice C_LIO_LIIdentifying a role in B cell effector functions C_LI

immunology↗

bioRxiv2023

Brain-wide Correspondence Between Neuronal Epigenomics and Long-Distance Projections

Single-cell genetic and epigenetic analyses parse the brains billions of neurons into thousands of "cell-type" clusters, each residing in different brain structures. Many of these cell types mediate their unique functions by virtue of targeted long-distance axonal projections to allow interactions between specific cell types. Here we have used Epi-Retro-Seq to link single cell epigenomes and associated cell types to their long-distance projections for 33,034 neurons dissected from 32 different source regions projecting to 24 different targets (225 source [->]target combinations) across the whole mouse brain. We highlight uses of this large data set for interrogating both overarching principles relating projection cell types to their transcriptomic and epigenomic properties and for addressing and developing specific hypotheses about cell types and connections as they relate to genetics. We provide an overall synthesis of the data set with 926 statistical comparisons of the discriminability of neurons projecting to each target for every dissected source region. We integrate this dataset into the larger, annotated BICCN cell type atlas composed of millions of neurons to link projection cell types to consensus clusters. Integration with spatial transcriptomic data further assigns projection-enriched clusters to much smaller source regions than afforded by the original dissections. We exemplify these capabilities by presenting in-depth analyses of neurons with identified projections from the hypothalamus, thalamus, hindbrain, amygdala, and midbrain to provide new insights into the properties of those cell types, including differentially expressed genes, their associated cis-regulatory elements and transcription factor binding motifs, and neurotransmitter usage.

neuroscience↗

bioRxiv2023

Single-cell DNA Methylome and 3D Multi-omic Atlas of the Adult Mouse Brain

Cytosine DNA methylation is essential in brain development and has been implicated in various neurological disorders. A comprehensive understanding of DNA methylation diversity across the entire brain in the context of the brains 3D spatial organization is essential for building a complete molecular atlas of brain cell types and understanding their gene regulatory landscapes. To this end, we employed optimized single-nucleus methylome (snmC-seq3) and multi-omic (snm3C-seq1) sequencing technologies to generate 301,626 methylomes and 176,003 chromatin conformation/methylome joint profiles from 117 dissected regions throughout the adult mouse brain. Using iterative clustering and integrating with companion whole-brain transcriptome and chromatin accessibility datasets, we constructed a methylation-based cell type taxonomy that contains 4,673 cell groups and 261 cross-modality-annotated subclasses. We identified millions of differentially methylated regions (DMRs) across the genome, representing potential gene regulation elements. Notably, we observed spatial cytosine methylation patterns on both genes and regulatory elements in cell types within and across brain regions. Brain-wide multiplexed error-robust fluorescence in situ hybridization (MERFISH2) data validated the association of this spatial epigenetic diversity with transcription and allowed the mapping of the DNA methylation and topology information into anatomical structures more precisely than our dissections. Furthermore, multi-scale chromatin conformation diversities occur in important neuronal genes, highly associated with DNA methylation and transcription changes. Brain-wide cell type comparison allowed us to build a regulatory model for each gene, linking transcription factors, DMRs, chromatin contacts, and downstream genes to establish regulatory networks. Finally, intragenic DNA methylation and chromatin conformation patterns predicted alternative gene isoform expression observed in a companion whole-brain SMART-seq3 dataset. Our study establishes the first brain-wide, single-cell resolution DNA methylome and 3D multi-omic atlas, providing an unparalleled resource for comprehending the mouse brains cellular-spatial and regulatory genome diversity.

