Siliceous scales in the centrohelid heliozoan Raphidocystis contractilis facilitate settlement to the substratum

Aguirre, L.E., Ouyang, L., Elfwing, A., Hedblom, M., Wulff, A., Olle Inganäs, O. 2018. Diatom

frustules protect DNA from ultraviolet light. Sci. Rep. 8: 5138.

https://doi.org/10.1038/s41598-018-21810-2

Chen, C.S., Shiu, R.F., Hsieh, Y.Y., Xu, C., Vazquez, C. I., Cui, Y., Hsu, I. C., Quigg, A., Santschi, P. H.,

Chin, W.C., 2021. Stickiness of extracellular polymeric substances on different surfaces via

magnetic tweezers. Sci. Total Environ. 757: 143766.

https://doi.org/10.1016/j.scitotenv.2020.143766.

Drachko, D., Shɨshkin, Y., Zlatogursky, V.V. 2020. Phenotypic masquerade: Polymorphism in the

life cycle of the centrohelid heliozoan Raphidiophrys heterophryoidea (Haptista:

Centroplasthelida). Eur. J. Protistol. 73: 125686.

https://doi.org/10.1016/j.ejop.2020.125686.

Drachko, D., Shɨshkin. Y., Zlatogursky, V.V. 2022. On the phylogenetic position of Raphidocystis

pallida with some notes on its life cycle. J. Eukaryot. Microbiol. 69(4):e12916.

https://doi.org/10.1111/jeu.12916

Lee, B.P., Messersmith, P.B., Israelachvili, J. N., Waite, J.H., 2011. Mussel-inspired adhesives and

coatings. Annu. Rev. Mater. Res. 41: 99–132. https://doi.org/10.1146/annurev-matsci062910-100429.

Kinoshita, E., Suzaki, T., Sugiyama, M., Shigenaka, Y., 1995. Ultrastructure and rapid axopodial

contraction of a heliozoan, Raphidiophrys contractilis sp. nov. J. Eukaryot. Microbiol. 42:

283–288. https://doi.org/10.1111/j.1550-7408.1995.tb01581.x.

Knoll, A., Kotrc, B., 2015. Protistan skeletons: A geologic history of evolution and constraint.In:

Hamm, C. (eds) Evolution of lightweight structures. Biologically-inspired systems, vol. 6.

Springer, Dordrecht. https://doi:10.1007/978-94-017-9398-8_1.

Nicholls K., Dürrschmidt M., 1985. Scale structure and taxonomy of some species of Raphidocystis,

Raphidiophrys, and Pompholyxophrys (Heliozoea) including description of six new taxa. Can.

J. Zool. 63: 1944–1961. https://doi.org/10.1139/z85-288.

Pančić, M., Torres, R.R., Almeda, R., Kiørboe, T. 2019. Silicified cell walls as a defensive trait in

diatoms. Proc. R. Soc. B. 286: 20190184. https://doi.org/10.1098/rspb.2019.0184

Patterson, D.J., Dürrschmidt, M. 1988. The Formation of Siliceous Scales by Raphidiophrys

Ambigua (Protista, Centroheliozoa). J. Cell Sci. 91: 33-39.

https://doi.org/10.1242/jcs.91.1.33

Sakaguchi, M., Suzaki, T., Khan, S.M.M.K., Hausmann, K., 2002. Food capture by kinetocysts in the

heliozoon Raphidiophrys contractilis. Europ. J. Protistol. 37: 453–458.

https://doi.org/10.1078/0932-4739-00847.

Seabra, S., Zenleser, T., Grosbusch, A.L., Hobmayer, B., Lengerer, B., 2022. The involvement of celltype-specific glycans in Hydra temporary adhesion revealed by a lectin screen. Biomimetics

7: 166. https://doi.org/10.3390/biomimetics7040166.

Sumper, M., Brunner, E., 2008. Silica biomineralisation in diatoms: The model organism

Thalassiosira pseudonana. ChemBioChem. 9: 1187–1194.

https://doi.org/10.1002/cbic.200700764.

Textor, J., Sinn, M., de Boer, R.J., 2013. Analytical results on the Beauchemin model of lymphocyte

migration. BMC Bioinformatics 14 (Suppl 6), S10. https://doi.org/10.1186/1471-2105-14-S6S10.

Waite, J., 2008. Mussel power. Nature Mater. 7: 8–9. https://doi.org/10.1038/nmat2087.

Yoshimura, C., Kobayashi, M., Khan, S.M.M.K., Islam, M.D.S., Matsubara, S., Chen, L., Higuchi, R.,

Suzaki, T., 2017. Development of a compact, highly-sensitive and low-cost biological

monitoring method using protozoa for detecting toxicants in aquatic environment. Int. J.

Environ. Agr. Res. 3: 41–44. https://doi.org/10.25125/agriculture-journal-IJOEAR-JUL-20177.

Zagumyonnyi, D.G., Radaykina, L.V., Keeling, P.J., Denis V. Tikhonenkov, D.V., 2022. Centrohelid

heliozoans of Ukraine with a description of a new genus and species (Haptista:

Centroplasthelida). Europ. J. Protistol. 86: 125916.

https://doi.org/10.1016/j.ejop.2022.125916.

