Protein-tyrosine phosphatases in zebrafish gastrulation

TY - THES

T1 - Protein-tyrosine phosphatases in zebrafish gastrulation

AU - van Eekelen, M.J.L.

N1 - Reporting year: 2011

PY - 2011

Y1 - 2011

N2 - Protein tyrosine phosphorylation plays a key role in relaying external stimuli and signals into the cell towards the appropriate responses. This process is mediated by protein-tyrosine kinases adding a phosphor group to a tyrosine residue and protein-tyrosine phosphatases removing a phosphor group from phosphorylated tyrosines. The addition and removal of a phosphor group to can result in changed protein folding, changed catalytic activity or changed binding affinity for interacting partners. The early development of the vertebrae is characterized by three cellular processes; proliferation, differentiation and cell migration. During gastrulation a few key cell movements can be distinguished, namely epiboly and emboly which result in the formation of the three different germ layers, and convergence and extension cell movements which result in the formation of the primary body axis. Convergence and extension cell movements are characterized be polarization and elongation of mesodermal and neurectodermal cells. These cells migrate towards the future dorsal midline, where they intercalate and elongate the dorsal midline. Impaired convergence and extension cell movements typically lead to less cells reaching the dorsal midline and decreased body axis elongation, hence shorter and fatter animals. Several pathways have been identified to play a role in convergence and extension cell movements, of which the non-canonical Wnt signaling pathway is the most important. This pathway is also known as the planar cell polarity pathway, and was first identified in the fly (drosophila), where it regulates the growth of wing hairs on the distal tips of epithelial cells. This path proved to be crucial to setup a second polarity (proximal-distal) within an already polarized tissue (apical-basal). Further findings show that the same pathway can regulate polarity in non-epidermal tissues like cells undergoing convergence/extension movements as well. The non-canonical Wnt signaling pathway has been shown to regulate and stimulate RhoA and Rac1 activity, as opposed to the canonical Wnt pathway which results in β-catenin stability and resulting transcriptional regulation. RhoA and Rac1 are small GTPases involved in actin skeletal rearrangements, cell polarity and cell migration. Recently several reports identified other pathways regulating these small GTPases, independent of the non-canonical Wnt signaling pathway. We set out to identify new pathways involved in convergence and extension cell movements by screening all classical protein-tyrosine phosphatases in the zebrafish. The zebrafish is a god model for such screens because of the fast development, transparency and ease of genetic manipulation by knockdown techniques. We started by identifying all classical protein-tyrosine phosphatase encoding genes in the zebrafish genome and establishing their expression patterns in early development. We next proceeded to knockdown each of these genes using synthetic antisense RNA (morpholinos), screening for possible convergence and extension related phenotypes (shorter and thicker fish embryos). keywords: Phosphatase, Tyrosine, Zebrafish, Gastrulation, Polarity, Cell movements, Convergence, Extension, Ephrin-B, Planar cell polarity

AB - Protein tyrosine phosphorylation plays a key role in relaying external stimuli and signals into the cell towards the appropriate responses. This process is mediated by protein-tyrosine kinases adding a phosphor group to a tyrosine residue and protein-tyrosine phosphatases removing a phosphor group from phosphorylated tyrosines. The addition and removal of a phosphor group to can result in changed protein folding, changed catalytic activity or changed binding affinity for interacting partners. The early development of the vertebrae is characterized by three cellular processes; proliferation, differentiation and cell migration. During gastrulation a few key cell movements can be distinguished, namely epiboly and emboly which result in the formation of the three different germ layers, and convergence and extension cell movements which result in the formation of the primary body axis. Convergence and extension cell movements are characterized be polarization and elongation of mesodermal and neurectodermal cells. These cells migrate towards the future dorsal midline, where they intercalate and elongate the dorsal midline. Impaired convergence and extension cell movements typically lead to less cells reaching the dorsal midline and decreased body axis elongation, hence shorter and fatter animals. Several pathways have been identified to play a role in convergence and extension cell movements, of which the non-canonical Wnt signaling pathway is the most important. This pathway is also known as the planar cell polarity pathway, and was first identified in the fly (drosophila), where it regulates the growth of wing hairs on the distal tips of epithelial cells. This path proved to be crucial to setup a second polarity (proximal-distal) within an already polarized tissue (apical-basal). Further findings show that the same pathway can regulate polarity in non-epidermal tissues like cells undergoing convergence/extension movements as well. The non-canonical Wnt signaling pathway has been shown to regulate and stimulate RhoA and Rac1 activity, as opposed to the canonical Wnt pathway which results in β-catenin stability and resulting transcriptional regulation. RhoA and Rac1 are small GTPases involved in actin skeletal rearrangements, cell polarity and cell migration. Recently several reports identified other pathways regulating these small GTPases, independent of the non-canonical Wnt signaling pathway. We set out to identify new pathways involved in convergence and extension cell movements by screening all classical protein-tyrosine phosphatases in the zebrafish. The zebrafish is a god model for such screens because of the fast development, transparency and ease of genetic manipulation by knockdown techniques. We started by identifying all classical protein-tyrosine phosphatase encoding genes in the zebrafish genome and establishing their expression patterns in early development. We next proceeded to knockdown each of these genes using synthetic antisense RNA (morpholinos), screening for possible convergence and extension related phenotypes (shorter and thicker fish embryos). keywords: Phosphatase, Tyrosine, Zebrafish, Gastrulation, Polarity, Cell movements, Convergence, Extension, Ephrin-B, Planar cell polarity

M3 - PhD thesis (Proefschrift)

SN - 978-90-393-55497

PB - Utrecht University

CY - Utrecht

ER -