Molecular Analysis of primary cilia proteins in human development

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The goal of the research in my laboratory is to study the genetic basis of human craniofacial and CNS malformations. Our efforts in both human and mouse genetics over the past several years have continually directed us towards the primary cilium as a critical hub in signaling for human health and disease. Ciliopathies are diseases associated with both severe congenital malformations as well as nonlethal craniofacial dysmorphology, intellectual disability and obesity (among other conditions). It is clear from the literature that modifying loci are crucial components in understanding much of human disease, but is especially true of the ciliopathies. The focus of this proposal is largely on the primary cilia gene tetratricopeptide repeat domain 21B (Ttc21b). Ttc21b homozygous mouse mutants have several striking features on their own but our preliminary data and the work of others clearly show that TTC21B is a hub in a human ciliopathy network. We have taken four of these candidate interactions from human genetics and recreated them in mouse. All four genes interact with Ttc21b but the cellular and mechanisms of the resulting phenotypes are not yet elucidated. We have also identified multiple novel interacting loci with a combination of ENU mutagenesis and a QTL analysis of the genetic background effects on the severity of the microcephaly phenotype. Thus, we have significant experience in the field and have identified four crucial gaps in knowledge we will address with the support of this MIRA award: 1) How does Ttc21b have such tissue specific effects on organ physiology and developmental signaling, 2) What are the genetic interactors and modifiers of Ttc21b which alter these ciliopathy phenotypes, 3) what is the cellular function of Ttc21b inside the primary cilium, 4) what is the role of Ttc21b outside the cilium. We will use a combination of genetics, molecular embryology, cell biology and biochemistry to address these topics. Many of the ciliary genes are identified to have roles within the primary cilium, but any function outside the cilium has not been elucidated. Identification of such roles would have very a significant effect on the field. Our favorite hypothesis based on preliminary data is that Ttc21b has significant roles in neuronal trafficking. It is clear that a better understanding of human disease will require knowledge of modifying loci and the underlying mechanism(s). Ttc21b is ripe for exploration as a crucial component in a ciliopathy genetic network. A combination of mouse embryology and cell biology inspired by human genomics is an ideal entry point. Our work is likely to not only contribute to knowledge about ciliopathies but point the way forward as a general experimental paradigm for a number of different pathophysiological contexts.