Optogenetic control of BMPRs with β3 integrin maintains the recruitment of ALK3 to focal adhesion sites and the exclusion of BMPRII from focal adhesions. (A and B) Representative images of MEFsv40 cells coexpressing opto-CAAX/opto-β3 integrin, opto-ALK3/opto-β3 integrin, opto-BMPRII/opto-β3 integrin, opto-ALK3Ca/opto-β3 integrin, or opto-ALK3Ci/opto-β3 integrin observed by TIRF imaging (A) before and (B) after pulses of blue laser light stimulation for 3 min (10-s frequency) to induce proximity between BMPRs and β3 integrin by the iLID/SspB system. Opto-CAAX and all opto-ALK3s were recruited to β3 integrin-containing FAs, whereas opto-BMPRII was not recruited. (C) The normalized intensity profile in FA sites shows the increase in RFP fluorescence intensity over time following the pattern of blue light illumination. Opto-ALK3Ca shows the highest recruitment level in FAs, whereas that of opto-CAAX and opto-ALK3Ci is three times lower. The mean ± SD is shown for 10 cells. Scale bar, 15 µm.