genomics↗

bioRxiv2023

A molecularly defined and spatially resolved cell atlas of the whole mouse brain

In mammalian brains, tens of millions to billions of cells form complex interaction networks to enable a wide range of functions. The enormous diversity and intricate organization of cells in the brain have so far hindered our understanding of the molecular and cellular basis of its functions. Recent advances in spatially resolved single-cell transcriptomics have allowed systematic mapping of the spatial organization of molecularly defined cell types in complex tissues1-3. However, these approaches have only been applied to a few brain regions1-11 and a comprehensive cell atlas of the whole brain is still missing. Here, we imaged a panel of >1,100 genes in [~]8 million cells across the entire adult mouse brain using multiplexed error-robust fluorescence in situ hybridization (MERFISH)12 and performed spatially resolved, single-cell expression profiling at the whole-transcriptome scale by integrating MERFISH and single-cell RNA-sequencing (scRNA-seq) data. Using this approach, we generated a comprehensive cell atlas of >5,000 transcriptionally distinct cell clusters, belonging to [~]300 major cell types, in the whole mouse brain with high molecular and spatial resolution. Registration of the MERFISH images to the common coordinate framework (CCF) of the mouse brain further allowed systematic quantifications of the cell composition and organization in individual brain regions defined in the CCF. We further identified spatial modules characterized by distinct cell-type compositions and spatial gradients featuring gradual changes in the gene-expression profiles of cells. Finally, this high-resolution spatial map of cells, with a transcriptome-wide expression profile associated with each cell, allowed us to infer cell-type-specific interactions between several hundred pairs of molecularly defined cell types and predict potential molecular (ligand-receptor) basis and functional implications of these cell-cell interactions. These results provide rich insights into the molecular and cellular architecture of the brain and a valuable resource for future functional investigations of neural circuits and their dysfunction in diseases.

neuroscience↗

bioRxiv2022

An In Vivo Analysis of the Functional Motifs of DEAD-box RNA Helicase Me31B in Drosophila Fertility and Germline Development

In Drosophila germline, Me31B is a putative ATP-dependent, RNA helicase that plays role in post-transcriptional RNA regulation to ensure the correct spatial and temporal expression of the mRNAs, a process crucial for proper germline development and fertility. However, Me31Bs in vivo working mechanism remains unclear. In this study, we aim to analyze the functions of Me31Bs key domains/motifs to understand how these domains/motifs operate to fulfill the proteins overall activities. We generated Drosophila strains mutant for six important motifs including three ATPase/helicase motifs (DEAD-box, DVLARAK, and HRIGR), the N-terminal domain (N-ter), the C-terminal domain (C-ter), and a protein-binding motif (FDF motif-binding motif). In characterizing these mutants, we observed that the three ATPase/helicase motif mutations cause dominant female sterility which is associated with developmental defects in oogenesis and embryogenesis. Follow-up examination of the DVLARAK motif mutant revealed its abnormalities in germline mRNA localization and transcript level. The Me31B N-ter domain (deletion of C-ter), C-ter domain (deletion of N-ter), and mutation of FDF motif-binding motif led to a decrease in female fertility and abnormal subcellular Me31B localizations in the egg chambers. Moreover, deletion of Me31B N-ter or C-ter motif results in a decrease of Me31B protein levels in the ovaries. This study indicates that these six motifs of Me31B play different roles to contribute to Me31Bs whole-protein functions like ATPase, RNA helicase, protein stability, protein localization, and partner protein binding, which are crucial for germline development and fertility. Considering Me31B protein familys conserved presence in both Drosophila germline and soma (for example, neurons) and in other organisms such as yeast, worm, mouse, and human, the results from this study could expand our understanding of Me31B helicase familys general working mechanisms in different cell types and species.

developmental biology↗

bioRxiv2021

Elys deficiency constrains Kras-driven tumour burden by amplifying oncogenic stress

The nucleoporin ELYS, encoded by AHCTF1, is a large multifunctional protein with essential roles in nuclear pore assembly and mitosis. Using a zebrafish model of hepatocellular carcinoma, in which the expression of an inducible mutant kras transgene (krasG12V) drives hepatocyte-specific hyperplasia and liver enlargement, we show that reducing ahctf1 gene dosage by 50% markedly shrinks tumour burden, while non-hyperplastic tissues are unaffected. We demonstrate that ahctf1 heterozygosity impairs nuclear pore formation, mitotic spindle assembly and chromosome segregation, leading to DNA damage and activation of TP53-dependent and independent mechanisms of cell death and cell cycle arrest. This selective vulnerability of cancer cells to mild disruption of Elys function uncovers a novel synthetic lethal interaction between ahctf1 and kras mutations that could be exploited therapeutically. Heterozygous expression of both ahctf1 and ranbp2, or treatment of heterozygous ahctf1 larvae with the nucleocytoplasmic transport inhibitor, Selinexor, completely blocked krasG12V-driven hepatocyte hyperplasia, revealing promising avenues for combinatorial treatments.

