Preparation of three‐notochord explants for imaging analysis of the cell movements of convergent extension during early Xenopus morphogenesis

Focal adhesion kinase (FAK) mediates vital cellular pathways during development. Despite its necessity, how FAK regulates and integrates with other signals during early embryogenesis remains poorly understood. We found that the loss of Fak1a impaired epiboly, convergent extension and hypoblast cell migration in zebrafish embryos. We also observed a clear disturbance in cortical actin at the blastoderm margin and distribution of yolk syncytial nuclei. In addition, we investigated a possible link between Fak1a and a well-known gastrulation regulator, Wnt5b, and revealed that the overexpression of fak1a or wnt5b could cross-rescue convergence defects induced by a wnt5b or fak1a antisense morpholino (MO), respectively. Wnt5b and Fak1a were shown to converge in regulating Rac1 and Cdc42, which could synergistically rescue wnt5b and fak1a morphant phenotypes. Furthermore, we generated several alleles of fak1a mutants using CRISPR/Cas9, but those mutants only revealed mild gastrulation defects. However, injection of a subthreshold level of the wnt5b MO induced severe gastrulation defects in fak1a mutants, which suggested that the upregulated expression of wnt5b might complement the loss of Fak1a. Collectively, we demonstrated that a functional interaction between Wnt and FAK signalling mediates gastrulation cell movements via the possible regulation of Rac1 and Cdc42 and subsequent actin dynamics.

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Furry is required for cell movements during gastrulation and functionally interacts with NDR1

10.1101/2020.05.08.083980 ◽

2020 ◽

Author(s):

Ailen S. Cervino ◽

Bruno Moretti ◽

Carsten Stuckenholz ◽

Hernán E. Grecco ◽

Lance A. Davidson ◽

...

Keyword(s):

Asymmetric Cell Division ◽

Cell Polarization ◽

Convergent Extension ◽

Cellular Functions ◽

Embryonic Axes ◽

Cell Movements ◽

Evolutionarily Conserved ◽

Axis Elongation ◽

Established Cell ◽

Division Cell

AbstractGastrulation is a key event in animal embryogenesis during which the germ layers precursors are rearranged and the embryonic axes are established. Cell polarization is essential during gastrulation driving asymmetric cell division, cell movements and cell shape changes. Furry (Fry) gene encodes an evolutionarily conserved protein with a wide variety of cellular functions mostly related to cell polarization and morphogenesis in invertebrates. However, little is known about its function in vertebrate development. Here we show that in Xenopus, Fry participates in the regulation of morphogenetic processes during gastrulation. Using morpholino knock-down, we demonstrate a role of Fry in blastopore closure and dorsal axis elongation. Loss of Fry function drastically affects the movement and morphological polarization of cells during gastrulation, in addition to dorsal mesoderm convergent extension, responsible for head-to-tail elongation. Finally, we demonstrate a functional interaction between Fry and NDR1 kinase, providing evidence of an evolutionarily conserved complex required for morphogenesis.

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Cell movements during epiboly and gastrulation in zebrafish

Development ◽

10.1242/dev.108.4.569 ◽

1990 ◽

Vol 108 (4) ◽

pp. 569-580 ◽

Cited By ~ 69

Author(s):

R.M. Warga ◽

C.B. Kimmel

Keyword(s):

Dorsal Side ◽

Epithelial Layer ◽

Convergent Extension ◽

Blastula Stage ◽

Cell Movements ◽

Yolk Cell ◽

Radial Cell ◽

Convergence And Extension ◽

Embryonic Ectoderm ◽

Anterior Posterior

Beginning during the late blastula stage in zebrafish, cells located beneath a surface epithelial layer of the blastoderm undergo rearrangements that accompany major changes in shape of the embryo. We describe three distinctive kinds of cell rearrangements. (1) Radial cell intercalations during epiboly mix cells located deeply in the blastoderm among more superficial ones. These rearrangements thoroughly stir the positions of deep cells, as the blastoderm thins and spreads across the yolk cell. (2) Involution at or near the blastoderm margin occurs during gastrulation. This movement folds the blastoderm into two cellular layers, the epiblast and hypoblast, within a ring (the germ ring) around its entire circumference. Involuting cells move anteriorwards in the hypoblast relative to cells that remain in the epiblast; the movement shears the positions of cells that were neighbors before gastrulation. Involuting cells eventually form endoderm and mesoderm, in an anterior-posterior sequence according to the time of involution. The epiblast is equivalent to embryonic ectoderm. (3) Mediolateral cell intercalations in both the epiblast and hypoblast mediate convergence and extension movements towards the dorsal side of the gastrula. By this rearrangement, cells that were initially neighboring one another become dispersed along the anterior-posterior axis of the embryo. Epiboly, involution and convergent extension in zebrafish involve the same kinds of cellular rearrangements as in amphibians, and they occur during comparable stages of embryogenesis.

