Dr. Lee A. Niswander

Grainyhead-like 2 and Neural Fold Fusion

Ray HJ, Niswander L. 2012. Mechanisms of tissue fusion during development. Development. 139(10):1701-11

Pyrgaki C, Trainor P, Hadjantonakis AK, Niswander L. Dynamic imaging of mammalian neural tube closure. Dev Biol. 2010 Aug 15;344(2):941-7.

Pyrgaki C, Liu A, Niswander L. 2011. Grainyhead-like 2 regulates neural tube closure and adhesion molecule expression during neural fold fusion. Dev Biol. 353(1):38-49.

Introduction

Very briefly tell us a bit about yourself, your career path over the years, and specifically what brought you to begin working on the mechanisms of neural tube closure?

Michael Barresi

Question 1

In class we learned that both E- and N-cadherins are involved in neural fold fusion, the paper mostly focuses on E-cadherins as they were shown to be the top targets of Grhl2. Do you think N-cadherins could be involved in the AP-2alpha pathway instead as the in AP-2alpha mutation shows normal expression of E-cadherins.

Belinda Nhundu

Question 2

In class we learned how differential expression of proteins in apical and basal regions can result in shape changes that produce bottle cells.  Do you have any hypotheses as to how the cells from opposite sides of the neural folds undergo what you call a “dramatic change in shape?”

Lisa Friedman

Question 3

Do you believe that the appropriate localization of β-catenin in the absence of E-cadherin is due to some compensatory mechanism, or does it point to a lack of E-cadherin involvement in the localization process?

Anne Slingerland

Question 4

If you disrupt the localization of B-catenin on the plasma membrane, what do you think the result will look like?

Gloria Kim

Question 5

If there is a close relationship between E-cadherin and another protein, is it possible that this other protein is downregulated in AP-2a mutants but present in Grhl2? In this way, each mutant would be lacking one main component necessary for adhesive properties of the non-neural ectoderm while normally expressing the other protein?

Elizabeth Summer

Question 6

In Table 1, p-value shows the significant difference of each gene expression, telling that all 10 genes are significantly downregulated. E-cdh shows the lowest p-value (0.0003) with certain fold reduction (-4.00). Rab25 had a significanly p-value, but not even close to E-cdh. However, Rab25 showed the biggest reduction in folding (-23.28) among 10 genes. Does this mean that Rab25 has most influence in folding among 10 gene?

Sam Kim

Question 7

In the dynamic imaging paper, a novel finding was of cellular bridges, and it was hypothesized that they may function as communication highways, transporting different elements between the two sides of the non-neural ectoderm.  If this is the case, what could be getting transported that would cause simultanous zippering or buttoning up of the two sides of the presumptive neural tube? Alternatively, could these bridges be exerting a physical force, such as pulling the two sides together?

Julia Johnstone

Question 8

Based on the Dynamic imaging of mammalian neural tube closure paper, what would you hypothesize would be the evolutionary cause for differential neural tube closure mechanisms in the hindbrain and midbrain? Why would they both not depend on the zipper-like mechanism or the buttoning up-like mechanism? And if the cellular bridges in the midbrain really are involved in potentially stabilizing intermediate closure points, then wouldn’t that be a more reliable way to avoid NTD then the zippering-like mechanism?

Sheila Sernoff

Question 9

If Grhl2 is critical to controlling adhesion to more anterior structures, is there a similarly-functioning ‘posterior’ version of Grhl2? Would Grhl3 be a candidate for this, as its loss-of-function causes spina bifida in 100% of mutants?.

Kristine Trotta

Question 10

In your paper, you have shown what is regulated by Grhl2, but what regulates Grhl2?

Risha Sinha

Question 11

The phenotypes expressed in the ENU-induced mutant (fig 3E) look like they lack dorsal lateral hinge points (DLHP); we learned from class that Sonic hedgehog (Shh) represses DLHP formation. Do you have a hypothesis for how Grhl2 may interact with the Shh pathway to cause this phenotype?

Bokang Rabasha

Question 12

In a reference to Werth et al., 2010, Grhl2 is found to directly bind to the promoters of E-cadherin and Claudin4—a target identified in the microarray—to regulate transcription. You mention that the expression of claudins during development is still being discovered, but they are expressed in E9.5 embryos and their expression is affected by Grhl2. Does this imply that claudins, at least Claudin4, play(s) a role in neural fold fusion? Or can this conclusion not be made from the results of your microarray of genes affected by Grhl2 loss?

Stephanie Bond

Question 13

Currently in your opinion what are the most pressing questions about neural tube closure, and what steps is your lab taking to address these questions?

Michael Barresi