Multiscale analysis of single and double maternal-zygotic Myh9 and Myh10 mutants during mouse preimplantation development
- Institut Curie, PSL Research University, Sorbonne Université, France
- Carl Zeiss SAS, France
Figures
Figure 1 with 1 supplement
Expression of non-muscle myosin II heavy chain (NMHC) paralogs during preimplantation development.
(A) RT-qPCR of Myh9 (red), Myh10 (blue), and Myh14 (grey) at the zygote (E0.5, n = 234), four-cell (E1.5, n = 159), morula (E2.5, n = 189), and blastocyst (E3.5, n = 152) stages from six independent …
Figure 1—figure supplement 1
Analysis of the expression of non-muscle myosin II heavy chain (NMHC) paralogs during preimplantation development based on single cell RNA sequencing and MYH9-GFP fluorescence.
(A) Mouse single cell RNA sequencing analysis adapted from Deng et al., 2014, showing the expression levels of Myh9 (red), Myh10 (blue), and Myh14 (grey) at the zygote, four-cell, morula, and …
Figure 2 with 6 supplements
Multiscale analysis of morphogenesis in single maternal-zygotic Myh9 or Myh10 mutant embryos.
(A) Representative images of long-term time-lapse of WT, mzMyh9, and mzMyh10 embryos at the end of the 4-cell stage, start and end of the 8-cell stage, at the initiation of blastocoel formation and …
Figure 2—figure supplement 1
Macroscopic shape analysis of maternal-zygotic Myh9 and Myh10 mutant embryos.
(A) Representative images of E1.5 WT, mzMyh9, mzMyh10, and mzMyh9;mzMyh10 embryos. The zona pellucida is highlighted with a red ellipse. (B) Boxplot of the short/long axis ratio of an ellipse …
Figure 2—figure supplement 2
Morphogenesis of embryos with blastomeres fused at the 4-cell stage.
(A) Schematic diagram of 4-cell stage embryos with four cells (control) or after fusion with three cells, two cells of equal or unequal sizes. Representative images of long-term time-lapse of …
Figure 2—video 1
Preimplantation development of WT, mzMyh9, and mzMyh10 embryos.
Long-term time-lapse imaging of WT (left), mzMyh9 (middle), and mzMyh10 (right) embryos from the 4-cell stage onwards. Images are taken every 30 min; scale bar is 20 µm. Time is given in hours …
Figure 2—video 2
Periodic waves of contraction in WT, mzMyh9, mzMyh10, and mzMyh9;mzMyh10 embryos.
Short-term time-lapse imaging of WT (top left), mzMyh9 (top right), mzMyh10 (bottom left), and mzMyh9;mzMyh10 (bottom right) embryos. Images are taken every 5 s; scale bar is 20 µm.
Figure 2—video 3
Failed cleavage in mzMyh9 embryos.
Long-term time-lapse imaging of an mzMyh9 embryo during the third cleavage wave. Images are taken every 30 min; scale bar is 20 µm.
Figure 2—video 4
Preimplantation development of control, ¾, and ½ cell number embryos.
Long-term time-lapse imaging of control (top left), ¾ cell number (top right), and ½ cell number (after the fusion events of two pairs of blastomeres on bottom left or after the fusion of three …
Figure 3 with 2 supplements
Analysis of TE and ICM lineages in single maternal-zygotic Myh9 or Myh10 mutant embryos.
(A) Representative images of WT, mzMyh9, and mzMyh10 embryos stained for TE and ICM markers CDX2 (magenta) and SOX2 (cyan), DAPI (yellow), and F-actin (grey). The same mutant embryos as in Figure 2A …
Figure 3—figure supplement 1
Analysis of YAP localization in maternal-zygotic Myh9 and Myh10 mutant embryos.
(A) Representative images of WT, mzMyh9, mzMyh10, and multiple- or single-celled Myh9;Myh10 mutant embryos stained for YAP (magenta), DAPI (yellow), and F-actin (grey). Scale bar, 20 µm. (B) Mean …
Figure 3—figure supplement 2
Lineage specification of embryos with blastomeres fused at the 4-cell stage.
(A) Schematic diagram of 4-cell stage embryos with four cells (control) or after fusion with three cells, two cells of equal or unequal sizes. Representative images of embryos with four cells …
Figure 4
Apico-basal polarity of single maternal-zygotic Myh9 or Myh10 mutant embryos.
Representative images of WT, mzMyh9, and mzMyh10 embryos stained for apico-basal polarity markers. From left to right: PRKCz, F-actin, ATP1A1, CDH1, and AQP3 shown in grey. Nuclei stained with DAPI …
Figure 5 with 1 supplement
Multiscale analysis of morphogenesis in double maternal-zygotic Myh9 and Myh10 mutant embryos.
