Jianjian Zhu, Ph.D.
Dr. Zhu utilizes genetic, genomic and embryological micro-manipulation approaches to study several important signaling pathways and transcription factors that interact and cooperate to regulate pattern formation and cell fate determination during embryonic development. Major focuses of research have been the role of Sonic Hedgehog and its signaling targets in early patterning of digit formation, and the function of Brachyury/T in notochord specification, maintenance, and differentiation. The working models include mouse mutant and reporter lines, chick embryos for micro-injection and beads implantation, bat limbs, as well as in vitro limb culture.
1) Hedgehog signaling, 2) short-hairpin RNA, 3) micro-manipulation, 4) tamoxifen-inducible Cre recombination
Dual roles of Shh in limb development: transient early requirement for patterning and sustained late requirement in cell expansion
Sonic Hedgehog (Shh) is a critical signaling molecule that is important for early embryonic development of multiple organs (nervous system, limb, etc) and when mis-regulated, is also involved in various cancers in adults (e.g. skin, nervous system, digestive system). In the developing limb, Shh determines digit number and digit identities (d1, thumb to d5, pinky, from anterior to posterior). The classical models include the morphogen gradient model and the integrated growth-morphogen model (either higher concentration or longer temporal integration of Shh signaling over time patterns more posterior digits). I previously analyzed the temporal requirement for Shh in digit patterning and growth using a conditional knockout allele of Shh (Shh CKO) and a tamoxifen-regulated Cre (Hoxb6CreER; Nguyen, Zhu et al, Dev. Dyn, 2009) and showed these roles can be uncoupled. I proposed a new bi-phasic model in which Shh is required only very early and transiently for patterning, but sustained activity over time is needed to promote cell survival and proliferation (Zhu et al, Dev. Cell, 2008).
This model is validated in several mutant lines with digit defects (Zhu and Mackem, Dev. Dyn, 2011), and is directly tested by restoring cell survival in Shh CKO embryos after transient Shh expression by concomitant removal of the pro-apoptotic genes Bax/Bak to intercept cell death, in order to rescue the proposed late Shh role. Half of the Bax/Bak DKO rescued embryos have up to 5 normal digits and long bones, compared to the single dysmorphic digit in non-rescued Shh CKO embryos. Nevertheless, a transient Shh pulse is essential for patterning; complete rescue of cell survival in germline Shh-/- embryos has no effect on the null phenotype. Our results reveal critical Shh roles in regulating growth and morphogenesis that are distinct and temporally separable, and point to 2 different classes of Hedgehog targets that require either transient or continuous signaling for their sustained activation.
Current work is focused on delineating these target classes and learning how Shh relays the downstream target signals to instruct limb morphogenesis. Elucidating Shh function in the context of these differentially regulated target classes will be relevant to Hedgehog signaling in other contexts, such as self-renewal of progenitors, and pathogenesis of tumors and has implications for deciphering and intercepting Hedgehog activity in cancer pathogenesis.
Role of Brachyury (T) in specification and maintenance of notochord progenitors
The transcription factor Brachyury (T gene) is expressed throughout primary mesoderm (primitive streak and notochord) during early embryonic development and has been strongly implicated in the genesis of chordoma, a sarcoma of notochord cell origin. Additionally, T expression has been found in, and proposed to play a role in promoting epithelial-mesenchymal transition (EMT) in various other types of human tumors.
I have generated a very efficient Cre-inducible T knockdown model (T-short hairpin RNA transgene) to analyze tissue-specific T function in mouse embryos. Notochord-specific Cre lines (ShhCre and Foxa2CreER) have been used, and results show that T function is essential for maintaining notochord cell fate and function. Progenitors adopt predominantly a neural fate in the absence of T. However, T function is dispensable for progenitor cell survival, proliferation and EMT, which has implications for the therapeutic targeting of T in chordoma and other cancers (Zhu et al, PNAS 2016). Collaboration is currently ongoing to use this inducible T-knockdown line for analysis of neural-mesodermal stem cells.
In collaboration, I have investigated EpCAM mutants and identified the placental defects causing early embryonic lethality (Nagao, Zhu et al, Plos One, 2009), and have analyzed TAF7 mutants (Gegonne et al, MCB, 2012). I have recently collaborated with scientists in Chemical Biology Lab to characterize a near-IR uncaging tamoxifen approach in tissue culture and embryonic development (Gorka et al, JACS, 2014).
Selected Key Publications
The putative oncogene Brachyury (T) is essential to specify cell fate, but dispensable for notochord progenitor proliferation and EMT.Proc Natl Acad Sci U S A. Mar 22: pii: 201601252. [Epub ahead of print], 2016. [ Journal Article ]
Cre-mediated recombination can induce apoptosis in vivo by activating the p53 DNA damage-induced pathway.Genesis. 50(2): 102-111, 2012. [ Journal Article ]
Analysis of mutants with altered Shh activity and posterior digit loss supports a biphasic model for Shh function as a morphogen and mitogen.Dev Dyn. 240: 1303-1310, 2011. [ Journal Article ]
Tamoxifen-dependent, inducible Hoxb6CreERT recombinase function in lateral plate and limb mesoderm, CNS isthmic organizer, posterior trunk neural crest, hindgut, and tailbud.Dev Dyn. 238: 467-474, 2009. [ Journal Article ]
- Dev Cell. 14: 624-632, 2008. [ Journal Article ]
Dr. Zhu obtained his Ph.D. in molecular biology from Peking University Health Science Center, China in 2001. After a brief postdoctoral fellowship in the Institute of Cardiovascular Research, Peking University, Dr. Zhu came to Dr. Susan Mackem’s lab in Laboratory of Pathology, National Cancer Institute in 2002, and joined the Cancer and Developmental Biology Laboratory, National Cancer Institute at Frederick in 2009. Since 2011, Dr. Zhu has served as a staff scientist at the National Cancer Institute at Frederick.