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Furry is required for cell movements during gastrulation and functionally interacts with NDR1

Scientific Reports ◽

10.1038/s41598-021-86153-x ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Ailen S. Cervino ◽

Bruno Moretti ◽

Carsten Stuckenholz ◽

Hernán E. Grecco ◽

Lance A. Davidson ◽

...

Keyword(s):

Asymmetric Cell Division ◽

Cell Polarization ◽

Convergent Extension ◽

Cellular Functions ◽

Embryonic Axes ◽

Cell Movements ◽

Evolutionarily Conserved ◽

Dorsal Mesoderm ◽

Axis Elongation ◽

Division Cell

AbstractGastrulation is a key event in animal embryogenesis during which germ layer precursors are rearranged and the embryonic axes are established. Cell polarization is essential during gastrulation, driving asymmetric cell division, cell movements, and cell shape changes. Thefurry(fry) gene encodes an evolutionarily conserved protein with a wide variety of cellular functions, including cell polarization and morphogenesis in invertebrates. However, little is known about its function in vertebrate development. Here, we show that inXenopus,Fry plays a role in morphogenetic processes during gastrulation, in addition to its previously described function in the regulation of dorsal mesoderm gene expression. Using morpholino knock-down, we demonstrate a distinct role for Fry in blastopore closure and dorsal axis elongation. Loss of Fry function drastically affects the movement and morphological polarization of cells during gastrulation and disrupts dorsal mesoderm convergent extension, responsible for head-to-tail elongation. Finally, we evaluate a functional interaction between Fry and NDR1 kinase, providing evidence of an evolutionarily conserved complex required for morphogenesis.

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Twinfilin1 controls lamellipodial protrusive activity and actin turnover during vertebrate gastrulation

10.1101/2020.09.03.281659 ◽

2020 ◽

Author(s):

Caitlin C. Devitt ◽

Chanjae Lee ◽

Rachael M. Cox ◽

Ophelia Papoulas ◽

José Alvarado ◽

...

Keyword(s):

Animal Cell ◽

Dynamic Control ◽

Deep Understanding ◽

Early Embryo ◽

Cell Polarization ◽

Actin Dynamics ◽

Convergent Extension ◽

Cell Movements ◽

Actin Turnover

AbstractThe dynamic control of the actin cytoskeleton is a key aspect of essentially all animal cell movements. Experiments in single migrating cells and in vitro systems have provided an exceptionally deep understanding of actin dynamics. However, we still know relatively little of how these systems are tuned in cell-type specific ways, for example in the context of collective cell movements that sculpt the early embryo. Here, we provide an analysis of the actin severing and depolymerization machinery during vertebrate gastrulation, with a focus on Twinfilin1. We confirm previous results on the role of Twf1 in lamellipodia and extend those findings by linking Twf1, actin turnover, and cell polarization required for convergent extension during vertebrate gastrulation.

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Twinfilin1 controls lamellipodial protrusive activity and actin turnover during vertebrate gastrulation

Journal of Cell Science ◽

10.1242/jcs.254011 ◽

2021 ◽

Author(s):

Caitlin C. Devitt ◽

Chanjae Lee ◽

Rachael M. Cox ◽

Ophelia Papoulas ◽

José Alvarado ◽

...

Keyword(s):

Animal Cell ◽

Dynamic Control ◽

Deep Understanding ◽

Early Embryo ◽

Actin Dynamics ◽

Convergent Extension ◽

Cell Movements ◽

Actin Cables

The dynamic control of the actin cytoskeleton is a key aspect of essentially all animal cell movements. Experiments in single migrating cells and in vitro systems have provided an exceptionally deep understanding of actin dynamics. However, we still know relatively little of how these systems are tuned in cell-type specific ways, for example in the context of collective cell movements that sculpt the early embryo. Here, we provide an analysis of the actin severing and depolymerization machinery during vertebrate gastrulation, with a focus on Twinfilin1 (Twf1). We find that Twf1 is essential for convergent extension, and loss of Twf1 results in a failure of lamellipodial dynamics and polarity. Moreover, Twf1 loss results in a failure to assemble polarized cytoplasmic actin cables essential for convergent extension. These data provide an in vivo complement to our more-extensive understanding of Twf1 action in vitro and provide new links between the core machinery of actin regulation and specialized cell behaviors of embryonic morphogenesis.

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