cancer biology↗

bioRxiv2021

Cellular resolution anatomical and molecular atlases for prenatal human brains

Increasing interest in studies of prenatal human brain development, particularly using new single-cell genomics and anatomical technologies to create cell atlases, creates a strong need for accurate and detailed anatomical reference atlases. In this study, we present two cellular-resolution digital anatomical atlases for prenatal human brain at post-conceptional weeks (PCW) 15 and 21. Both atlases were annotated on sequential Nissl-stained sections covering brain-wide structures on the basis of combined analysis of cytoarchitecture, acetylcholinesterase staining and an extensive marker gene expression dataset. This high information content dataset allowed reliable and accurate demarcation of developing cortical and subcortical structures and their subdivisions. Furthermore, using the anatomical atlases as a guide, spatial expression of 37 and 5 genes from the brains respectively at PCW 15 and 21 was annotated, illustrating reliable marker genes for many developing brain structures. Finally, the present study uncovered several novel developmental features, such as the lack of an outer subventricular zone in the hippocampal formation and entorhinal cortex, and the apparent extension of both cortical (excitatory) and subcortical (inhibitory) progenitors into the prenatal olfactory bulb. These comprehensive atlases provide useful tools for visualization, targeting, imaging and interpretation of brain structures of prenatal human brain, and for guiding and interpreting the next generation of cell census and connectome studies.

neuroscience↗

bioRxiv2020

Single-cell RNA-seq uncovers shared and distinct axes of variation in dorsal LGN neurons in mice, non-human primates and humans

Abundant anatomical and physiological evidence supports the presence of at least three distinct types of relay glutamatergic neurons in the primate dorsal lateral geniculate nucleus (dLGN) of the thalamus, the brain region that conveys visual information from the retina to the primary visual cortex. Relay neuron diversity has also been described in the mouse dLGN (also known as LGd). Different types of relay neurons in mice, humans and macaques have distinct morphologies, distinct connectivity patterns, and convey different aspects of visual information to the cortex. To investigate the molecular underpinnings of these cell types, and how these relate to other cellular properties and differences in dLGN between human, macaque, and mice, we profiled gene expression in single nuclei and cells using RNA-sequencing. These efforts identified four distinct types of relay neurons in the primate dLGN, magnocellular neurons, parvocellular neurons, and two cell types expressing canonical marker genes for koniocellular neurons. Surprisingly, despite extensive documented morphological and physiological differences between magno- and parvocellular neurons, we identified few genes with significant differential expression between transcriptomic cell types corresponding to these two neuronal populations. We also detected strong donor-specific gene expression signatures in both macaque and human relay neurons. Likewise, the dominant feature of relay neurons of the adult mouse dLGN is high transcriptomic similarity, with an axis of heterogeneity that aligns with core vs. shell portions of mouse dLGN. Together, these data show that transcriptomic differences between principal cell types in the mature mammalian dLGN are subtle relative to striking differences in morphology and cortical projection targets. Finally, we align cellular expression profiles across species and find homologous types of relay neurons in macaque and human, and distinct relay neurons in mouse.

neuroscience↗

bioRxiv2020

The roseoloviruses downregulate the protein tyrosine phosphatase PTPRC(CD45)

