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. 2016 May 16;36(11):1639–1654. doi: 10.1128/MCB.00020-16

FIG 8.

FIG 8

The turnover rate of Neph1 at the podocyte cell membrane is decreased upon loss of Myo1c binding. (A) The turnover rate of Neph1 at the podocyte cell membrane was analyzed in cultured podocytes stably transfected with mCherry-Neph1-wt and mCherry-Neph1-K761A by fluorescence recovery after photobleaching (FRAP). A region of interest (ROI) at the cell junction was selected and photobleached. The recovery of Neph1 at the photobleached region was recorded over a period of 5 min. Arrows indicate the localization of mCherry-Neph1-wt at the cell-cell junctions. (B) The FRAP analysis suggests a high turnover rate for Neph1-wt with >50% fluorescence recovery (n = 3 experiments) at the membrane within 3 min of recovery time, whereas the Myo1c-binding Neph1-K761A mutant displayed <10% (n = 3 experiments) recovery under similar conditions. (C) The observed mobile fraction for mCherry-Neph1-wt was about 80%, compared to 20% for the Neph1-K761A mutant. Data are presented as means ± SEM.