Inhibition of the serine protease HtrA1 by SerpinE2 suggests a role for an extracellular proteolytic pathway in the control of neural crest migration

Reviewer #1 (Public Review):

Summary:

A novel serine protease and an inhibitor pair regulate cell migration in the neural crest. This is a very important study that describes a novel pathway controlling neural crest migratory behavior through a pair of protease and inhibitor regulators that act in the extracellular space. Using very high technical standards in Xenopus embryos they show that knockdown of the inhibitor SerpinE2 prevents cell migration and that this is restored by simultaneous knockdown of the serine protease HtrA1.

Strengths:

The reproduction of classical cranial neural crest extirpations and their phenocopy by SerpinE2 morpholino is remarkable. The experiments provided must represent many years of work, and the paper is written in a very scholarly fashion. The data is of the highest quality.

Weaknesses:

The paper is very long and contains many years of experiments, making it at times difficult to read. The paper contains so much data that it would help the readership if the present version were revised in order to make it more digestible.

Reviewer #2 (Public Review):

Summary:

The authors conducted research on the role of SerpinE2 and HtrA1 in neural crest migration using Xenopus embryos. The data presented in this study was of high quality and supported the authors' conclusions. The discovery of the potential molecular connection between SerpinE2 and HtrA1 in neural crest cell migration in vivo is significant, as understanding this pathway could potentially lead to treatments for aggressive cancers and pregnancy-related disorders.

Strengths:

Previous research has shown that SerpinE2 and HtrA1 can have both positive and negative effects on cell migration, but their molecular interplay and role in neural crest migration are not well-established. This study is the first to reveal a potential connection between these two proteins in neural crest cell migration in vivo. The authors found that SerpinE2 promotes neural crest migration by inhibiting HtrA1. Additionally, overexpression of Sdc4 partly alleviates neural crest migration issues caused by SerpinE2 knockdown or HtrA1 overexpression. These findings suggest that the SeprinE2-HtrA1-Sdc4 pathway is crucial for neural crest migration.

Weaknesses:

To further increase the study's credibility, the authors could use techniques like western blotting, qRT-PCR, or in situ hybridization to verify the efficiency of SerpinE2 and HtrA1 knockdown and/or overexpression. Furthermore, determining whether the observed craniofacial phenotypes in SerpinE2 and/or HtrA1 mutants were solely due to modified cranial neural crest migration or affected by other factors such as cell proliferation, cell survival, and chondrogenic differentiation could provide more clarity. Lastly, it is unclear whether the SeprinE2-HtrA1-Sdc4 pathway is constant in both cranial and trunk neural crest migration.