Wnt11 family dependent morphogenesis during frog gastrulation is marked by the cleavage furrow protein anillin

2022 ◽  
Author(s):  
Elizabeth S Van Itallie ◽  
Christine M Field ◽  
Timothy J Mitchison ◽  
Marc W Kirschner

Wnt11 family proteins are ligands that activate a type of Dishevelled-mediated, non-canonical Wnt signaling pathway. Loss of function causes defects in gastrulation and/or anterior-posterior axis extension in all vertebrates. Non-mammalian vertebrate genomes encode two Wnt11 family proteins whose distinct functions have been unclear. We knocked down zygotic Wnt11b and Wnt11, separately and together, in Xenopus laevis. Single morphants exhibited very similar phenotypes of delayed blastopore closure, but they had different phenotypes at the tailbud stage. In response to their very similar gastrulation phenotypes, we chose to characterize dual morphants. Using dark field illuminated time-lapse imaging and kymograph analysis, we identified a failure of dorsal blastopore lip maturation that correlated with slower blastopore closure and failure to internalize the endoderm at the dorsal blastopore lip. We connected these externally visible phenotypes to cellular events in the internal tissues – including the archenteron – by imaging intact embryos stained for anillin and microtubules. The cleavage furrow protein anillin provided an exceptional cytological marker for blastopore lip and archenteron morphogenesis and the consequent disruption through loss of Wnt11 signaling. These cytological changes suggest a novel role for the regulation of contractility and stiffness of the epithelial cells that result in dramatic shape changes and are important in gastrulation.

2004 ◽  
Vol 166 (1) ◽  
pp. 49-60 ◽  
Author(s):  
Yoshihiro H. Inoue ◽  
Matthew S. Savoian ◽  
Takao Suzuki ◽  
Endre Máthé ◽  
Masa-Toshi Yamamoto ◽  
...  

We address the relative roles of astral and central spindle microtubules (MTs) in cytokinesis of Drosophila melanogaster primary spermatocytes. Time-lapse imaging studies reveal that the central spindle is comprised of two MT populations, “interior” central spindle MTs found within the spindle envelope and “peripheral” astral MTs that probe the cytoplasm and initiate cleavage furrows where they contact the cortex and form overlapping bundles. The MT-associated protein Orbit/Mast/CLASP concentrates on interior rather than peripheral central spindle MTs. Interior MTs are preferentially affected in hypomorphic orbit mutants, and consequently the interior central spindle fails to form or is unstable. In contrast, peripheral MTs still probe the cortex and form regions of overlap that recruit the Pav-KLP motor and Aurora B kinase. orbit mutants have disorganized or incomplete anillin and actin rings, and although cleavage furrows initiate, they ultimately regress. Our work identifies a new function for Orbit/Mast/CLASP and identifies a novel MT population involved in cleavage furrow initiation.


2018 ◽  
Vol 114 (8) ◽  
pp. 1115-1131 ◽  
Author(s):  
Marina Leone ◽  
Gentian Musa ◽  
Felix Benedikt Engel

Abstract Aims After birth mammalian cardiomyocytes initiate a last cell cycle which results in binucleation due to cytokinesis failure. Despite its importance for cardiac regenerative therapies, this process is poorly understood. Here, we aimed at a better understanding of the difference between cardiomyocyte proliferation and binucleation and providing a new tool to distinguish these two processes. Methods and results Monitoring of cell division by time-lapse imaging revealed that rat cardiomyocyte binucleation stems from a failure to properly ingress the cleavage furrow. Astral microtubule required for actomyosin ring anchorage and thus furrow ingression were not symmetrically distributed at the periphery of the equatorial region during anaphase in binucleating cardiomyocytes. Consequently, RhoA, the master regulator of actomyosin ring formation and constriction, non-muscle myosin IIB, a central component of the actomyosin ring, as well as IQGAP3 were abnormally localized during cytokinesis. In agreement with improper furrow ingression, binucleation in vitro and in vivo was associated with a failure of RhoA and IQGAP3 to localize to the stembody of the midbody. Conclusion Taken together, these results indicate that naturally occurring cytokinesis failure in primary cardiomyocytes is due to an aberrant mitotic microtubule apparatus resulting in inefficient anchorage of the actomyosin ring to the plasma cell membrane. Thus, cardiomyocyte binucleation and division can be discriminated by the analysis of RhoA as well as IQGAP3 localization.


2021 ◽  
Vol 7 (34) ◽  
pp. eabg2712
Author(s):  
Eslam M. F. Mehina ◽  
Stephanie Taylor ◽  
Roobina Boghozian ◽  
Emily White ◽  
Sun Eui Choi ◽  
...  

