Fig. 4. LAP1 enables repeated constrained migration and invasion.
a, Schematic of TOR1AIP1 human gene transcription and translation. The gene comprises ten exons and encodes a long protein isoform (584 amino acids), LAP1B, and a short protein isoform (462 amino acids), LAP1C, resulting from the use of an alternative translation initiation site at position 122. b, Representative immunoblot of LAP1 expression levels in the indicated melanocytes, primary melanoma cells and metastatic melanoma cells. c, Quantification of LAP1B and LAP1C protein expression in panel of cell lines in b, normalized to GAPDH. d, Representative immunoblot of LAP1 expression levels in WM983B cells upon transfection with pooled LAP1-targeting siRNA. e,f, Percentage of WM983B cells that translocated their nuclei (e) and displayed NE blebs (f) after a second round of transwell migration through 5 μm pores upon LAP1 depletion with siRNA pool. n = 872 and 705 cells, respectively. g,h, 3D invasion index (number of invading cells at 50 μm/total number of cells) of WM983B cells in 3D collagen I matrices upon LAP1 depletion with siRNA pool. Representative images of WM983B cells stained for DNA at 0 μm and 50 μm into collagen upon LAP1 depletion with siRNA pool. Scale bars, 30 μm. n = 673 and 539 cells, respectively. Bar charts show the mean and error bars represent s.e.m. from N = 3 independent experiments. P values calculated by two-tailed unpaired t-test (e–g); **P < 0.01, ***P < 0.001. Unprocessed western blots, numerical data and exact P values are available in source data.