Honors & Awards
Albert J. Ryan fellowship, Harvard University (2011)
Edward C. Horn Memorial Prize for Excellence in Biology, Duke University (2008)
Graduation with High Distinction in Biology, Duke University (2008)
Summa Cum Laude, Duke University (2008)
American Foundation for Aging Research (AFAR) Scholarship, AFAR (2007)
Phi Beta Kappa Honor Society, Duke University (2007)
Doctor of Philosophy, Harvard University (2014)
Bachelor of Science, Duke University (2008)
Current Research and Scholarly Interests
Studying the role of the p53 tumor suppressor in embryonic development
The p53 family members have distinct roles during mammalian embryonic development.
Cell death and differentiation
The p53 tumor suppressor is a member of a multi-protein family, including the p63 and p73 transcription factors. These proteins can bind to the same consensus sites in DNA and activate the same target genes, suggesting that there could be functional redundancy between them. Indeed, double mutant mice heterozygous for any two family member-encoding genes display enhanced cancer phenotypes relative to single heterozygous mutants. However, whether the family members play redundant roles during embryonic development has remained largely unexplored. Although p53(-/-); p73(-/-) mice are born and manifest phenotypes characteristic of each of the single mutants, the consequences of combined deficiency of p63 and either p53 or p73 have not been elucidated. To examine the functional overlap of p53 family members during development, we bred and analyzed compound mutant embryo phenotypes. We discovered that double knockout embryos and five allele knockout embryos only displayed obvious defects accounted for by loss of single p53 family members. Surprisingly, at mid-gestation (E11), we identified a single viable triple knockout embryo that appeared grossly normal. Together, these results suggest that the p53 family is not absolutely required for early embryogenesis and that p53 family members are largely non-redundant during early development.
View details for DOI 10.1038/cdd.2016.128
View details for PubMedID 28211873
View details for PubMedCentralID PMC5384018
SHP2 Regulates Chondrocyte Terminal Differentiation, Growth Plate Architecture and Skeletal Cell Fates
2014; 10 (5)
Loss of PTPN11/SHP2 in mice or in human metachondromatosis (MC) patients causes benign cartilage tumors on the bone surface (exostoses) and within bones (enchondromas). To elucidate the mechanisms underlying cartilage tumor formation, we investigated the role of SHP2 in the specification, maturation and organization of chondrocytes. Firstly, we studied chondrocyte maturation by performing RNA-seq on primary chondrocyte pellet cultures. We found that SHP2 depletion, or inhibition of the ERK1/2 pathway, delays the terminal differentiation of chondrocytes from the early-hypertrophic to the late-hypertrophic stage. Secondly, we studied chondrocyte maturation and organization in mice with a mosaic postnatal inactivation of Ptpn11 in chondrocytes. We found that the vertebral growth plates of these mice have expanded domains of early-hypertrophic chondrocytes that have not yet terminally differentiated, and their enchondroma-like lesions arise from chondrocytes displaced from the growth plate due to a disruption in the organization of maturation and ossification zones. Furthermore, we observed that lesions from human MC patients also display disorganized chondrocyte maturation zones. Next, we found that inactivation of Ptpn11 in Fsp1-Cre-expressing fibroblasts induces exostosis-like outgrowths, suggesting that loss of SHP2 in cells on the bone surface and at bone-ligament attachment sites induces ectopic chondrogenesis. Finally, we performed lineage tracing to show that exostoses and enchondromas in mice likely contain mixtures of wild-type and SHP2-deficient chondrocytes. Together, these data indicate that in patients with MC, who are heterozygous for inherited PTPN11 loss-of-function mutations, second-hit mutations in PTPN11 can induce enchondromas by disrupting the organization and delaying the terminal differentiation of growth plate chondrocytes, and can induce exostoses by causing ectopic chondrogenesis of cells on the bone surface. Furthermore, the data are consistent with paracrine signaling from SHP2-deficient cells causing SHP2-sufficient cells to be incorporated into the lesions.
