Eroshkin FM et al. (2024), Mechanical Tensions Regulate Gene Expression in...

XB-ART-60541

Int J Mol Sci 2024 Jan 10;252:. doi: 10.3390/ijms25020870.

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Mechanical Tensions Regulate Gene Expression in the Xenopus laevis Axial Tissues.

Eroshkin FM , Fefelova EA , Bredov DV , Orlov EE , Kolyupanova NM , Mazur AM , Sokolov AS , Zhigalova NA , Prokhortchouk EB , Nesterenko AM , Zaraisky AG .

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During gastrulation and neurulation, the chordamesoderm and overlying neuroectoderm of vertebrate embryos converge under the control of a specific genetic programme to the dorsal midline, simultaneously extending along it. However, whether mechanical tensions resulting from these morphogenetic movements play a role in long-range feedback signaling that in turn regulates gene expression in the chordamesoderm and neuroectoderm is unclear. In the present work, by using a model of artificially stretched explants of Xenopus midgastrula embryos and full-transcriptome sequencing, we identified genes with altered expression in response to external mechanical stretching. Importantly, mechanically activated genes appeared to be expressed during normal development in the trunk, i.e., in the stretched region only. By contrast, genes inhibited by mechanical stretching were normally expressed in the anterior neuroectoderm, where mechanical stress is low. These results indicate that mechanical tensions may play the role of a long-range signaling factor that regulates patterning of the embryo, serving as a link coupling morphogenesis and cell differentiation.

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Species referenced: Xenopus laevis
Genes referenced: adamts7 cd82 cdx4 cebpa cer1 foxc1 foxc2 frzb2 fst gbx2 gsc irx3 lef1 mmp21 myf5 nalcn otx2 prss8l.4 sfrp1 shh six3 socs3 tbx6 tbxt xmc
GO keywords: neural tube closure [+]

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Phenotypes: Xla Wt + neurectoderm explant + stretch (Fig. 3. B)

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	Figure 1. Genetically encoded mechanical sensor VinTS reveals different mechanical tensions in the head and trunk parts of the neuroectoderm. (A–D) VinTS mechanosensor. (E–H) VinTSΔC control. (A,E) Schema of sensor. (B,F) Images of the Xenopus embryo, microinjected with the VinTS mRNA, in the green channel. (C,G) The result of computer processing of images in green and cyan channels. Red color corresponds to low degree of MTs (high FRET); yellow and blue color corresponds to high degree of MTs (low FRET). (D,H) Vertical plot showing the distribution of signal intensity (green) and FRET (orange) along the anterior–posterior axis of the embryo.
	Figure 2. Morphometric analysis of tissue deformation. (A,B) To evaluate tissue deformation, we performed time-lapse imaging of cells in head (A) and trunk (B) regions of st. 14–15 Xenopus neurula during a 20 min interval. Insets in the right row show the position of segmented regions (enclosed with white dashed line) on scheme (A′,B′) and in imaged embryos (A″,B″) under lower magnification. Cell trajectories in (A,B) are color-coded: yellow–green indicates the initial part of the trajectory, blue—central, purple–red—terminal. (C,D) Tissue-level deformation evaluated by shape change of integral ROI including all segmented cells. Green and red counters reflect ROI shape at the first and last frame, respectively, of the time-lapse sequence. Individual trajectories of selected cells are visualized as black lines; arrows indicate direction of cell displacements inside ROIs. (C′,D′) and (C″,D″): To quantitatively describe spatial distribution of cell displacements underlying ROI deformation presented in (C,D), we calculated cosine of the angle between anteroposterior axis and direction of each cell displacement during two initial (C′,D′) and two terminal (C″,D″) frames in sequence and plotted this cosine value against ordinate (in pixels). We denote 0° (360°) displacement angle as shift towards anterior pole, and 180°—towards posterior. Thus, cosine values close to 1 indicate displacement towards the anterior pole, and values close to −1—towards the posterior pole. Note the marked translational shift of ROI in the head region without significant elongation on the contrary to the trunk region, that mostly remained in the field of view and was subjected to bidirectional ((D″) black dotted boxes) elongation. (E,F) To estimate cellular shape changes underlying observed ROI deformations, we analyzed distribution of cell eccentricities (E) for head (Nembryos = 1, ncells = 136) and trunk (Nembryos = 1, ncells = 56) regions at the start (blue) and the end (grey) of time-lapse series as well as area changes (F) of cells (blue) and small ROIs, including 10–11 cells (gray); columns show mean, whiskers—95% confidence interval. Inset in (E) (reprinted under Creative Commons 3.0 license) shows typical cell and respective ellipse shape for eccentricity = 0.6 and for eccentricity = 0.8. Note the slight (but statistically significant) increase in mean eccentricity in the trunk region while the apical cell area remains the same, reflecting cell elongation in the trunk region. At the same time, mean eccentricity in the head region did not change, while the apical cell area decreased, reflecting apical constriction.
	Figure 3. Convergent extension of the neurectoderm and schema of experiments in finding of genes regulated by mechanical tensions. (A) Photos of FLD-injected neural plate at different successive stages of development. White dashed lines connect at different stages the same noticeable features at the embryo surface. The red dashed line on the image of the midgastrula embryo indicates the approximate border of the neuroectodermal explants which had been excised at this stage. (B) Schema of experiments of explants stretching followed by NGS (see main text for details). Red dashed lines indicate the boundaries of the explants.
	Figure 4. Analysis of the influence of the neuroectodermal explants stretching on gene expression. (A) Examples of explants nonstretched and stretched according to the schema shown in Figure 3B. (B) Genes whose expression changed in the explants upon the stretching, according to qRT-PCR data. Bars indicate the standard deviation. (C) Percentage expression distribution of selected MT-responsive genes in the indicated on the right groups of cells in midgastrula embryo (stage 14) according to the Jamboree single-cell sequencing database [24]. The group anterior neural includes such cell types as anterior neural plate; the group anterior non-neural includes such cell types as anterior placodal area, cement gland primordium, eye primordium, and anterior neural crest; the group posterior neural includes such cell types as posterior neural plate; the group posterior non-neural includes such cell types as presomitic mesoderm, somite, and tail bud; the group other includes other cell types (see Xenopus single-cell transcriptomic data analysis in Section 4.6 for details).

References [+] :

Agricola, Identification of genes expressed in the migrating primitive myeloid lineage of Xenopus laevis. 2016, Pubmed, Xenbase