Like all herpesviruses, the roseoloviruses (HHV6A, -6B, and -7) establish lifelong infection within their host, requiring these viruses to evade host anti-viral responses. One common host-evasion strategy is the downregulation of host-encoded, surface-expressed glycoproteins. Roseoloviruses have been shown to evade host the host immune response by downregulating NK-activating ligands, MHC class I, and the TCR/CD3 complex. To more globally identify glycoproteins that are differentially expressed on the surface of HHV6A-infected cells, we performed cell surface capture of N-linked glycoproteins present on the surface of T cells infected with HHV6A, and compared these to proteins present on the surface of uninfected T cells. We found that the protein tyrosine phosphatase CD45 is downregulated in T cells infected with HHV6A. We also demonstrated that CD45 is similarly downregulated in cells infected with HHV-7. CD45 is essential for signaling through the T cell receptor and as such, is necessary for developing a fully functional immune response. Interestingly, the closely related {beta}-herpesviruses human cytomegalovirus (HCMV) and murine cytomegalovirus (MCMV) have also separately evolved unique mechanisms to target CD45. While HCMV and MCMV target CD45 signaling and trafficking, HHV6A acts to downregulate CD45 transcripts. ImportanceHuman herpesviruses-6 and -7 infect essentially 100% of the worlds population before the age of 5 and then remain latent or persistent in their host throughout life. As such, these viruses are among the most pervasive and stealthy of all viruses. Host immune cells rely on the presence of surface-expressed proteins to identify and target virus-infected cells. Here, we investigated the changes that occur to proteins expressed on the cell surface of T cells after infection with human herpesvirus-6A. We discovered that HHV-6A infection results in a reduction of CD45 on the surface of infected cells. Targeting of CD45 may prevent activation of these virus-infected T cells, possibly lengthening the life of the infected T cell so that it can harbor latent virus.

microbiology↗

bioRxiv2020

Regional, layer, and cell-class specific connectivity of the mouse default mode network

The evolutionarily conserved default mode network (DMN) is characterized by temporally correlated activity between brain regions during resting states. The DMN has emerged as a selectively vulnerable network in multiple disorders, so understanding its anatomical composition will provide fundamental insight into how its function is impacted by disease. Reproducible rodent analogs of the human DMN offer an opportunity to investigate the underlying brain regions and structural connectivity (SC) with high spatial and cell type resolution. Here, we performed systematic analyses using mouse resting state functional magnetic resonance imaging to identify the DMN and whole brain axonal tracing data, co-registered to the 3D Allen Mouse Common Coordinate Framework reference atlas. We identified the specific, predominantly cortical, brain regions comprising the mouse DMN and report preferential SC between these regions. Next, at the cell class level, we report that cortical layer (L) 2/3 neurons in DMN regions project almost exclusively to other DMN regions, whereas L5 neurons project to targets both in and out of the DMN. We then test the hypothesis that in- and out-DMN projection patterns originate from distinct L5 neuron sub-classes using an intersectional viral tracing strategy to label all the axons from neurons defined by a single target. In the ventral retrosplenial cortex, a core DMN region, we found two L5 projection types related to the DMN and mapped them to unique transcriptomically-defined cell types. Together, our results provide a multi-scale description of the anatomical correlates of the mouse DMN.

neuroscience↗

bioRxiv2020

Identifying the spatial and temporal dynamics of molecularly-distinct glioblastoma sub-populations

Glioblastomas (GBMs) are the most aggressive primary brain tumours and have no known cure. Each individual tumour comprises multiple sub-populations of genetically-distinct cells that may respond differently to targeted therapies and may contribute to disappointing clinical trial results. Image-localized biopsy techniques allow multiple biopsies to be taken during surgery and provide information that identifies regions where particular sub-populations occur within an individual GBM, thus providing insight into their regional genetic variability. These sub-populations may also interact with one another through a competitive or cooperative nature; it is important to ascertain the nature of these interactions, as they may have implications for responses to targeted therapies. We combine genetic information from biopsies with a mechanistic model of interacting GBM sub-populations to characterise the nature of interactions between two commonly occurring GBM sub-populations, those with EGFR and PDGFRA genes amplified. We study population levels found across image-localized biopsy data from a cohort of 25 patients and compare this to model outputs under competitive, cooperative and neutral interaction assumptions. We explore other factors affecting the observed simulated sub-populations, such as selection advantages and phylogenetic ordering of mutations, which may also contribute to the levels of EGFR and PDGFRA amplified populations observed in biopsy data.

cancer biology↗

bioRxiv2020

A taxonomy of transcriptomic cell types across the isocortex and hippocampal formation