View details for DOI 10.1371/journal.pgen.1004364
View details for Web of Science ID 000337145100055
View details for PubMedID 24875294
Sclerostin Inhibition Reverses Skeletal Fragility in an Lrp5-Deficient Mouse Model of OPPG Syndrome
SCIENCE TRANSLATIONAL MEDICINE
2013; 5 (211)
Osteoporosis pseudoglioma syndrome (OPPG) is a rare genetic disease that produces debilitating effects in the skeleton. OPPG is caused by mutations in LRP5, a WNT co-receptor that mediates osteoblast activity. WNT signaling through LRP5, and also through the closely related receptor LRP6, is inhibited by the protein sclerostin (SOST). It is unclear whether OPPG patients might benefit from the anabolic action of sclerostin neutralization therapy (an approach currently being pursued in clinical trials for postmenopausal osteoporosis) in light of their LRP5 deficiency and consequent osteoblast impairment. To assess whether loss of sclerostin is anabolic in OPPG, we measured bone properties in a mouse model of OPPG (Lrp5(-/-)), a mouse model of sclerosteosis (Sost(-/-)), and in mice with both genes knocked out (Lrp5(-/-);Sost(-/-)). Lrp5(-/-);Sost(-/-) mice have larger, denser, and stronger bones than do Lrp5(-/-) mice, indicating that SOST deficiency can improve bone properties via pathways that do not require LRP5. Next, we determined whether the anabolic effects of sclerostin depletion in Lrp5(-/-) mice are retained in adult mice by treating 17-week-old Lrp5(-/-) mice with a sclerostin antibody for 3 weeks. Lrp5(+/+) and Lrp5(-/-) mice each exhibited osteoanabolic responses to antibody therapy, as indicated by increased bone mineral density, content, and formation rates. Collectively, our data show that inhibiting sclerostin can improve bone mass whether LRP5 is present or not. In the absence of LRP5, the anabolic effects of SOST depletion can occur via other receptors (such as LRP4/6). Regardless of the mechanism, our results suggest that humans with OPPG might benefit from sclerostin neutralization therapies.
View details for DOI 10.1126/scitranslmed.3006627
View details for Web of Science ID 000327012900005
View details for PubMedID 24225945
Perspectives for identification of mutations in the zebrafish: Making use of next-generation sequencing technologies for forward genetic approaches
2013; 62 (3): 185-196
The ability to identify a phenotype causing mutation is essential for successful use of mutagenesis screens in many model organisms. Mapping mutations was for a long time a bottleneck in zebrafish research, as the standard method for mapping and identification of mutations was time consuming and expensive. The development of new sequencing technologies in the last couple of years has enabled the rapid and cost-effective sequencing of whole genomes. This has led to the establishment of new strategies for mapping and identification of mutations in several model organisms. The application of these techniques to the zebrafish model, with its large genome and the high level of variation in and between strains, was not trivial. Several techniques have been developed recently, taking the specific characteristics of the zebrafish genome into account. Here we give an overview on how to plan a mapping experiment, detail the critical parameters and discuss available tools for mapping and identification of mutations in zebrafish using next-generation sequencing. Using these methods, zebrafish mutants can now be mapped in a couple of weeks for a fraction of the costs. The increased efficiency of identification of mutations in the zebrafish broadens the utility of the model and allows for systematic analysis of gene function in a vertebrate model.
View details for DOI 10.1016/j.ymeth.2013.05.015
View details for Web of Science ID 000324454000002
View details for PubMedID 23748111
Presphenoidal synchondrosis fusion in DBA/2J mice
2013; 24 (1-2): 54-62
Cranial base growth plates are important centers of longitudinal growth in the skull and are responsible for the proper anterior placement of the face and the stimulation of normal cranial vault development. We report that the presphenoidal synchondrosis (PSS), a midline growth plate of the cranial base, closes in the DBA/2J mouse strain but not in other common inbred strains. We investigated the genetics of PSS closure in DBA/2J mice by evaluating F1, F1 backcross, and/or F1 intercross offspring from matings with C57BL/6J and DBA/1J mice, whose PSS remain open. We observed that PSS closure is genetically determined, but not inherited as a simple Mendelian trait. Employing a genome-wide SNP array, we identified a region on chromosome 11 in the C57BL/6J strain that affected the frequency of PSS closure in F1 backcross and F1 intercross offspring. The equivalent region in the DBA/1J strain did not affect PSS closure in F1 intercross offspring. We conclude that PSS closure in the DBA/2J strain is complex and modified by different loci when outcrossed with C57BL/6J and DBA/1J mice.
View details for DOI 10.1007/s00335-012-9437-8
View details for Web of Science ID 000314298600005
View details for PubMedID 23179633
Identification of mutations in zebrafish using next-generation sequencing.
Current protocols in molecular biology / edited by Frederick M. Ausubel ... [et al.]
2013; 104: 7 13 1-7 13 33
Whole-genome sequencing (WGS) has been used in many invertebrate model organisms as an efficient tool for mapping and identification of mutations affecting particular morphological or physiological processes. However, the application of WGS in highly polymorphic, larger genomes of vertebrates has required new experimental and analytical approaches. As a consequence, a wealth of different analytical tools has been developed. As the generation and analysis of data stemming from WGS can be unwieldy and daunting to researchers not accustomed to many common bioinformatic analyses and Unix-based computational tools, we focus on how to manage and analyze next-generation sequencing datasets without an extensive computational infrastructure and in-depth bioinformatic knowledge. Here we describe methods for the analysis of WGS for use in mapping and identification of mutations in the zebrafish. We stress key elements of the experimental design and the analytical approach that allow the use of this method across different sequencing platforms and in different model organisms with annotated genomes. Curr. Protoc. Mol. Biol. 104:7.13.1-7.13.33. © 2013 by John Wiley & Sons, Inc.
