Date: Friday, 10 May 2019

Venue: Seminar Room 1, G/F
Laboratory Block, Faculty of Medicine Building
21 Sassoon Road, Hong Kong

Time: 5:00 p.m.

Title: The Role of ephrin-A1/EphA4 signaling in Enteric Neural Crest Cell Migration
Speaker: Mr. LUI Jeffrey (MPhil candidate)

Summary:
Enteric neural crest cells (ENCCs) migrate and colonize the gut during enteric nervous system (ENS) development, in which Ret and p75 support ENCC survival, proliferation, migration and differentiation. Cell-cell interaction is largely regulated by Eph/ephrin signaling that induces attraction or repulsion between cells, crucial to cell segregation and thus cell migration in nervous system development. However, the role of Eph/ephrin signaling in regulating ENCC migration remains unknown. Mutation of Ephs and ephrins have been reported in Hirschsprung disease (HSCR) patients, suggesting the involvement of Eph/ephrin signaling. I examined the role of ephrin-A1/EphA4 signaling using 293T cells to investigate their interaction and regulation of cell segregation. Immunoprecipitation shows that ephrin-A1 can interact with Ret and p75. Ephrin-A1 mutant cloned to mimic human mutation reported is also able to interact with Ret and p75, and perform ligand-receptor binding interaction with EphA4. Cell segregation assay using stably-transfected cells shows that ephrin-A1 mutant-expressing cells demonstrate abnormal cell segregation when co-culturing with EphA4-expressing cells. These findings suggest that ephrin-A1 mutant could disrupt cell segregation regulated by ephrin-A1/EphA4 signaling, while interaction of ephrin-A1 with Ret and p75 suggests that it potentially plays an important role in this cell-cell interaction to regulate ENCC migration.

Title: A knock-in strategy to study protein localization in human induced pluripotent stem cell (hiPSC)-derived neuron through genome editing
Speaker: Mr. LYU Quanwei (MPhil candidate)

Summary:
The emergence of human induced pluripotent stem cells (hiPSCs) and their ability to differentiate into multiple subtypes of neuron present the attractive potential as a cellular model to study human brain disorders. Furthermore, the graft of hiPSC-derived neurons into the mouse brains potentially allows exploration of how their synaptic connections form in vivo. Many disorders on the central nervous system involve mis-localization of proteins in neurons. Here I use CRISPR-Cas9 to create a knock-in epitope tag to study protein localization in hiPSC-derived neurons. We have developed the differentiation protocol to generate cortical neurons that express synaptic markers and produce miniature excitatory postsynaptic current. As a proof-of-principle investigation, we test the possibility of labeling endogenous β-actin by adding a hemagglutinin (HA) tag to the N-terminal of b-actin through genome editing. After differentiation of the genome-edited hiPSCs into cortical neurons, the enrichment of b-actin in the growth cones and dendritic spines were visualized by immunostaining through anti-HA antibody. Our findings suggest the feasibility of this knock-in strategy to label synaptic proteins for the study of protein localization during synaptogenesis in hiPSC-derived cortical neurons. I will also discuss the use of this approach to create and screen for mutant iPSCs for the study of amyotrophic lateral sclerosis (ALS).

ALL ARE WELCOME