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Zachary Smith, PhD

Assistant Professor of Genetics

Contact Information

Zachary Smith, PhD

Mailing Address

  • Yale School of Medicine

    Yale Stem Cell Center, 10 Amistad Street

    New Haven, CT 06519

    United States

Research Summary

Our overarching goal is to understand moments in developmental time where the genome undergoes dramatic changes in regulation, as well as their purpose and underlying molecular machinery. We are specifically interested in how covalent modifications to chromatin are coordinated to control organismal phenotype epigenetically, including possible impacts of the fetal environment. To study these phenomena, we apply a variety of cutting-edge technological approaches to measure changes in genome regulation as well as advanced micromanipulation techniques to perturb native processes within the early mouse embryo. Our efforts are focused on the first three major transitions in mammalian embryogenesis: fertilization, implantation, and gastrulation.

Extensive Research Description

During fertilization, a full meter of genetic material is delivered by sperm as a highly dense, protamine-compacted particle and rapidly outfitted with chromatin over the first cell cycle. During this period, this formerly inert genome must be activated and made competent for essentially every life process, including transcription, replication, and mitosis. Simultaneously, pre-existing epigenetic modifications within the paternal genome, primarily cytosine methylation, are globally erased. Failure to appropriately reconstruct a functioning genome may also have downstream consequences that affect embryo viability or long term phenotypes. Nonetheless, the exact sequences and molecular pathways that govern these processes are still only minimally understood and hampered by the delicacy and transience of the single cell embryo. We seek to apply a suite of next generation sequencing approaches and advanced micromanipulation techniques to specifically measure and perturb the fertilization process to understand the very first moments of life.

The maternal-fetal interface is established during implantation. During this process, mammalian genomes undergo two distinct waves of global genome reprogramming depending upon their ultimate developmental lineage. Within the embryonic lineage, the pre-implantation inner cell mass (ICM) matures to form the epiblast, which remains capable of forming all subsequent tissues but appears to have fundamentally different genome regulation. This transition (frequently referred to as “naïve to primed”) includes the initial establishment of a chromatin landscape that resembles all subsequent somatic cells in the body and is characterized by high global levels of cytosine methylation and regulation of developmnental gene promoters by the Polycomb Repressive Complexes.

Simultaneously, the pre-implantation trophectodermal lineage matures to form the Extraembryonic Ectoderm (ExE) which will go on to invade maternal tissues and form the placenta. As it develops, the ExE aquires a highly unusual mode of genome regulation characterized by an erratic, highly dynamic DNA methylation landscape that includes terminal silencing of Polycomb targets. While unobserved within the embryonic lineage, a highly similar mode of genome regulation emerges in nearly every observed mammalian cancer, regardless of its mutational profile or tissue of origin. We are interested in understanding how these two highly distinct modes of genome regulation are established and employ a variety of approaches to study the epigenetics and genome biology of implantation in vivo and in vitro. For example, we employ Cas9-mediated genome editing to perturb key epigenetic regulators in zygotes and examine changes to the epigenome or transcriptome in early post-implantation tissues. Similarly, we hope to develop novel massive parallel reporter assays (MPRA) to identify how specific epigenetic states are initiated and maintained.

After implantation, the pluripotent epiblast is triggered to differentiate by ExE-supported morphogen-gradients to initiate gastrulation. Although the initial waves of mammalian development require approximately one week, the establishment of the three germ layers (ectoderm, mesoderm, and endoderm) and foundational embryonic body axes is comparatively rapid. Within two days, the mouse embryo proceeds from a fairly rudimentary conceptus comprised of several hundred cells to a highly sophisticated structure with dozens of unique cell types and well over 100,000 cells. Notably, many chromatin regulators are absolutely essential for this process and produce embryonic lethal knockout phenotypes within the gastrulation window. However, it is exceedingly unclear how this class of regulators, which are constitutively expressed and recognize highly generic substrates, help orchestrate such intricate and highly specific developmental outcomes. To study this seeming paradox, we have innovated a novel platform that combines zygotic Cas9-based mutation and single cell transcriptional analysis to examine cohorts of complex mutant embryos. Using this approach, we hope to develop quantitative models that explain the specific roles of epigenetic regulators in gastrulation. Moreover, we hope to expand on these principles to understand sources of phenotypic variation, particularly environmental factors that may alter the epigenome and result in fetal or congenital disorders.

Finally, we are constantly seeking to develop new technological approaches for studying these processes. For example, we recently helped innovate a novel molecular recorder to capture historical information between thousands of single cells within the post-gastrulation embryo. We hope to utilize this strategy to quantify progenitor field dynamics, the coordinated creation and consumption of multipotent progenitor cells to generate embryonic structures. We are continuing to optimize this technology to improve its resolution and reproducibility. In parallel, we hope to apply it to understand complex developmental transitions, as well as to reconstruct cell lineages within normal and experimentally perturbed cell lineages.

Research Interests

Biotechnology; Cell Nucleus; Chromatin; Embryonic and Fetal Development; Genetics; Molecular Biology; Reproduction; Epigenetic Repression

Selected Publications