Quantitative assessment of chemotropism in pollen tubes using microslit channel filters

Yanagisawa, Naoki; Higashiyama, Tetsuya

doi:10.1063/1.5023718

Cited by 9 publications

(6 citation statements)

References 22 publications

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“…In the last years, several microfluidic devices were designed for the small-scale investigation of growth responses of bacteria, tumor cell lines, yeast but also multicellular organisms by the application of fluids, which overflow the biological material (Moore et al, 2008;Brett et al, 2012;Choi et al, 2012;Yanagisawa and Higashiyama, 2018). Unicellular organisms can be investigated quite easily with these systems: Liquid chemical gradients are established and can be changed by the utilization of pumps applying solutions from source to sink.…”

Section: Discussionmentioning

confidence: 99%

“…Cells, which aim to reach the ovary, have to grow toward a solid matrix of agar and fulfill two directional changes. Due to this dual direction change, the number of false results is minimized (Yanagisawa and Higashiyama, 2018). Similar as described for pollen tubes, fungi are prone to grow toward gradients established in solid matrices by polar elongation.…”

Section: Discussionmentioning

confidence: 99%

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A 3D Printed Device for Easy and Reliable Quantification of Fungal Chemotropic Growth

2020

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Chemical gradients are surrounding living organisms in all habitats of life. Microorganisms, plants and animals have developed specific mechanisms to sense such gradients. Upon perception, chemical gradients can be categorized either as favorable, like nutrients or hormones, or as disadvantageous, resulting in a clear orientation toward the gradient and avoiding strategies, respectively. Being sessile organisms, fungi use chemical gradients for their orientation in the environment. Integration of this data enables them to successfully explore nutrient sources, identify probable plant or animal hosts, and to communicate during sexual reproduction or early colony development. We have developed a 3D printed device allowing a highly standardized, rapid and low-cost investigation of chemotropic growth processes in fungi. Since the 3D printed device is placed on a microscope slide, detailed microscopic investigations and documentation of the chemotropic process is possible. Using this device, we provide evidence that germlings derived from oval conidia of the hemibiotrophic plant pathogen Colletotrichum graminicola can sense gradients of glucose and reorient their growth toward the nutrient source. We describe in detail the method establishment, probable pitfalls, and provide the original program files for 3D printing to enable broad application of the 3D device in basic, agricultural, medical, and applied fungal science.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

A 3D Printed Device for Easy and Reliable Quantification of Fungal Chemotropic Growth

2020

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“…In the previously reported culture chamber for Torenia fournieri pollen tubes, a cut pistil was directly inserted into a reservoir in the device in order to guide them inside the fluidic channels. 11 This approach, however, is most likely not applicable for plants whose pistil's length is very short, such as A. thaliana , the most popular model species for plant biology. Although a cut pistil of Arabidopsis can be accommodated in an open microfluidic chamber, 20 the fluid manipulation involved with sample plug injection in such an open microfluidic format is not feasible.…”

Section: Resultsmentioning

confidence: 99%

“…However, widely used electrokinetic injection methods are not desirable for studies that involve living cells due to their exposure to electric elds. Therefore, all the reported microuidic devices for pollen tube chemotropism assays are performed under static 11,12 or pressure-driven ow 13 (PDF) conditions. While pressure gradients can also be generated on-chip by integrating electro-osmotic (EO) pumps into the device, 14 dynamic PDF control in microchannels through the operation of EO pumps is challenging or not even feasible when physiologically relevant cell culture medium (e.g., highly conductive liquid medium) is used for the experiments.…”

Section: Introductionmentioning

confidence: 99%

Pulsatile reverse flow actuated microfluidic injector: toward the application for single-molecule chemotropism assay

2021

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“…A variety of microfluidic devices has been developed that can generate in vitro chemical gradients to study diffusion of signaling molecules with a range of different length and time scales [9][10][11] or can perform quantitative analyses of pollen tube guidance. 12,13 Microfluidic devices have been used to study the mating response in yeast and provided new insights. [14][15][16][17] For example, Brett et al managed to produce and rapidly rotate stable pheromone gradients in a microfluidic device allowing the observation of initial orientation and re-orientation in yeast chemotropism.…”

Section: Introductionmentioning

confidence: 99%

Chemotropism among populations of yeast cells with spatiotemporal resolution in a biofabricated microfluidic platform

Shah

Choy

et al. 2020

Biomicrofluidics

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Chemotropism is an essential response of organisms to external chemical gradients that direct the growth of cells toward the gradient source. Chemotropic responses between single cells have been studied using in vitro gradients of synthetically derived signaling molecules and helped to develop a better understanding of chemotropism in multiple organisms. However, dynamic changes including spatial changes to the gradient as well as fluctuations in levels of cell generated signaling molecules can result in the redirection of chemotropic responses, which can be difficult to model with synthetic peptides and single cells. An experimental system that brings together populations of cells to monitor the population-scale chemotropic responses yet retain single cell spatiotemporal resolution would be useful to further inform on models of chemotropism. Here, we describe a microfluidic platform that can measure the chemotropic response between populations of mating yeast A-and α-cells with spatiotemporal programmability and sensitivity by positioning cell populations side by side in calcium alginate hydrogels along semipermeable membranes with micrometer spatial control. The mating phenotypes of the yeast populations were clearly observed over hours. Three distinct responses were observed depending on the distance between the A-and α-cell populations: the cells either continued to divide, arrest, and develop a stereotypical polarized projection termed a "shmoo" toward the cells of opposite mating type or formed shmoos in random directions. The results from our studies of yeast mating suggest that the biofabricated microfluidic platform can be adopted to study population-scale, spatial-sensitive cell-cell signaling behaviors that would be challenging using conventional approaches.

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Quantitative assessment of chemotropism in pollen tubes using microslit channel filters

Cited by 9 publications

References 22 publications

A 3D Printed Device for Easy and Reliable Quantification of Fungal Chemotropic Growth

A 3D Printed Device for Easy and Reliable Quantification of Fungal Chemotropic Growth

Pulsatile reverse flow actuated microfluidic injector: toward the application for single-molecule chemotropism assay

Chemotropism among populations of yeast cells with spatiotemporal resolution in a biofabricated microfluidic platform

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