Much of what distinguishes the vertebrates from their invertebrate relatives is a small group of embryonic cells known as the neural crest. In all vertebrate embryos the neural crest gives rise to a dazzling array of cell types and structures, including cartilage and bone of the head, face and neck, many of the sensory neurons throughout the head and trunk, striking patterns of pigmentation, the covering of teeth, major vessels that circulate blood from the heart, and much more. Despite their importance, the evolutionary origins of neural crest cells have remained enigmatic. My dissertation research is structured to address a set of fundamental questions concerning the evolutionary origins of this important cell type. To this end, I focused on deciphering the molecular, cellular and genetic features underpinning neural crest embryology in lampreys, a group of primitively jawless vertebrates that are derived from some of the earliest branching lineage of vertebrate animals. I then compared these mechanisms in lampreys with similar ones in jawed vertebrates. Because all vertebrates are either jawed or jawless, this comparative embryology method allows one to infer the nature of neural crest cells in the last common ancestor of these two vertebrate groups—in this case, the last common ancestor of all vertebrates. This method not only provides insights into early vertebrate history, but also tells us how changes in neural crest cell function over time could have driven major evolutionary transitions in the vertebrate lineage. My dissertation is divided into six chapters (summarized below) that discuss and address fundamental, unresolved issues in the field of neural crest and vertebrate development and evolution. These include: the origin and evolution of vertebrates and vertebrate neural crest cells (chapter 1), mechanisms of neural crest cell migration (chapters 2, 3); evolution of developmental mechanisms that guide and organize neural crest cells into anatomical structures and organs (chapter 4); evolution of neural crest gene function (chapter 5); and the origins of the neural crest cell population in early vertebrates (chapter 6). Taken together, my work provides important insights into the evolution of neural crest cells and the nature of our earliest vertebrate ancestors. In chapter 1, I review ideas on the evolutionary origins of the vertebrates, vertebrate neural crest cells, and the neural crest gene regulatory network. I then discuss how comparative embryology between jawed and jawless vertebrate systems, coupled with modern molecular genetic tools such as CRISPR/Cas9, can be leveraged to address long-standing hypotheses related to early vertebrate evolution. One of the most striking features of neural crest cells is their ability to embark on long- distance migration throughout vertebrate embryos. However, the evolutionary-genetic basis for this migratory feat is unknown. In chapter 2, I survey the neural crest literature and describe cellular and genetic features controlling migration that are common to neural crest cells in all vertebrates. I then describe similar features operating in a variety of migratory cells in invertebrates and propose that neural crest cells share a common molecular genetic “signature” with several other migratory cell types. This new synthesis predicts that neural crest cells evolved their impressive migratory capabilities by activating a core genetic toolkit for cell migration that originated in the last common ancestor of all animals. In chapter 3, I explore the evolutionary origin of genetic mechanisms controlling neural crest migration in vertebrates. To this end, I analyze in lamprey the embryonic function of a gene called Snail, which initiates the earliest stages of neural crest migration in jawed vertebrates. I show that lampreys use a fundamentally different mechanism to initially detach neural crest cells from the neural tube before migrating, but that Snail gene activity is still required start the physical process of cell migration. This work is important because it sheds light on the ancestral nature of neural crest migratory mechanisms in the first vertebrates. Neural crest cells migrate extensively throughout vertebrate embryos, but how did this population of cells become organized into the structures and organ systems that have made the vertebrates an evolutionary success? In chapter 4, I show that the appearance of a new cellular communication system known as Sema3F-Nrp was a pivotal event in early vertebrate and neural crest evolution. This cellular system is active in lamprey embryos when neural crest cells are migrating and gradually sculpts specific groups of neural crest cells into key anatomical structures and organs in the lamprey head (e.g., head skeleton, sensory nerve cells). Based on similarities with other vertebrates, I propose that Sema3F-Nrp evolved in the first lineage of vertebrates. The origins of this cell-cell communication system allowed our early vertebrate ancestors to organize and pattern neural crest cells for the first time into entirely new structures and may have been an important mechanism for continually generating evolutionary change in the vertebrate body during the past 500 million years. A major challenge in the field of neural crest biology is to identify how genes in invertebrates, which lack neural crest cells, evolved new roles for neural crest development in vertebrates. For example, the gene Snail is a key regulator of neural crest cells in jawed vertebrates and CNS neurons among invertebrates, but the fact that these two very different cell populations both use Snail genes is thought to be purely coincidental, rather than suggestive of a common evolutionary origin. In chapter 5, I show in lamprey—a jawless vertebrate spanning the invertebrate-jawed vertebrate divide—that the Snail gene regulates lamprey neural crest development as in other vertebrates, but also regulates CNS development similar to invertebrates. Thus, lampreys seem to bridge the evolutionary-genetic gap between invertebrates and jawed vertebrates by using Snail for the simultaneous development of both of these cell populations. This study provides evidence that the genetic control of neural crest development by Snail genes in vertebrates likely evolved from an ancient function for CNS development among invertebrates. In chapter 6, I address the evolutionary origins of the neural crest cell population and its migratory properties. The paradigm in the field of neural crest biology for the past 150 years has been that these cells form within and then migrate from the dorsal part of the embryonic neural tube. Because this process occurs the same way in all vertebrate embryos studied to date it is also thought to be an accurate reflection of what neural crest cells were like in our earliest vertebrate ancestors. In contrast to this paradigm, I demonstrate in lampreys that multiple transcriptional regulators of neural crest identity are expressed throughout much of the entire embryonic neural tube, not just the dorsal-most region. Using cell lineage tracing and live microscopic imaging, I show that neural crest cells expressing these genes can migrate from almost any position along the neural tube dorsal-ventral axis. In light of these findings, I propose a new evolutionary model in which the first vertebrate neural crest cells formed within and migrated from almost any position in the embryonic neural tube. This new model suggests that the neural crest that forms only in the dorsal neural tube of jawed vertebrates is not an ancestral vertebrate feature as has been thought, but should instead be viewed as a relatively recent evolutionary innovation.