Peer review of: Feeding-dependent tentacle development in the sea anemone Nematostella vectensis (2020)


 
 

Reviewers' Comments: 
 
Reviewer #1: 
Remarks to the Author: 
In the revised manuscript, Ikmi et al. made considerable efforts to address my 
concerns/comments. They chose, however, to provide only partial answers to some points. For the 
origin of the FGFRb expressing cell clusters it is now nicely shown that these expression domains 
are not an artefact. In remains unclear how the number of these clusters relates to the developing 
tentacles. In Fig S10D there are three such clusters visible and labelled, but a later stage (S10F) 
there are only two that label the new tentacle buds. The authors point out the latter observation 
but I still find it unclear whether the so-called “pre-metamorphic” clusters are really the origin of 
the new tentacle buds. The authors also did SU5402 treatments to show that the development of 
the tentacle buds requires FGF signalling after the first four tentacles are formed. This approach 
may also have been useful for earlier treatments to test whether the formation of the FGFRb 
clusters (and new tentacle buds) is affected before or during metamorphosis. There is now a better 
description of the phenotype of the FGFRb mutants at polyp stage, but how the phenotype 
develops over time, is not addressed. 
These are non-essential points, but if the authors have the relevant data available, they could 
further strengthen the manuscript. 
 
The authors have also provided a clearer description of the conceptual background and its 
relevance. 
 
Other points: 
line 113 micromeres should read microcnemes 
line 121 – Please replace “adult organogenesis” with “tentacle development”. While definitions of 
what can be called an organ are vague, the use of this term here is in my eyes an attempt to 
attract attention to animals that are typically said to lack organ-level organization. 
 
 
 
Reviewer #3: 
Remarks to the Author: 
The authors failed to reference several of the seminal works on this topic (including Stephenson 
1935 and Crowell 1946) but many of the observations they present have been published 
previously. Nematostella can have variable numbers of tentacles, which has been described, but 
the fact that wild populations typically have 16 (or fewer) tentacles suggests that the 20-, 22- and 
24-tentacle animals the authors describe are aberrations that may not actually reflect the biology 
of this animal. 
The authors continue to provide data that are of insufficient magnification and quality (and without 
proper counterstains) to evaluate their claims. 
 
Figures 1-3 are fine, they have not changed from the first version. 
 
Figure 4 The authors added two arrowheads pointing to “unspecific” EdU labeling in the first image 
in panel F but at the magnification they have provided, there is no way for any reader to 
discriminate “specific” from “unspecific” staining in these images. Thus, we are required to trust 
the authors’ interpretation. The authors provided new panels showing quantification of the green 
signal in their images but have already pointed out that this label is not specific and seems to vary 
across treatments. 
 
Figure 5 Panel C is identical in magnification and quality to the previous version and is still of 
insufficient quality to evaluate whether any of these labels (pRPS6 or EdU) are 
intracellular/nuclear. Panel D (or E?) claims to report RPS6 “intensity” (according to the rebuttal 
letter) but really these figures demonstrate EdU “intensity” (and I have already discussed my 

concerns with this) and a count of the polyps the authors considered to be positive for pRPS6 
expression. 
 
Figure 6 Panel A is lovely, but panel B shows staining in some unidentified tissue at an oblique 
angle that is not interpretable, and panels C/D/E have no nuclear counterstain making it nearly 
impossible for anyone to know whether those cells are ectodermal or endodermal. Panel E seems 
to show ectodermal expression in the top panels (unfed) and endoderm in the bottom panels (fed) 
but it is not possible to be sure from the images provided. 
 
Figure 7 Panel E shows two different planes from control and experimental animals so it is not 
possible for the reader to draw any reasonable conclusions about whether the “oral cell clusters” 
are still there or not. Furthermore, there is a fundamental difference in the staining represented in 
the top and bottom parts of panel F that is not mentioned. Either the antibody is responding 
differently to these tissues in different treatments or the authors have provided images from 
significantly different focal planes that reflect different parts of the cell/tissue biology of these 
animals. 
 
Supplemental Figure 7 is impossible to interpret as there is no cellular level resolution. 
 
Supplemental Figure 8 Both panels are presented in insufficient quality to see cellular level 
resolution. Furthermore, the images in the bottom of panel B claim to show a cross section but 
seems to be showing body wall ectoderm. 
 
Supplemental Figure 12 Panel E is not presented in high enough magnification to evaluate their 
claims – are the cells of the (ectodermal) septal filaments gone or has the epithelium just become 
cuboidal instead of the normal columnar morphology?