High-throughput imaging of 3D chromatin regulation events in the nervous system

Project Summary/Abstract
Understanding the mechanisms of gene and chromatin regulation and their roles within a
multicellular organism has relevance across many disciples such as synthetic biology, medicine,
developmental biology and neuroscience 1–3. Large-scale efforts of the genomics community
have identified many of the functional genes and gene regulatory elements (GREs) including
recent atlases with the specific expression of genes and their putative regulatory regions within
different cell types of complex tissues 4,5. However, it remains unclear how the 3D organization
of chromatin impacts gene regulation and vice versa. To build a mechanistic understanding of
the interplay between chromatin organization and gene regulation, we would ideally
simultaneously measure all the key elements - DNA sequences, regulatory proteins, and the
transcribed RNA - at the genomic-scale, while maintaining information about cell type identity.
To address this challenge, I will develop an imaging platform that can simultaneously measure
the 3D structure of DNA together with the RNA expression of the regulated genes and their
interaction with key structural proteins (Aim 1). While this method can be applied to many
systems, a particularly suited example is the peripheral olfactory system. Olfaction, one of the
main mammalian senses, is controlled by the largest family of genes comprising more than
1000 olfactory receptors 6,7. Large networks of regulatory sequences interact across the
genome to establish more than 1000 neuronal types, each expressing one and only one
receptor8. I will apply this imaging method to address the longstanding question: how do
different olfactory sensory neurons establish their receptor expression? These integrated
measurements relating chromatin organization and regulatory protein structures to
transcriptional activity will provide a model of olfactory gene regulation. Aim 2, is to dissect this
model and the roles of GRE-promoter interactions in achieving cell-type specific expression
using a high-throughput synthetic biology approach. I will infect the olfactory epithelium with
large pools of viral vectors that combine different regulatory elements and promoters, and
determine the precise cell-type expression of these vectors using multiplexed imaging. There is
an additional synergy between the two aims - the first aim provides measurements of the
endogenous chromatin structure-transcription relationship which will be used to design
transgenic control of specific subpopulation of cells. I will explore this capability to
activate/inhibit specific sub-populations of olfactory receptor neurons and determine the
behavior consequences of these manipulations.

Grant Number: 5DP5OD031878-05
NIH Institute/Center: NIH
Principal Investigator: Bogdan Bintu