(A) Representative images of long-term time-lapse of WT and mzMyh9;mzMyh10 embryos at the end of the 4-cell stage, start and end of the 8-cell stage, at the initiation of blastocoel formation, and …
Figure 5—video 1
Preimplantation development of mzMyh9;mzMyh10 embryos.
Long-term time-lapse imaging of mzMyh9;mzMyh10 embryos forming an ICM (left), or without ICM (right) from the equivalent of the 2-cell stage onwards. Images are taken every 30 min; scale bar is 20 …
Figure 6
Analysis of TE and ICM lineage in double maternal-zygotic Myh9 and Myh10 mutant embryos.
(A) Representative images of WT and mzMyh9;mzMyh10 embryos stained for TE and ICM markers CDX2 (magenta) and SOX2 (cyan), DAPI (yellow), and F-actin (grey). The same mutant embryos as in Figure 5A …
Figure 7
Apico-basal polarity of maternal Myh9;Myh10 mutant embryos composed of multiple cells or a single cell.
Representative images of WT, multiple- or single-celled Myh9;Myh10 mutant embryos stained for apico-basal polarity markers. From left to right: PRKCz, F-actin, ATP1A1, CDH1, and AQP3 shown in grey. …
Figure 8 with 3 supplements
Single-celled embryos at the blastocyst stage.
(A) Representative images of long-term time-lapse of WT and single-celled mMyh9;mMyh10 embryos at the onset of fluid accumulation (Figure 8—video 1). Scale bar, 20 µm. (B) Embryo growth curves …
Figure 8—figure supplement 1
Morphogenesis and lineage specification of embryos with all blastomeres fused at the late morula stage.
(A) Representative images of long-term time-lapse of control and fused single-celled embryos at the onset of fluid accumulation (Figure 8—video 2). A schematic diagram of the cell fusion process is …
Figure 8—video 1
Preimplantation development of an mMyh9;mMyh10 embryo failing all successive cleavages.
Long-term time-lapse imaging of an mMyh9;mMyh10 embryo failing all successive cleavages. Images are taken every 30 min; scale bar is 20 µm. Time is given in hours following the last putative …
Figure 8—video 2
Fluid accumulation in control and single-celled fused embryos.
Long-term time-lapse imaging of control (left) and single-celled fused (right) embryos around the time of the fifth wave of cleavage onwards. Images are taken every 30 min; scale bar is 20 µm. Time …
Tables
Appendix 1—table 1
Statistical analysis of compaction data.
| Time after last third cleavage | Time before intial fourth cleavage | Time after last fourth cleavage | Time before intial fifth cleavage | |||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Mean | SEM | Embryos | p value | Mean | SEM | Embryos | p value | Mean | SEM | Embryos | p value | Mean | SEM | Embryos | p value | |||
| WT | Time after 3rd cleavage [h] | 0 | 0 | 23 | NA | 7.0 | 0.3 | 23 | NA | 10.9 | 0.3 | 21 | NA | 16.7 | 0.4 | 22 | NA | |
| Contact angle [°] | 87 | 3 | NA | 147 | 2 | NA | 140 | 3 | NA | 148 | 2 | NA | ||||||
| mzMyh9 | Time after 3rd cleavage [h] | 0 | 0 | 15 | NA | 9.8 | 0.5 | 15 | 0.0001 | 13.8 | 0.6 | 10 | 0.002 | 18.8 | 0.5 | 8 | 0.01 | |
| Contact angle [°] | 85 | 2 | 0.5 | 125 | 4 | 0.00001 | 104 | 5 | 0.00001 | 131 | 8 | 0.02 | ||||||
