Fig. 4

Extracellular AFAP1-AS1 is transferred through incorporating into exosomes. a RNA fluorescence in situ hybridization (RNA-FISH) verified that AFAP1-AS1 was mainly distributed in nuclear of SKBR-3-TR and BT474-TR cells. b Extracellular AFAP1-AS1 was degraded by treatment with RNAse A and Triton simultaneously, ^**P < 0.01. c TEM scanning showed the exosomes images released by SKBR-3-TR and SKBR-3 cells. d Size distribution of exosomes were analyzed by Zetasizer. e Exosomal protein markers (TSG101 and CD81) detection by Western blot from purified exosomes and exosome-depleted cell extracts. f qRT-PCR analysis of lncRNA AFAP1-AS1 expression in exosomes, culture medium with or without exosomes, ^***P < 0.001. g RT-qPCR analysis showed that exosomal AFAP1-AS1 was upregulated in trastuzumab-resistant cells in contrast to sensitive cells, ^**P < 0.01. h RNA immunoprecipitation assay using anti-HNRNPA2B1 antibody showed that AFAP1-AS1 was associated with HNRNPA2B1 protein. i HNRNPA2B1 was silenced by specific oligonucleotides in both transcript and protein levels, ^**P < 0.01. j Knockdown of HNRNPA2B1 decreased AFAP1-AS1 expression in SKBR-3-TR and BT474-TR cells, ^*P < 0.05. k qRT-PCR and western blot verified the upregulation of HNRNPA2B1 by transfection of specific plasmids, ^**P < 0.01. l AFAP1-AS1 was upregulated by overexpression of HNRNPA2B1 in SKBR-3-TR and BT474-TR cells, ^*P < 0.05