[
Methods,
2003]
We show that structural protein arrays consisting largely of collagen, myosin, and tubulin, and their associated proteins can be imaged in three dimensions with high contrast and resolution by laser-scanning second harmonic generation (SHG) microscopy. SHG is a nonlinear optical scheme and this form of microscopy shares several common advantages with multiphoton excited fluorescence, namely, intrinsic three-dimensionality and reduced out-of-plane photobleaching and phototoxicity. SHG does not arise from absorption and in-plane photodamage considerations are therefore also greatly reduced. In particular, structural protein arrays that are highly ordered and birefringent produce large SHG signals without the need for any exogenous labels. We demonstrate that thick tissues including muscle and bone can be imaged and sectioned through several hundred micrometers of depth. Combining SHG with two-photon excited green fluorescent protein (GFP) imaging allows inference of the molecular origin of the SHG contrast in Caenorhabditis elegans sarcomeres. Symmetry and organization of microtubule structures in dividing C. elegans embryos are similarly studied by comparing the endogenous tubulin contrast with that of GFP::tubulin fluorescence. It is found that SHG provides molecular level data on radial and lateral symmetries that GFP constructs cannot. The physical basis of SHG is discussed and compared with that of two-photon excitation as well as that of polarization microscopy. Due to the intrinsic sectioning, lack of photobleaching, and availability of molecular level data, SHG is a powerful tool for in vivo imaging.
[
Front Cell Dev Biol,
2020]
Cell invasion is defined by the capability of cells to migrate across compartment boundaries established by basement membranes (BMs). The development of complex organs involves regulated cell growth and regrouping of different cell types, which are enabled by controlled cell proliferation and cell invasion. Moreover, when a malignant tumor takes control over the body, cancer cells evolve to become invasive, allowing them to spread to distant sites and form metastases. At the core of the switch between proliferation and invasion are changes in cellular morphology driven by remodeling of the cytoskeleton. Proliferative cells utilize their actomyosin network to assemble a contractile ring during cytokinesis, while invasive cells form actin-rich protrusions, called invadopodia that allow them to breach the BMs. Studies of developmental cell invasion as well as of malignant tumors revealed that cell invasion and proliferation are two mutually exclusive states. In particular, anchor cell (AC) invasion during <i>Caenorhabditis elegans</i> larval development is an excellent model to study the transition from cell proliferation to cell invasion under physiological conditions. This mini-review discusses recent insights from the <i>C. elegans</i> AC invasion model into how G1 cell-cycle arrest is coordinated with the activation of the signaling networks required for BM breaching. Many regulators of the proliferation-invasion network are conserved between <i>C. elegans</i> and mammals. Therefore, the worm may provide important clues to better understand cell invasion and metastasis formation in humans.
[
Biology of the Cell,
1999]
In the Caenorhabditis elegans hermaphrodite, the establishment of the egg-laying system requires the connection of two epithelial tubes: the uterus of the gonad and the vulva in the underlying ectoderm. A specialized uterine cell, the anchor cell (AC), plays a central role in specifying the fates of the uterine and vulval precursor cells via the EGF-Ras-MAP kinase and the Notch/Delta signaling pathways. This central and common inducing source ensures that the two sets of cells are in register and it specifies the cell types that build the T-shaped connection between uterus and vulva. On either side, progeny of the induced cells form lumen structures and undergo stereotyped cell-to-cell fusion, thereby building epithelial tubes. Finally, the anchor cell fuses with a uterine syncytium and thus leaves only a thin cellular process between the lumen of the uterus and the vulva. In the adult, the fertilized eggs exit the lumen of the uterus through the vulva. This relatively simple developmental process serves as a model to study the biology of cells during organogenesis, such as intercellular signaling, cell polarity, invasion of basal laminae and epithelia, cell recognition and cell fusion. The anchor cell is a particularly interesting cell as it coordinates the development of its neighboring cells by using different signaling pathways at different times.