The isocortex and hippocampal formation are two major structures in the mammalian brain that play critical roles in perception, cognition, emotion and learning. Both structures contain multiple regions, for many of which the cellular composition is still poorly understood. In this study, we used two complementary single-cell RNA-sequencing approaches, SMART-Seq and 10x, to profile [~]1.2 million cells covering all regions in the adult mouse isocortex and hippocampal formation, and derived a cell type taxonomy comprising 379 transcriptomic types. The completeness of coverage enabled us to define gene expression variations across the entire spatial landscape without significant gaps. We found that cell types are organized in a hierarchical manner and exhibit varying degrees of discrete or continuous relatedness with each other. Such molecular relationships correlate strongly with the spatial distribution patterns of the cell types, which can be region-specific, or shared across multiple regions, or part of one or more gradients along with other cell types. Glutamatergic neuron types have much greater diversity than GABAergic neuron types, both molecularly and spatially, and they define regional identities as well as inter-region relationships. For example, we found that glutamatergic cell types between the isocortex and hippocampal formation are highly distinct from each other yet possess shared molecular signatures and corresponding layer specificities, indicating their homologous relationships. Overall, our study establishes a molecular architecture of the mammalian isocortex and hippocampal formation for the first time, and begins to shed light on its underlying relationship with the development, evolution, connectivity and function of these two brain structures.

neuroscience↗

bioRxiv2019

Distinct transcriptomic cell types and neural circuits of the subiculum and prosubiculum along the dorsal-ventral axis

Subicular region plays important roles in spatial processing and many cognitive functions and these were mainly attributed to subiculum (Sub) rather than prosubiculum (PS). Using single-cell RNA-sequencing (scRNA-seq) technique we have identified up to 27 distinct transcriptomic clusters/cell types, which were registered to anatomical sub-domains in Sub and PS. Based on reliable molecular markers derived from transcriptomic clustering and in situ hybridization data, the precise boundaries of Sub and PS have been consistently defined along the dorsoventral (DV) axis. Using these borders to evaluate Cre-line specificity and tracer injections, we have found bona fide Sub projections topographically to structures important for spatial processing and navigation. In contrast, PS along DV axis sends its outputs to widespread brain regions crucial for motivation, emotion, reward, stress, anxiety and fear. Brain-wide cell-type specific projections of Sub and PS have also been revealed using specific Cre-lines. These results reveal two molecularly and anatomically distinct circuits centered in Sub and PS, respectively, providing a consistent explanation to historical data and a clearer foundation for future functional studies. HighlightsO_LI27 transcriptomic cell types identified in and spatially registered to "subicular" regions. C_LIO_LIAnatomic borders of "subicular" regions reliably determined along dorsal-ventral axis. C_LIO_LIDistinct cell types and circuits of full-length subiculum (Sub) and prosubiculum (PS). C_LIO_LIBrain-wide cell-type specific projections of Sub and PS revealed with specific Cre-lines. C_LI In BriefDing et al. show that mouse subiculum and prosubiculum are two distinct regions with differential transcriptomic cell types, subtypes, neural circuits and functional correlation. The former has obvious topographic projections to its main targets while the latter exhibits widespread projections to many subcortical regions associated with reward, emotion, stress and motivation.

neuroscience↗

bioRxiv2019

Endocardial differentiation in zebrafish occurs during early somitogenesis and is dependent on BMP and etv2 signalling

Abstract The endocardium and adjacent vascular endothelial network share a number of molecular markers however there are distinct physiological functions of these tissues. What distinguishes these lineages on a molecular level remains an important, unanswered question in cardiovascular biology. We have identified the Gt(SAGFF27C); Tg(4xUAS:egfp) line as a marker of early endocardial development and used this line to examine endocardial differentiation. Our results show that the endocardium emerges from the anterior lateral plate mesoderm at the 8-somite stage (13 hpf). Analysis in a number of loss-of-function models showed that whilst nkx2.5, hand2 and tal1 loss-of-function have no effect on the endocardial progenitor domain, both etv2 loss-of-function and inhibition of BMP signalling reduce the endocardial domain. Furthermore, manipulating BMP signalling alters etv2 expression. Together, these results describe the onset of endocardial molecular identity and suggest a signalling cascade whereby BMP signalling acts upstream of etv2 to direct differentiation of endocardial progenitors.