| mMyh9 | Time after 3rd cleavage [h] | 0 | 0 | 8 | NA | 10.4 | 0.7 | 8 | 0.001 | 14.6 | 0.8 | 8 | 0.002 | 18.7 | 0.7 | 3 | 0.002 | |
| Contact angle [°] | 86 | 5 | 0.5 | 115 | 6 | 0.0001 | 112 | 7 | 0.003 | 113 | 21 | 0.1 | ||||||
| mzMyh10 | Time after 3rd cleavage [h] | 0 | 0 | 11 | NA | 5.4 | 0.4 | 11 | 0.004 | 10.2 | 0.6 | 11 | 0.2 | 15.8 | 0.4 | 11 | 0.1 | |
| Contact angle [°] | 87 | 5 | 0.8 | 121 | 4 | 0.0001 | 126 | 5 | 0.03 | 148 | 4 | 0.8 | ||||||
| mMyh10 | Time after 3rd cleavage [h] | 0 | 0 | 20 | NA | 6.3 | 0.4 | 20 | 0.2 | 10.8 | 0.3 | 20 | 0.5 | 16.8 | 0.6 | 20 | 0.9 | |
| Contact angle [°] | 85 | 3 | 0.3 | 128 | 5 | 0.0004 | 128 | 3 | 0.006 | 149 | 2.8 | 0.5 | ||||||
| mzMyh9;mzMyh10 | Time after 3rd cleavage [h] | 0 | 0 | 7 | NA | 9.7 | 0.8 | 3 | 0.07 | 12 | NA | 1 | NA | NA | ||||
| Contact angle [°] | 89 | 6 | 0.8 | 117 | 4 | 0.006 | 84.5 | NA | NA | |||||||||
| mzMyh9;mMyh10 | Time after 3rd cleavage [h] | 0 | 0 | 7 | NA | 6.9 | 1.7 | 4 | 1.0 | 9 | 0.5 | 2 | 0.1 | 12 | NA | 1 | NA | |
| Contact angle [°] | 83 | 5 | 0.6 | 100 | 5 | 0.002 | 108 | 14 | 0.07 | 113 | NA | NA | ||||||
| mMyh9;mMyh10 | Time after 3rd cleavage [h] | 0 | 0 | 7 | NA | 8 | 1.3 | 5 | 0.5 | 11 | 0.5 | 3 | 1.0 | NA | ||||
| Contact angle [°] | 76 | 7 | 0.1 | 110 | 5 | 0.0006 | 105 | 3 | 0.002 | |||||||||
Appendix 1—table 2
Statistical analysis of cell cycle duration data.
Mean durations of 8-cell stage, fourth wave of cleavage, and 16-cell stage related to Figures 2B and 5B with associated SEM, embryo number, and p value from unpaired two-tailed Welch’s t test …
| Duration of 8-cell stage [h] | Duration of fourth wave of cleavages [h] | Duration of 16-cell stage [h] | ||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Mean | SEM | Embryos | p value | Mean | SEM | Embryos | p value | Mean | SEM | Embryos | p value | |
| WT | 7.0 | 0.3 | 23 | NA | 4.0 | 0.3 | 23 | NA | 5.5 | 0.4 | 23 | NA |
| mzMyh9 | 9.8 | 0.5 | 15 | 0.0001 | 4.8 | 0.7 | 15 | 0.3 | 5.7 | 0.8 | 14 | 0.8 |
| mMyh9 | 10.4 | 0.7 | 8 | 0.001 | 4.2 | 1.2 | 8 | 0.9 | 7.0 | 1.1 | 8 | 0.1 |
| mzMyh10 | 5.4 | 0.4 | 11 | 0.004 | 4.8 | 0.8 | 11 | 0.2 | 5.6 | 0.5 | 11 | 0.9 |
| mMyh10 | 6.3 | 0.4 | 20 | 0.2 | 4.5 | 0.4 | 20 | 0.3 | 6.1 | 0.5 | 20 | 0.4 |
| mzMyh9;mzMyh10 | 10.3 | 1.4 | 8 | 0.06 | 2.0 | 0.8 | 3 | 0.1 | ||||
| mzMyh9;mMyh10 | 8.1 | 1.4 | 6 | 0.5 | 4.3 | 1.1 | 3 | 0.8 | NA | |||
| mMyh9;mMyh10 | 9.3 | 1.0 | 8 | 0.06 | 3.6 | 0.7 | 5 | 0.6 | ||||
Appendix 1—table 3
Statistical analysis of periodic contraction data.
Mean amplitude of periodic movements and contact angle related to Figure 2H with associated SEM, embryo number, and p value from Mann-Whitney U test against WT.
| Maximum amplitude within 50–120 s oscillation period range [µm] | Contact angle [°] | ||||||
|---|---|---|---|---|---|---|---|
| Embryos | Mean | SEM | p value | Mean | SEM | p value | |
| WT | 21 | 0.140 | 0.009 | NA | 126 | 5 | NA |
| mzMyh9 | 15 | 0.039 | 0.002 | 0.0000005 | 78 | 3 | 0.000003 |
| mMyh9 | 8 | 0.045 | 0.003 | 0.00005 | 82 | 6 | 0.001 |
| mzMyh10 | 11 | 0.094 | 0.009 | 0.006 | 103 | 6 | 0.02 |
| mMyh10 | 20 | 0.111 | 0.013 | 0.01 | 108 | 5 | 0.02 |
