Disheveled-1 Interacts with Claudin-5 and Contributes to Norrin-Induced Endothelial Barrier Restoration
, Amy Wang 2, Xuwen Liu 2 and David A. Antonetti 2,*Round 1
Reviewer 1 Report
In this manuscript, the author effectively demonstrates that Disheveled-1 (DVL1) interacts with claudin-5 (CLDN5) and contributes to norrin-2-induced endothelial barrier restoration. Additionally, they discovered that DVL1 regulates tight junction organization through its PDZ domain binding to CLDN5. Their findings provide valuable insights into the study of tight junction organization. I have a few minor questions and suggestions:
Please avoid using red-green labeled figures, as they are not very user-friendly for individuals with visual impairments.
In Figure 1, the immunofluorescence conducted with the antibodies should be validated for specificity using knockdown cell lines.
In Figure 2, when the author attempted to knock down DVL1, the immunofluorescence data should also be presented.
The manuscript should discuss the impact of DVL1 overexpression on basal barrier function.
In panels 5 of Figure 1a and b, why does the wild-type (WT) DVL1 binding appear lower than DVL-deltaC7?
How does the knockout mouse model of DVL1 affect tight junction organization?
Author Response
"Please see the attachment"
Author Response File: 
Author Response.pdf
Reviewer 2 Report
Díaz-Coránguez and colleagues sought to investigate the role of disheveled-1 (DVL1) in Norrin-induced blood-retinal barrier restoration. The authors found a novel, non-canonical signaling pathway and demonstrated that DVL1 binds to claudin 5 to promote restoration of endothelial barrier properties after VEGF-induced permeability. The study also identified the mechanism of which DVL1 interacts with the PDZ-BM binding domain of CLDN5 for tight junction organization in the vascular endothelium. This is an interesting and well-performed study.
1. The authors demonstrated the mechanism of which DVL1 regulates tight junction organization in the endothelial cells. They showed that DVL proteins are highly expressed in the retinal capillaries, as well as in the isolated BRECs. In addition, knockdown of Dvl1 ablated norrin-induced increase in the resistance of BRECs. It would strengthen the study showing whether knockdown or supplementation of Dvl1 actually increases or decreases the leakage of retinal capillaries in vivo after VEGF/norrin treatment.
2. Figure 2, what is the reason that siDvl1a significantly increased the Axin2 mRNA expression (~3-fold increase, fig. 2c) while siDvl1b did not have an effect on Axin2 mRNA level (fig. 2f), although both siRNAs similarly knockdown the expression of Dvl1 mRNA?
Minor:
1. Page 8, line 303, please correct “Dv1”.
Minor editing of English language and spell check required
Author Response
"Please see the attachment"
Author Response File: Author Response.pdf