developmental biology↗

bioRxiv2019

Transcriptomic evidence that von Economo neurons are regionally specialized extratelencephalic-projecting excitatory neurons

von Economo neurons (VENs) are bipolar, spindle-shaped neurons restricted to layer 5 of human frontoinsula and anterior cingulate cortex that appear to be selectively vulnerable to neuropsychiatric and neurodegenerative diseases, although little is known about other VEN cellular phenotypes. Single nucleus RNA-sequencing of frontoinsula layer 5 identified a transcriptomically-defined cell cluster that contained VENs, but also fork cells and a subset of pyramidal neurons. Cross-species alignment of this cell cluster with a well-annotated mouse classification shows strong homology to extratelencephalic (ET) excitatory neurons that project to subcerebral targets. This cluster also shows strong homology to a putative ET cluster in human temporal cortex, but with a strikingly specific regional signature. Together these results predict VENs are a regionally distinctive type of ET neuron, and we additionally describe the first patch clamp recordings of VENs from neurosurgically-resected tissue that show distinctive intrinsic membrane properties relative to neighboring pyramidal neurons.

neuroscience↗

bioRxiv2019

Epigenetic landscape and AAV targeting of human neocortical cell classes

Viral genetic tools to target specific brain cell types in humans and non-genetic model organisms will transform basic neuroscience and targeted gene therapy. Here we used comparative epigenetics to identify thousands of human neuronal subclass-specific putative enhancers to regulate viral tools, and 34% of these were conserved in mouse. We established an AAV platform to evaluate cellular specificity of functional enhancers by multiplexed fluorescent in situ hybridization (FISH) and single cell RNA sequencing. Initial testing in mouse neocortex yields a functional enhancer discovery success rate of over 30%. We identify enhancers with specificity for excitatory and inhibitory classes and subclasses including PVALB, LAMP5, and VIP/LAMP5 cells, some of which maintain specificity in vivo or ex vivo in monkey and human neocortex. Finally, functional enhancers can be proximal or distal to cellular marker genes, conserved or divergent across species, and could yield brain-wide specificity greater than the most selective marker genes.

neuroscience↗

bioRxiv2019

Prospective, brain-wide labeling of neuronal subclasses with enhancer-driven AAVs

The rapid pace of cell type identification by new single-cell analysis methods has not been met with efficient experimental access to the newly discovered types. To enable flexible and efficient access to specific neural populations in the mouse cortex, we collected chromatin accessibility data from individual cells and clustered the single-cell data to identify enhancers specific for cell classes and subclasses. When cloned into adeno-associated viruses (AAVs) and delivered to the brain by retro-orbital injections, these enhancers drive transgene expression in specific cell subclasses in the cortex. We characterize several enhancer viruses in detail to show that they result in labeling of different projection neuron subclasses in mouse cortex, and that one of them can be used to label the homologous projection neuron subclass in human cortical slices. To enable the combinatorial labeling of more than one cell type by enhancer viruses, we developed a three-color Cre-, Flp- and Nigri-recombinase dependent reporter mouse line, Ai213. The delivery of three enhancer viruses driving these recombinases via a single retroorbital injection into a single Ai213 transgenic mouse results in labeling of three different neuronal classes/subclasses in the same brain tissue. This approach combines unprecedented flexibility with specificity for investigation of cell types in the mouse brain and beyond.

neuroscience↗

bioRxiv2018

Conserved cell types with divergent features between human and mouse cortex

Elucidating the cellular architecture of the human neocortex is central to understanding our cognitive abilities and susceptibility to disease. Here we applied single nucleus RNA-sequencing to perform a comprehensive analysis of cell types in the middle temporal gyrus of human cerebral cortex. We identify a highly diverse set of excitatory and inhibitory neuronal types that are mostly sparse, with excitatory types being less layer-restricted than expected. Comparison to a similar mouse cortex single cell RNA-sequencing dataset revealed a surprisingly well-conserved cellular architecture that enables matching of homologous types and predictions of human cell type properties. Despite this general conservation, we also find extensive differences between homologous human and mouse cell types, including dramatic alterations in proportions, laminar distributions, gene expression, and morphology. These species-specific features emphasize the importance of directly studying human brain.

neuroscience↗