Figure 6

CSE1L is a microvesicle membrane protein and anti-CSE1L antibodies can target tumors. (A) Representative immunofluorescence show preferential accumulation of CSE1L (arrowheads) in microvesicles. MMP-2 distribution was analyzed for comparison. (B) Localization of CSE1L in microvesicle membranes (arrowhead) analyzed by immunofluorescence. (C) A representative immunogold electron microscopic image shows CSE1L (12-nm gold, arrowheads) distributions in microvesicle in B16-Ras cells. (D) Representative immunofluorescence shows CSE1L (arrowheads) staining in microvesicles harvested from the sera of colorectal cancer patients. Control immunofluorescence was performed with the samples stained with secondary antibodies coupled with Alexa 568 (lower panel). (E) Representative images show near-infrared fluorescence signals of mice bearing B16-CSE1L cell-derived tumors injected with Qdot-conjugated anti-CSE1L antibodies (right, n = 3) or Qdot-conjugated anti-mouse IgG (left, n = 3) at 4 h after injection. (F) Pathway of Ras-ERK-CSE1L signaling in the regulation of microvesicle biogenesis. Ras activated by the upstream signaling such as the ErbB2 results in ERK activation. ERK interacts with CSE1L, which in turn mediates microvesicle biogenesis. CSE1L-induced microvesicle biogenesis is not inhibited by PD98059, indicating that there is a signaling pathway other than ERK that also interacts with CSE1L and regulates microvesicle generation. ERK activation also results in MLCK activation, which in turn modulates microvesicle shedding.