Polyclonal architecture of the mammalian head

Doctoral thesis English OPEN
Lapage, John M. J.
  • Subject: QP

While much of modern developmental biology has focussed upon molecularly defined cell populations, relatively little is understood about how clonal groups within these broad cell populations organise complex tissues. In this thesis, I explore the clonal architecture of the jaws and teeth, and of the dermal bones of the calvaria, revealing cryptic modules as novel developmental features in both. I combine Confetti multicolour genetic lineage labelling with novel analytical techniques in order to map clonal populations in 3D and provide quantitative parameters for clonal expansions.\ud \ud Tooth identity within the mandible is thought to be encoded by the an initial proximodistal position within the branchial arch, which implies that cells do not undergo migration. I observe distal Hand2-Cre labelled cells in the proximal territory, which necessitates migration. These distal cells give rise to the mandible, alveolar bone and a small proportion of odontoblasts, while unlabelled proximal cells were found in the distal territory of the incisors in different proportions. The clonal composition of teeth and jaw bones is dissected by novel analysis of mixed cell populations. I find odontogenic and alveolar bone populations to share a common lineage, comprising a cryptic developmental unit distinct from the mandible, a feature that I can also verify in another transgenic for the upper jaw. I also find that the initial tooth composition radically changes in ontogenetic time. Starting from similar compositions of distal and proximal cells I find that in incisors the distal population expands while the proximal wanes, while in molars the opposite occurs. This is the first evidence for a temporally changing cell population structure underlying the well defined heterodonty between incisors and molars and allows a reinterpretation of early tooth specification events.\ud \ud The dermal bones of the calvaria are thought to grow in thickness by static osteoblasts depositing matrix appositionally and growth is supposed to occur exclusively at sutures. Whole calvaria single-cell clonal lineage analysis of cranial neural crest cells with Wnt1-Cre and Confetti labelling reveals an extensively dynamic program of invasive growth distributed throughout all parts of the dermal bone. Cryptic clonal modules grow laterally, with invasion through and into the bone primarily organised around the centres of these `patches'. The process of bone maturation is revealed to consist of a series of invasions between the three layers of the bone, with the innermost compacta layer driving initial thickness growth by invasion into the middle spongy layer, and the outermost (dermis-adjacent) compacta layer driving later growth. Conversely there is no evidence to suggest that the sutures are principal generative regions, as they do not share a common clonal lineage with adjacent bone. I also investigate muscle attachment regions and find that the same clones traverse tissue boundaries from bone into muscle connective tissues, thus a clonal model predicated on joint 'attachment point precursor cells' can now explain patterns of skeletomuscular connectivity previously found at the population level by my supervisor. A novel generative model of bone growth from cryptic clonal patch modules is proposed, allowing us for the first time to understand thickness growth and the evolutionary transition from a micromeric to a macromeric dermal bone condition, events first visible in crown gnathostomes (placoderms).
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