The role of hdac4 in zebrafish craniofacial development Vishesh Khanna University of Oregon PI: Prof. Charles Kimmel Mentor: Dr. April DeLaurier Hypothesis: hdac4 patterns anterior craniofacial development through upregulation of shh

hdac = Histone Deacetylase Transcriptional repressors SNPs in the human hdac4 gene are linked to cleft palate-like defects (Park et al. 2006) and skeletal defects in mice (Vega et al. 2004) Zebrafish may serve as a good model by which to study craniofacial disorders in humans Images from Ida.org.in and Kimmel et. al 2001

Hypothesis: hdac4 patterns neurocranium formation through upregulation of shh Images from Kimmel et al. 2001

Morpholino (MO) knockdowns of hdac4 mRNA result in neurocranial defects WT 6dpf hdac4 MO 6dpf Photos courtesy of Dr. April DeLaurier hdac4 MO 6dpf hdac4 MO 6dpf hdac4 MO 6dpf

sonic hedgehog (shh) and hdac4 shh plays a crucial role in craniofacial development Zebrafish shh - mutants show variable neurocranium defects (Eberhart et al. 2006) Loss of Hdac proteins results in reduction of shh expression in chick limb buds (Zhao et al. 2009) 4 dpf -- Photo from Wada et al, 2005 WT

shh and hdac4 Two shh signals are required for neurocranium development: 1)Shh signals to the stomodeal (mouth) precursors 2)Stomodeum signals to post-migratory neural crest cells Images adapted from Eberhart et al. 2006

At 10 and 14 hpf, loss of hdac4 ≠ loss of shh shh WT 10 hpfMO 10 hpf shh WT 14 hpfMO 14 hpf

At 36 hpf, loss of hdac4 = loss of shh 36 hpf shh WT MO

Conclusion Hypothesis: hdac4 patterns anterior craniofacial development through upregulation of shh. hdac4 knockdown does not result in downregulation of shh at early stages (10 & 14 hpf) but may affect shh expression at later (36 hpf) stages of development My hypothesis can be rejected at early stages but may hold true for later stages

Future Directions Repeat in-situ hybridizations of shh expression at later stages Look for a genetic interaction between hdac4 and pdgfra (platelet-derived growth factor receptor alpha) and factors in the pdgf pathway – Factors in the pdgf pathway is important for neural crest cell migration and its loss is associated with anterior neurocranium defect s

Acknowledgments Professor Chuck Kimmel Dr. April DeLaurier The Kimmel Lab Dr. Peter O’Day Blakely Strand The SPUR participants