Cells are illuminated with pulses of blue light (illumination routine comprises 3 minutes imaging, 11 moments of activation and imaging – 1 activation every 45 mere seconds -, and 20 moments of imaging only)

Cells are illuminated with pulses of blue light (illumination routine comprises 3 minutes imaging, 11 moments of activation and imaging – 1 activation every 45 mere seconds -, and 20 moments of imaging only)

Cells are illuminated with pulses of blue light (illumination routine comprises 3 minutes imaging, 11 moments of activation and imaging – 1 activation every 45 mere seconds -, and 20 moments of imaging only). optoGEF-RhoA transmission. The bottom cluster (blue square) was illuminated (one activation every 10 mere seconds for 190 mere seconds) whereas the top cluster served Eniluracil as control (no illumination). Right: differential tractions relative to the -50s time point. Scale pub is definitely 20 m. Red scale bar is definitely 250 Pa and yellow scale bar is definitely 175 Pa. ncomms14396-s4.avi (33M) GUID:?4BE2FD87-F333-4270-AACF-B4848B01AC7D Supplementary Movie 4 OptoGEF-RhoA recruitment to mitochondria decreases contractility. Remaining: time development of traction causes exerted by MDCK cells expressing mito-CIBN51 GFP and optoGEF-RhoA during an activation protocol. Causes are overlaid within the optoGEF-RhoA transmission. The bottom cell (blue square) was illuminated (one activation every 10 mere seconds for 190 mere seconds) whereas the top cells served as control (no illumination). Right: differential tractions relative to the -50 mere seconds time point. Level bar is definitely 20 m. Red scale bar is definitely 1000 Pa and yellow scale bar is definitely 500 Pa. ncomms14396-s5.avi (15M) GUID:?4B2790DD-3753-476A-8AE8-C61D8C6A3A52 Supplementary Movie 5 Translocation of optoGEF-RhoA to the cellular membrane leads to basal actin accumulation. iRFP Eniluracil transmission of MDCK cells expressing lifeact-iRFP, CIBN-GFP-CAAX and optoGEF-RhoA. Cells are Rabbit Polyclonal to AKT1/2/3 (phospho-Tyr315/316/312) illuminated with pulses of blue light (illumination routine comprises 3 minutes imaging, 11 moments of activation and imaging – one activation every 45 mere seconds – and 20 moments of imaging only). Blue point shows when blue light is definitely on. Scale pub is definitely 10 m. ncomms14396-s6.avi (2.4M) GUID:?A0B3C77E-8840-4934-9EC4-8615FF45E2AF Supplementary Movie 6 Translocation 69 of optoGEF-RhoA to mitochondria is usually followed by actin depolymerization in the basal aircraft. iRFP transmission of MDCK cells expressing lifeact-iRFP, mito-CIBN-GFP and optoGEF-RhoA. Cells are illuminated with pulses of blue light (illumination routine consists of 3 minutes imaging, 11 moments of activation and imaging – one activation every 45 mere seconds-, and 20 moments of imaging only). Blue point shows when blue light is definitely on. Scale pub is definitely 10 m. ncomms14396-s7.avi (1.3M) GUID:?1B397FB8-9FB3-455A-A002-31DCF1012C5A Supplementary Movie 7 Translocation of optoGEF-RhoA to mitochondria is followed by loss of focal adhesions. iRFP transmission of MDCK cells expressing vinculin-iRFP, Eniluracil mito-CIBN-GFP and optoGEF-RhoA. Cells are illuminated with pulses of blue light (illumination routine comprises 3 minutes imaging, 11 moments of activation and imaging – one activation every 45 mere seconds -, and 20 moments of imaging only). Blue point shows when blue light is definitely on. Scale pub is definitely 10 m. ncomms14396-s8.avi (4.3M) GUID:?CEFCAE7D-84FA-4E2F-92BB-B316EE450D78 Supplementary Movie 8 A local increase in contractility induces tissue compaction. Myr-iRFP transmission (plasma membrane) of a confluent MDCK cell monolayer expressing myriRFP, CIBN-GFP-CAAX and optoGEF-RhoA. The area displayed from the blue dashed rectangle was illuminated at time t=0 (one pulse every 20 mere seconds for 5 minutes). PIV analysis of membrane velocity was performed every 40 mere seconds and is displayed by reddish arrows. Scale pub is definitely 20 m. Red scale bar is definitely 10? m/min. ncomms14396-s9.avi (17M) GUID:?D4977B11-1151-40FE-8334-6528F12956D7 Supplementary Movie 9 A local decrease in contractility triggers tissue expansion. Bright field images of a confluent MDCK cell monolayer co-expressing mito-CIBN-GFP and optoGEF-RhoA. The area displayed from the blue dashed rectangle was illuminated at time t=0 (one pulse every 20 mere seconds for 5 minutes). PIV analysis of velocity was performed every 40 mere seconds and is displayed by reddish arrows. Scale pub is definitely 20 m. Red scale bar is definitely 10 m/min. ncomms14396-s10.avi (17M) GUID:?8B45725A-768B-4393-A123-2388F33B42D6 Supplementary Movie 10 Cell contraction increases nuclear YAP. iRFP-YAP transmission of MDCK cells expressing Eniluracil iRFP-YAP, CIBN-GFP-CAAX and optoGEFRhoA. Cells are subjected to two activation periods, each of them followed by 60 moments without any activating light (activation routines consist of 7.5 minutes of imaging followed by 52.5 minutes of imaging and activation, one activation pulse.