Fig. 5: ECM-related factors contained in GBM migrasomes.

a Volcano plot of the quantification of TMT-labelled proteins through mass spectrometry. Red dots indicate a migrasome with a cell abundance ratio of >2 (P < 0.05), while blue dots indicate a migrasome with a cell abundance ratio of <0.5 (P < 0.05). The experiment was performed independently three times (n = 3). PAK4 and LAMA4 are labeled separately. A two-tailed, two-sample, unequal variance t-test was used in Excel to calculate the P values. b, d The abundance of the indicated proteins in 5a was analyzed. P values were calculated using a one-tailed, two-sample unequal variance t-test. The P values were as follows: histone (P = 0.000215), integrin (P = 0.000379), tetraspanin (P = 0.000556), cytokines (P = 0.001711), PAK4 (P = 0.001885), LAMA4 (P = 0.008333), and ELP6 (P = 0.000181). c For the gene ontology (GO) enrichment analysis at the biological process (BP) level of the red-dot proteins in 5a, the pathways associated with migration are highlighted in red. e Western blot was used to analyze cell body and migrasome proteins with anti-LAMA4 and anti-PAK4 antibodies. Two batches of cell body and migrasome proteins were extracted. CB: cell body. Mig: migrasome. f LN229 cells were visualized using confocal microscopy after staining with WGA, anti-LAMA4, and anti-PAK4 antibodies. Scale bar, 10 μm. High magnification images of the migrasomes are shown in the white boxes. Scale bar, 2 μm. For the co-localization analysis of the boxed region, the arrows indicate co-localization. g Staining of cells migrating/invading to the bottom of Transwell chambers using crystal violet. Cells were treated with PBS or recombinant proteins PAK4 and LAMA4. ***P < 0.001. ****P < 0.0001.