Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Mol Cells
2024 Jun 01;476:100076. doi: 10.1016/j.mocell.2024.100076.
Show Gene links
Show Anatomy links
Actin depolymerizing factor destrin governs cell migration in neural development during Xenopus embryogenesis.
Kim Y
,
Lee HK
,
Park KY
,
Ismail T
,
Lee H
,
Ryu HY
,
Cho DH
,
Kwon TK
,
Park TJ
,
Kwon T
,
Lee HS
.
???displayArticle.abstract???
The actin-based cytoskeleton is considered a fundamental driving force for cell differentiation and development. Destrin (Dstn), a member of the actin-depolymerizing factor family, regulates actin dynamics by treadmilling actin filaments and increasing globular actin pools. However, the specific developmental roles of dstn have yet to be fully elucidated. Here, we investigated the physiological functions of dstn during early embryonic development using Xenopus laevis as an experimental model organism. dstn is expressed in anterior neural tissue and neural plate during Xenopus embryogenesis. Depleting dstn promoted morphants with short body axes and small heads. Moreover, dstn inhibition extended the neural plate region, impairing cell migration and distribution during neurulation. In addition to the neural plate, dstn knockdown perturbed neural crest cell migration. Our data suggest new insights for understanding the roles of actin dynamics in embryonic neural development, simultaneously presenting a new challenge for studying the complex networks governing cell migration involving actin dynamics.
Fig. 1. dstn is expressed in anterior neural tissues and the epidermis during Xenopus embryogenesis. (A) Temporal expression of dstn analyzed by real-time qPCR at 9 landmark stages of embryonic development. (B) Spatial expression patterns of dstn were examined by whole-mount in situ hybridization (WISH). np, neural plate; an, anterior neural; ey, eye; ba, branchial arch; qPCR, quantitative polymerase chain reaction.
dstn (destrin, actin depolymerizing factor) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 16, dorsal view, anterior up.
dstn (destrin, actin depolymerizing factor) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 16, anterior view, dorsal up.
dstn (destrin, actin depolymerizing factor) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 22, lateral view, dorsal up.
dstn (destrin, actin depolymerizing factor) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 32, lateral view, dorsal up.
Fig. 2. dstn knockdown embryos show developmental malformations: short body axis and small-sized heads. (A) A graphical representation indicating dstn morpholino oligonucleotide (MO), dstn MO binding mRNA (dstn MOB)-Flag, dstn MO non-binding mRNA (dstn*)-Flag construct that codes for destrin but possess a mutated 5'-UTR, which dstn MO cannot target. Co-delivery of the dstn MO and rescue mRNA produced the same phenotypes as wild-type dstn mRNA. (B) Western blotting confirmed the efficacy of dstn MO. Anti-Beta-actin was used as a loading control. (C) dstn MO (40ng) was injected into the Xenopus embryos at the 2-cell stage, and the embryos were then fixed at developmental stages 31 and 41. White arrows represent the defects in tail development, and black arrows represent head and eye anomalies compared with control embryos. The statistical analysis showed that more than 60% of dstn morphants exhibited short and bent axes compared to control embryos. (D) Analysis of cartilage formation in dstn MO-injected embryos was conducted by fixing the embryos at stage 47, followed by staining with Alcian blue. The graphical representation indicated that more than 50% of dstn morphants showed small-sized heads compared to the control embryos. (E) Rescue experiments were performed by microinjecting the dstn** mRNA with dstn MO at the 2-cell stage of embryos. The developing embryos fixed at stage 32 showed that the phenotypic malformations of the small-sized head and bent axis were effectively recovered by co-injecting the dstn* and dstn MO. A graphical representation indicated that microinjection of dstn* mRNA with dstn MO considerably rescued phenotypic malformations as observed in dstn morphants. The significance levels are shown as *P<.05, **P<.01, ***P<.001, ****P<.0001, ns, not significant.
Fig. 3. dstn mediates neural plate extension and perturbs cell migration. (A) dstn MO was coinjected with beta-galactosidase (-gal) mRNA into 1 blastomere of 2-cell staged embryos, and the embryos were then fixed at the late neurula stage (NF. St.16). -gal staining indicated the injected side of the embryos. The length of the white-headed arrows exhibits the neural plate expansion in the dstn MO-injected side of the embryos compared to the shorter white-headed arrows in the control MO-injected side, as shown in the anterior and dorsal views. A graphical representation showed that approximately 50% of the dstn MO-injected embryos exhibited wider neural plates than the control embryos (scale bar: 250m). (B) Cross-sectional analysis of neurula stage embryos (NF. St.19) showed a difference in mesoderm distribution and position between the MO-injected side (B) and the uninjected side (A). Further, phalloidin immunostaining exhibited an accumulation of actin filaments (increased intensity) on the injected side of the embryos compared with the uninjected side. A graph representing the mean intensity of phalloidin staining showed a significant change in phalloidin intensity in dstn morphants compared to the control. Black arrows represent variation in the distribution and position of mesodermal cells (scale bar: 250m), no (notochord). (C) The dstn MO and -gal mRNA microinjections indicated that cell migration was highly limited. Black arrows represent the reduced and restricted cell migration in dorsal (only dorsal view for early-stage embryos) and lateral views of developing embryos. Cell migration was restricted only to the head regions in the dstn-depleted embryos compared to cell migration from the head to the tail axis in control embryos. Statistical analysis of dstn morphants and control embryos revealed significant cell migration restriction in dstn-depleted embryos compared to control embryos (scale bar: 250m). *P<.05, **P<.01, ***P<.001.
Fig. 4. dstn knockdown effects on neural crest specifiers during Xenopus embryogenesis. (A) Embryos were microinjected with dstn MO and -gal mRNA and analyzed for neural crest-specific genes, sox9, and snail2. The uninjected side of the embryos served as an internal control. Statistical analysis showed no significant change in the expression of neural crest specifiers on the dstn MO-injected side of the embryos compared to the uninjected and control MO-injected sides (scale bar: 250m). (B) WISH analysis of twist in stage 27 embryos injected with control MO or dstn MO. Statistical analysis indicated that 80% of dstn morphants had defective neural crest migration compared to control MO-injected embryos (scale bar: 500m). nc, neural crest. ***P<.001, ns, not significant.
