Journal
COMMUNICATIONS BIOLOGY
Volume 5, Issue 1, Pages -Publisher
NATURE PORTFOLIO
DOI: 10.1038/s42003-022-03835-6
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Funding
- Ministry of Science and Technology, Taiwan [MOST 109-2113-M-001 -034 -MY3, 109-2628M-001-001-MY4, 110-2321-B-002-012, MOST 110-2636-B-007-011, MOST 111-2636-B-007-009]
- Academia Sinica, Taipei, Taiwan [AS-IA-110-M04, AS-CDA-107-M08]
- National Tsing Hua University, Hsinchu, Taiwan [111Q2713E1]
- Academia Sinica Core Facility and Innovative Instrument Project [AS-CFII-108-115]
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This study improves lattice lightsheet microscopy (LLSM) to manipulate cellular behavior with subcellular resolution through optogenetic activation (optoLLSM). By integrating a position-controllable Bessel beam as a stimulation source, spatiotemporal photoactivation can be achieved by changing spatial light modulator (SLM) patterns. The study demonstrates the ability to control membrane ruffling and cell migration behavior using light-triggered recruitment of phosphoinositide 3-kinase and guided cell migration with low energy power.
Lattice lightsheet microscopy (LLSM) featuring three-dimensional recording is improved to manipulate cellular behavior with subcellular resolution through optogenetic activation (optoLLSM). A position-controllable Bessel beam as a stimulation source is integrated into the LLSM to achieve spatiotemporal photoactivation by changing the spatial light modulator (SLM) patterns. Unlike the point-scanning in a confocal microscope, the lattice beams are capable of wide-field optical sectioning for optogenetic activation along the Bessel beam path.We show that the energy power required for optogenetic activations is lower than 1 nW (or 24 mWcm(-2)) for time-lapses of CRY2olig clustering proteins, and membrane ruffling can be induced at different locations within a cell with subcellular resolution through light-triggered recruitment of phosphoinositide 3-kinase. Moreover, with the epidermal growth factor receptor (EGFR) fused with CRY2olig, we are able to demonstrate guided cell migration using optogenetic stimulation for up to 6 h, where 463 imaging volumes are collected, without noticeable cellular damages. Using a Bessel beam as a simulation source allows the use of lattice lightsheet microscopy for spatiotemporal control of photoactivation, illustrated by the control of cellular migration behavior.
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