The cellular events that dictate the repair of damaged vessels in the brain, especially in those with vascular risk factors such as diabetes, is poorly understood. Here, we dissected the role of resident microglia and infiltrative macrophages in determining the repair of ruptured cerebral microvessels. Using in vivo time-lapse imaging, gene expression analysis, and immunohistochemistry, we identified a unique population of phagocytic Galectin 3 (Gal3) expressing macrophages, distinct from resident microglia, which infiltrated and aggregated at the site of injury in diabetic mice and were associated with the elimination of microvessels. Depletion of these infiltrative macrophages in diabetic mice attenuated phagocytic activity and prevented the loss of blood vessels after injury. These findings highlight a previously unknown role for infiltrative Gal3 expressing macrophages in promoting vessel elimination after brain injury and provide impetus for future studies to determine whether depleting these cells can facilitate vascular repair in at risk populations.


Author(s):  
A. M. Grigoriev ◽  
I. V. Kholodenko ◽  
A. Y. Lupatov ◽  
R. V. Kholodenko ◽  
L. A. Kirsanova ◽  
...  

Objective: to obtain long-lived proliferating cells with progenitor features by dedifferentiation of mature rat hepatocytes using combinations of small molecules.Materials and Methods. Hepatocytes isolated from rat liver by perfusion were cultured in the presence of a cocktail of three small molecules – Wnt signaling pathway activator (CHIR99021), TGF-β inhibitors (A83-01) and ROCK kinase (Y27632). The morphological characteristics and growth features of the culture were assessed using fluorescence and phase-contrast microscopy during cell culture. Cell proliferative activity was analyzed using real-time time-lapse imaging. The expression of surface and intracellular markers was analyzed using flow cytometry and high-resolution fluorescence microscopy.Results. Using a cocktail of small molecules, Y-27632, A-83-01, and CHIR99021, long-lived proliferating cells that express progenitor cell markers, such as α-fetoprotein and HNF4α, were obtained from mature rat hepatocytes. The cells had hepatocyte-like morphology and formed discrete clusters of proliferating cells, forming a single cell layer during culturing. Removal of the small molecules from the medium led to expansion of fibroblast-like cells and elimination of potentially progenitor hepatocyte-like cells.Conclusion. Proliferating progenitor cells can be obtained by dedifferentiation of mature hepatocytes.


2008 ◽  
Vol 4 (2) ◽  
pp. 71-81 ◽  
Author(s):  
Sarah Kucenas ◽  
Heather Snell ◽  
Bruce Appel

During development, multipotent neural precursors give rise to oligodendrocyte progenitor cells (OPCs), which migrate and divide to produce additional OPCs. Near the end of embryogenesis and during postnatal stages, many OPCs stop dividing and differentiate as myelinating oligodendrocytes, whereas others persist as nonmyelinating cells. Investigations of oligodendrocyte development in mice indicated that the Nkx2.2 transcription factor both limits the number of OPCs that are formed and subsequently promotes their differentiation, raising the possibility that Nkx2.2 plays a key role in determining myelinating versus nonmyelinating fate. We used in vivo time-lapse imaging and loss-of-function experiments in zebrafish to further explore formation and differentiation of oligodendrocyte lineage cells. Our data show that newly specified OPCs are heterogeneous with respect to gene expression and fate. Whereas some OPCs express the nkx2.2a gene and differentiate as oligodendrocytes, others that do not express nkx2.2a mostly remain as nonmyelinating OPCs. Similarly to mouse, loss of nkx2.2a function results in excess OPCs and delayed oligodendrocyte differentiation. Notably, excess OPCs are formed as a consequence of prolonged OPC production from neural precursor cells. We conclude that Nkx2.2 promotes timely specification and differentiation of myelinating oligodendrocyte lineage cells from species representing different vertebrate taxa.


Acta Naturae ◽  
2016 ◽  
Vol 8 (3) ◽  
pp. 88-96
Author(s):  
Yu. K. Doronin ◽  
I. V. Senechkin ◽  
L. V. Hilkevich ◽  
M. A. Kurcer

In order to estimate the diversity of embryo cleavage relatives to embryo progress (blastocyst formation), time-lapse imaging data of preimplantation human embryo development were used. This retrospective study is focused on the topographic features and time parameters of the cleavages, with particular emphasis on the lengths of cleavage cycles and the genealogy of blastomeres in 2- to 8-cell human embryos. We have found that all 4-cell human embryos have four developmental variants that are based on the sequence of appearance and orientation of cleavage planes during embryo cleavage from 2 to 4 blastomeres. Each variant of cleavage shows a strong correlation with further developmental dynamics of the embryos (different cleavage cycle characteristics as well as lengths of blastomere cycles). An analysis of the sequence of human blastomere divisions allowed us to postulate that the effects of zygotic determinants are eliminated as a result of cleavage, and that, thereafter, blastomeres acquire the ability of own syntheses, regulation, polarization, formation of functional contacts, and, finally, of specific differentiation. This data on the early development of human embryos obtained using noninvasive methods complements and extend our understanding of the embryogenesis of eutherian mammals and may be applied in the practice of reproductive technologies.


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