Single Cell Transcriptomics Classification

PI: John J. NgaiNgai Lab
University of California Berkeley
Title:
“Classification of Cortical Neurons by Single Cell Transcriptomics”
BRAIN Category: Census of Cell Types (RFA MH-14-215)

To understand what makes neurons distinct, Dr. Ngai’s team will explore one major type of mouse brain cell, pinpointing genes responsible for differentiating them into subtypes and will also test whether each subtype has unique functions, using a new technique that labels them with tagged genes.

NIH Webpage

Ngai Lab BRAIN Initiative project represents a multidisciplinary collaboration between 10 research groups at UC Berkeley.

Ngai Lab BRAIN Initiative project represents a multidisciplinary collaboration between 10 research groups at UC Berkeley.

Project Description

Unraveling the complexity of the mammalian brain is one of the most challenging problems in biology today. A major goal of neuroscience is to understand how circuits of neurons and non-neuronal cells process sensory information, generate movement, and subserve memory, emotion and cognition. Elucidating the properties of neural circuits requires an understanding of the cell types that comprise these circuits and their roles in processing and integrating information. However, since the description of diverse neuronal cell types over a century ago by Ramon y Cajal, we have barely scratched the surface of understanding the diversity of cell types in the brain and how each individual cell type contributes to nervous system function. Current approaches for classifying neurons rely upon features including the differential expression of small numbers of genes, cell morphology, anatomical location, physiology, and connectivity – important descriptive properties that nonetheless are insufficient to fully describe or predict the vast number of different cell types that comprise the mammalian brain. Here we propose a suite of technologies for identifying and classifying the myriad cell types present in the brain. Our method will be developed using layer 5 pyramidal cells from mouse somatosensory cortex as a model system. First, we will exploit the latest developments in DNA sequencing technologies to characterize gene expression profiles on single layer 5 neurons at high throughput. This information will be used to classify individual cells based on their transcriptome “fingerprints.” Second, genes found to define newly discovered neuronal subtypes will be used to gain genetic access to these cells using Cas9/CRISPR-mediated genome engineering to create transgenic reporter lines. Development of this technology promises to open a pipeline for the rapid generation of multigenic mouse reporter strains in which specific neuronal subtypes are uniquely labeled by combinations of tagged genes. Third, we will use these genetically engineered mice to confirm that our taxonomy represents distinct functional properties of the classified neurons. Our approach can ultimately be scaled up to generate a complete census of cell types in the brain, a critically needed resource for dissecting nervous system function with modern investigative tools.

From Ngai Lab webpage

A major goal of neuroscience is to understand how circuits of neurons and non-neuronal cells process sensory information, generate movement, and subserve memory, emotion and cognition. Elucidating the properties of neural circuits requires an understanding of the cell types that comprise these circuits and their roles in processing and integrating information. However, since the description of diverse neuronal cell types over a century ago by Ramon y Cajal, we have barely scratched the surface of understanding the diversity of cell types in the brain and how each individual cell type contributes to nervous system function. Current approaches for classifying neurons rely upon features including the differential expression of small numbers of genes, cell morphology, anatomical location, physiology, and connectivity – important descriptive properties that nonetheless are insufficient to fully describe or predict the vast number of different cell types that comprise the mammalian brain. This NIH-supported BRAIN Initiative project aims to provide a suite of technologies for identifying and classifying the diverse cell types in the mammalian nervous system. We are developing our method using layer 5 pyramidal cells from mouse somatosensory cortex as a model system.

First, we will exploit the latest developments in DNA sequencing technologies to characterize gene expression profiles on single layer 5 neurons at high throughput. This information will be used to classify individual cells based on their transcriptome “fingerprints.”

Second, genes found to define newly discovered neuronal subtypes will be used to gain genetic access to these cells using Cas9/CRISPR-mediated genome engineering to create transgenic reporter lines. Development of this technology promises to open a pipeline for the rapid generation of multigenic mouse reporter strains in which specific neuronal subtypes are uniquely labeled by combinations of tagged genes.

Third, we will use these genetically engineered mice to confirm that our taxonomy represents distinct functional properties of the classified neurons.

Our approach can ultimately be scaled up to generate a complete census of cell types in the brain, a critically needed resource for dissecting nervous system function with modern investigative tools.

The Ngai Lab BRAIN Initiative project represents a multidisciplinary collaboration between 10 research groups at UC Berkeley working in the following areas:

Genomics

·       John Ngai – PI (MCB)

Neurobiology

·       Hillel Adesnik (MCB)

·       Helen Bateup (MCB)

·       Dan Feldman (MCB)

Genome Engineering and Mouse Transgenesis

·       Jennifer Doudna (MCB, Chemistry, HHMI, LBNL)

·       Dirk Hockemeyer (MCB)

·       Russell Vance (MCB, HHMI)

Statistics and Bioinformatics

·       Sandrine Dudoit (Biostatistics)

·       Elizabeth Purdom (Statistics)

·       Nir Yosef (Computer Science)

Public Health Relevance Statement

The human brain contains billions of neurons which in turn are thought to comprise hundreds if not thousands of distinct cell types, each tailored for a specific functional role in the processing of sensory information, generation of motor output, and for providing the biological basis of memory, behavior and consciousness. Current technologies so far have been unable to provide a detailed census of the myriad neuronal cell types in the mammalian brain, information that is critical for providing tools for unraveling and probing the computational logic of the brain. This BRAIN Initiative application proposes a suite of cutting edge technologies – including single cell transcriptome profiling and genome engineering – that can be developed and scaled up to catalog all of the neuronal cell types in the mammalian brain.

NIH Spending Category

Genetics; Human Genome; Mental Health; Neurosciences

Project Terms

Anatomy; Atlases; base; Base of the Brain; Behavior; Biological; Biological Models; Biology; Brain; Brain Mapping; Brain region; Cataloging; Catalogs; Cell physiology; Cell Separation; cell type; Cells; Cellular Morphology; Censuses; Classification; Classification Scheme; Clustered Regularly Interspaced Short Palindromic Repeats; Cognition; Complement; Complex; Conscious; Development; DNA Sequence; Emotions; Equipment and supply inventories; Fingerprint; Future; Gene Expression; Gene Expression Profile; Generations; Genes; Genetic; Genetic Techniques; Genetically Engineered Mouse; Genome; Genome engineering; Genotype; Goals; hippocampal pyramidal neuron; Human; In Situ; Individual; insight; Label; Location; Logic; Maps; Mediating; Memory; Methods; Molecular; molecular marker; Molecular Profiling; Morphology; Motor output; Mouse Strains; Movement; Mus; Nervous System Physiology; neural circuit; Neuroglia; Neurons; Neurosciences; optogenetics; Organ; Organism; Pattern; Physiological; Physiology; Population; Process; Property; Proteins; public health relevance; Pyramidal Cells; recombinase; Reporter; Research; research study; Resources; RNA; Role; Sampling; scale up; Sensory; Sensory Process; Somatosensory Cortex; Staging; Statistical Methods; Surface; Taxonomy; Technology; technology development; Testing; Tissues; tool; transcriptomics; Transgenic Mice; Transgenic Organisms; Validation

 

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