Zlatogursky, V.V., Drachko, D., Klimov, V.I., Shishkin, Y., 2018. On the phylogenetic position of the

genus Raphidocystis (Haptista: Centroplasthelida) with notes on the dimorphism in

centrohelid life cycle. Eur. J. Protistol. 64: 82–90.

https://doi.org/10.1016/j.ejop.2018.03.006.

Table 1. Free-fall velocity of wild-type and scale-deficient cells in water (10% ASW)

Strain

Velocity* (μm/s)

Sample number

Wild-type

5.22 ± 1.89

12

Scale-deficient strain

0.70 ± 0.30**

12

*mean ± standard deviation

**significantly different from the wild-type (Welch’s t-test, p < 0.01)

Fig. 1. Wild-type and scale-deficient strain cells of Raphidocystis contractilis. A and C show optical

micrographs of the wild-type strain (A) and the scale-deficient strain (C). Arrows indicate

phagocytic vesicles and arrowheads in A indicate scales found only in the wild strain. A magnified,

contrast-enhanced photograph of the cell surface of the scale-deficient strain (indicated by the

rectangle) is shown as an inset in C. In the scale-deficient strain, no scales are observed, but fine

needle-like structures are seen covering the cell surface (arrowheads in the inset in C, and in D). B

and D show transmission electron micrographs of negatively stained cell surface structures of the

wild-type (B) and the scale-deficient strain (D). Scale bars: A and C: 5 μm; B and D: 1 μm.

Fig. 2. Whole-mount specimens of the cell surface layers of Raphidocystis contractilis were

examined using a transmission electron microscope (JSM-7100F, JEOL) equipped with an energydispersive X-ray spectroscope (EDS, JED-2300, JEOL). The spectra in A and B show the signals for the

elements from the entire region shown in C for the wild and scale-deficient strains, respectively. C

shows two-dimensional elemental mapping images of C, O, Si, and Ca for wild-type and scaledeficient cells; the leftmost pictures in C show transmission electron microscope images of the

region where elemental analysis was performed. The scale of the wild strain showed silicon (Si) and

oxygen (O) signals (A and upper column in C). On the other hand, little silicon signal was detected in

the needle-like surface structure of the scale-deficient strain, where C and O were the major

elements detected.

Fig. 3. Relationship between the presence of scales and the degree of cell adhesion to the surface

of the coverslip. Wild-type or scale-deficient cells were placed on the coverslips and turned over

after a certain time (1 h or 1 d). The percentage of cells adhering to the coverslip was then

determined and shown as a box-and-whisker diagram. Both after 1 h and 1 d, the wild strain

showed stronger adhesion (*: Welch’s t-test, p < 0.01). The numbers below the boxes indicate the

number of independent measurements. Individual data are indicated by open circles.

Fig. 4. Percoll density-gradient centrifugation of cells. Wild-type and scale-deficient cells were

centrifuged in a discrete gradient of Percoll solution prepared by mixing 100% Percoll (density =

1.13 g/mL) and 10% ASW for 30 min at 2,300 × g. After centrifugation, wild-type cells were

separated into interfaces of different Percoll concentrations, forming five layers (A–E, with pictures

of cells corresponding to each layer shown). At the bottom of the tube, a pellet of scales detached

from the cells was observed (F), indicating that the density of scales is higher than 1.13 g/mL. In

contrast, scale-deficient cells accumulated at the interface alone between 20% and 30% Percoll

layers (G). Scale bars: 2 μm.

Fig. 5. A 3D image of a Raphidocystis contractilis wild-type cell showing its adhesion to the glass

surface. The cell suspension was placed on a coverslip and allowed to stand for 1 d. The cells were

then flipped upside down and consecutive photographs of the cells on the coverslip were taken

with a differential interference microscope at 1.5 μm intervals from the adherent surface to the

center of the cell body. The obtained photographs were reconstructed in 3D, in which orange

shows scales, green shows axopodia, and light blue shows the cell body. For clarity, the lower half

of the cell alone is depicted. As also shown in the reconstructed rotational movie

(https://youtu.be/h30k9aFHcOk), the cell was attached to the coverslip using scales in addition to

the tips of the axopodia. On the surface of the glass substratum, some scales were observed that

had fallen off from the cell body and remained attached to the substratum (arrows).

Fig. 6. The trajectory of the migratory movement of cells on the glass substratum traced at 10 s

intervals for 10 min. The trajectories of 16 cells were superimposed using the cell position at the

start of imaging as the origin. The central enlargement is shown on the right for wild-type (A) and

scale-deficient strain (B), respectively. Wild-type cells show less movement than scale-deficient

cells do. In C, root mean square (RMS) displacement of cells was plotted as a function of time. In

both wild-type and scale-deficient cells, RMS displacement was linear, which is a characteristic of

directed motion with cell migration velocities of 0.04 μm/s in the wild-type and 0.39 μm/s in the

scale-deficient cells.

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