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Development of Microfluidic Devices to Study the Elongation Capability of Tip-growing Plant Cells in Extremely Small Spaces

In vivo, tip-growing plant cells need to overcome a series of physical barriers; however, researchers lack the methodology to visualize cellular behavior ...

Nagoya University

We herein report a 52-year-old man infected with human immunodeficiency virus (HIV) who was referred to our hospital due to the development of severe neurocognitive disorders and bilateral leukoencephalopathy. He has been treated with antiretroviral agents for 17 years, but low-level viremia has been detected consistently prior to admission. Drug resistant testing of the serum and the cerebrospinal fluid (CSF) both demonstrated a M184V mutation. A brain biopsy revealed perivascular CD8(+) T-lymphocyte infiltration, leading to the diagnosis of CD8 encephalitis. The clinical symptoms improved drastically after changing to a nucleoside reverse transcriptase inhibitor sparing regimen, which subsequently decreased the HIV viral load to an undetectable level in both the serum and CSF. 

CD8 Encephalitis Caused by Persistently Detectable Drug-resistant HIV.

Plant cells are covered with rigid cell walls, yet tip-growing cells can elongate by providing new cell wall material to their apical regions. Studies of the mechanical properties of tip-growing plant cells typically involve measurement of the turgor pressure and stiffness of the cells' apical regions. These experiments, however, do not address how living tip-growing cells react when they encounter physical obstacles that are not substantially altered by turgor pressure. To investigate this issue, we constructed microfabricated platforms with a series of artificial gaps as small as 1 μm, and examined the capability of tip-growing plant cells, including pollen tubes, root hairs, and moss protonemata, to penetrate into these gaps. The cells were grown inside microfluidic chambers and guided towards the gaps using microdevices customized for each cell type. All types of tip-growing cells could grow through the microgaps with their organelles intact, even though the gaps were much smaller than the cylindrical cell diameter. Our findings reveal the dramatic physiological and developmental flexibility of tip-growing plant cells. The microfluidic platforms designed in this study provide novel tools for the elucidation of the mechanical properties of tip-growing plant cells in extremely small spaces.

Capability of tip-growing plant cells to penetrate into extremely narrow gaps.

Stabilisation of minus ends of microtubules (MTs) is critical for organising MT networks in land plant cells, in which all MTs are nucleated independent of centrosomes. Recently, Arabidopsis SPIRAL2 (SPR2) protein was shown to localise to plus and minus ends of cortical MTs, and increase stability of both ends. Here, we report molecular and functional characterisation of SPR2 of the basal land plant, the moss Physcomitrella patens. In protonemal cells of P. patens, where non-cortical, endoplasmic MT network is organised, we observed SPR2 at minus ends, but not plus ends, of endoplasmic MTs and likely also of phragmoplast MTs. Minus end decoration was reconstituted in vitro using purified SPR2, suggesting that moss SPR2 is a minus end-specific binding protein (-TIP). We generated a loss-of-function mutant of SPR2, in which frameshift-causing deletions/insertions were introduced into all four paralogous SPR2 genes by means of CRISPR/Cas9. Protonemal cells of the mutant showed instability of endoplasmic MT minus ends. These results indicate that moss SPR2 is a MT minus end stabilising factor.Key words: acentrosomal microtubule network, microtubule minus end, P. patens, CAMSAP/Nezha/Patronin.

Naoki Yanagisawa

Division of Biological Science,
Graduate School of Science,
Division of Biological Science, Graduate School of Science

Nagoya University

Development of Microfluidic Devices to Study the Elongation Capability of Tip-growing Plant Cells in Extremely Small Spaces

Naoki Yanagisawa

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Division of Biological Science,Graduate School of Science,Division of Biological Science, Graduate School of Science

Nagoya University

Development of Microfluidic Devices to Study the Elongation Capability of Tip-growing Plant Cells in Extremely Small Spaces

Division of Biological Science,
Graduate School of Science,
Division of Biological Science, Graduate School of Science