Fig. 4: Circp53–209aa-R175 is identified as the site-specific target of CypD to trigger mPTP opening in MM cells. | Experimental & Molecular Medicine

Fig. 4: Circp53–209aa-R175 is identified as the site-specific target of CypD to trigger mPTP opening in MM cells.

From: Extracellular vesicle-mediated delivery of circp53 suppresses the progression of multiple cancers by activating the CypD/TRAP/HSP90 pathway

Fig. 4: Circp53–209aa-R175 is identified as the site-specific target of CypD to trigger mPTP opening in MM cells.

a, GO analysis of RNA-sequencing data revealed that the circp53–209aa is highly associated with mitochondrial pathways. b, Go analysis of RNA-sequencing data revealed that the circp53–209aa is highly associated with apoptotic process signaling pathways. c, A graphic illustration of circp53–209aa interacting with CypD and releasing CypD from the CypD/TRAP1/HSP90 complex, thereby activating its isomerase activity and inducing mPTP opening. d, A Co-IP assay confirmed the direct interaction between circp53–209aa and CypD in MM cells. e,Representative confocal images of HA and CypD demonstrate the interaction between circp53–209aa and CypD. f, Statistical analysis of the colocation of circp53-209aa and CypD in H929 and XG1 cells. g, A Co-IP assay showed a weaker interaction between CypD and HSP90 in circp53-OE MM cells compared with Ctrl cells. h, Distribution of calcein in H929 and MM1.S cells. i, Distribution of calcein in XG1 and RPMI 8226 cells. j, IOD quantitative statistics of Calcein in Ctrl and circp53–209aa-OE MM cells. k, ATP levels were significantly decreased in circp53–209aa-OE cells. l, Statistical analysis of ATP levels in different cells (P < 0.001). m, A model showing the high confidence interval of the CypD protein. n, A model of the circp53–209aa-CypD complex. R175 of circp53–209aa binds to E43, E34 and Q163 of CypD through hydrogen bonds and/or ionic interactions. o, Circp53–209aa-R175 is involved in hydrogen-bond interactions with CypD, and these interactions are disrupted upon mutation of arginine to a histidine, as demonstrated by Co-IP using an HA antibody as bait in circp53-OE cells, followed by WB analysis. p, Circp53–209aa-R175H-OE MM cells were constructed successfully, as shown by WB analysis. q, Circp53–209aa-R175H did not influence the proliferation of MM cells. r, Circp53–209aa interacts with CypD and releases it from the CypD/TRAP1/HSP90 complex, as evidenced by Co-IP. s, Colocalization dynamics of CypD/HSP90 in H929 and MM1.S cells. t, Colocalization dynamics of CypD/HSP90 in XG1 and 8226 cells. u, Statistical analysis of the colocation of CypD and HSP90 in H929 and MM1.S cells. v, Statistical analysis of the colocation of CypD and HSP90 in XG1 and RPMI 8226 cells. w, IOD quantitative statistics of CypD and HSP90 in MM cells. x, Distribution of calcein in circp53-OE and circp53–209aa-R175 MM cells. y, The mPTP was not opened in the circp53–209aa-R175H group by quantitative statistics. The data are presented as mean ± s.d. (n = 3 cultures for each group, *P < 0.05, **P < 0.01, ***P < 0.001 and ns, no statistical significance).

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