View details for DOI 10.1002/0471142727.mb0713s104
View details for PubMedID 24510885
Somatic Mosaic Activating Mutations in PIK3CA Cause CLOVES Syndrome
AMERICAN JOURNAL OF HUMAN GENETICS
2012; 90 (6): 1108-1115
Congenital lipomatous overgrowth with vascular, epidermal, and skeletal anomalies (CLOVES) is a sporadically occurring, nonhereditary disorder characterized by asymmetric somatic hypertrophy and anomalies in multiple organs. We hypothesized that CLOVES syndrome would be caused by a somatic mutation arising during early embryonic development. Therefore, we employed massively parallel sequencing to search for somatic mosaic mutations in fresh, frozen, or fixed archival tissue from six affected individuals. We identified mutations in PIK3CA in all six individuals, and mutant allele frequencies ranged from 3% to 30% in affected tissue from multiple embryonic lineages. Interestingly, these same mutations have been identified in cancer cells, in which they increase phosphoinositide-3-kinase activity. We conclude that CLOVES is caused by postzygotic activating mutations in PIK3CA. The application of similar sequencing strategies will probably identify additional genetic causes for sporadically occurring, nonheritable malformations.
View details for DOI 10.1016/j.ajhg.2012.05.006
View details for Web of Science ID 000305262600016
View details for PubMedID 22658544
Efficient Mapping and Cloning of Mutations in Zebrafish by Low-Coverage Whole-Genome Sequencing
2012; 190 (3): 1017-U229
The generation and analysis of mutants in zebrafish has been instrumental in defining the genetic regulation of vertebrate development, physiology, and disease. However, identifying the genetic changes that underlie mutant phenotypes remains a significant bottleneck in the analysis of mutants. Whole-genome sequencing has recently emerged as a fast and efficient approach for identifying mutations in nonvertebrate model organisms. However, this approach has not been applied to zebrafish due to the complicating factors of having a large genome and lack of fully inbred lines. Here we provide a method for efficiently mapping and detecting mutations in zebrafish using these new parallel sequencing technologies. This method utilizes an extensive reference SNP database to define regions of homozygosity-by-descent by low coverage, whole-genome sequencing of pooled DNA from only a limited number of mutant F(2) fish. With this approach we mapped each of the five different zebrafish mutants we sequenced and identified likely causative nonsense mutations in two and candidate mutations in the remainder. Furthermore, we provide evidence that one of the identified mutations, a nonsense mutation in bmp1a, underlies the welded mutant phenotype.
View details for DOI 10.1534/genetics.111.136069
View details for Web of Science ID 000301531900013
View details for PubMedID 22174069
Loss-of-Function Mutations in PTPN11 Cause Metachondromatosis, but Not Ollier Disease or Maffucci Syndrome
2011; 7 (4)
Metachondromatosis (MC) is a rare, autosomal dominant, incompletely penetrant combined exostosis and enchondromatosis tumor syndrome. MC is clinically distinct from other multiple exostosis or multiple enchondromatosis syndromes and is unlinked to EXT1 and EXT2, the genes responsible for autosomal dominant multiple osteochondromas (MO). To identify a gene for MC, we performed linkage analysis with high-density SNP arrays in a single family, used a targeted array to capture exons and promoter sequences from the linked interval in 16 participants from 11 MC families, and sequenced the captured DNA using high-throughput parallel sequencing technologies. DNA capture and parallel sequencing identified heterozygous putative loss-of-function mutations in PTPN11 in 4 of the 11 families. Sanger sequence analysis of PTPN11 coding regions in a total of 17 MC families identified mutations in 10 of them (5 frameshift, 2 nonsense, and 3 splice-site mutations). Copy number analysis of sequencing reads from a second targeted capture that included the entire PTPN11 gene identified an additional family with a 15 kb deletion spanning exon 7 of PTPN11. Microdissected MC lesions from two patients with PTPN11 mutations demonstrated loss-of-heterozygosity for the wild-type allele. We next sequenced PTPN11 in DNA samples from 54 patients with the multiple enchondromatosis disorders Ollier disease or Maffucci syndrome, but found no coding sequence PTPN11 mutations. We conclude that heterozygous loss-of-function mutations in PTPN11 are a frequent cause of MC, that lesions in patients with MC appear to arise following a "second hit," that MC may be locus heterogeneous since 1 familial and 5 sporadically occurring cases lacked obvious disease-causing PTPN11 mutations, and that PTPN11 mutations are not a common cause of Ollier disease or Maffucci syndrome.
View details for DOI 10.1371/journal.pgen.1002050
View details for Web of Science ID 000289977000031
View details for PubMedID 21533187