Loss of Rab25 promotes the development of intestinal neoplasia in mice and is associated with human colorectal adenocarcinomas
Ki Taek Nam, … , Robert J. Coffey, James R. Goldenring
Ki Taek Nam, … , Robert J. Coffey, James R. Goldenring
Published February 8, 2010
Citation Information: J Clin Invest. 2010;120(3):840-849. https://doi.org/10.1172/JCI40728.
View: Text | PDF
Research Article Oncology

Loss of Rab25 promotes the development of intestinal neoplasia in mice and is associated with human colorectal adenocarcinomas

Abstract

Transformation of epithelial cells is associated with loss of cell polarity, which includes alterations in cell morphology as well as changes in the complement of plasma membrane proteins. Rab proteins regulate polarized trafficking to the cell membrane and therefore represent potential regulators of this neoplastic transition. Here we have demonstrated a tumor suppressor function for Rab25 in intestinal neoplasia in both mice and humans. Human colorectal adenocarcinomas exhibited reductions in Rab25 expression independent of stage, with lower Rab25 expression levels correlating with substantially shorter patient survival. In wild-type mice, Rab25 was strongly expressed in cells luminal to the proliferating cells of intestinal crypts. While Rab25-deficient mice did not exhibit gross pathology, ApcMin/+ mice crossed onto a Rab25-deficient background showed a 4-fold increase in intestinal polyps and a 2-fold increase in colonic tumors compared with parental ApcMin/+ mice. Rab25-deficient mice had decreased β1 integrin staining in the lateral membranes of villus cells, and this pattern was accentuated in Rab25-deficient mice crossed onto the ApcMin/+ background. Additionally, Smad3+/– mice crossed onto a Rab25-deficient background demonstrated a marked increase in colonic tumor formation. Taken together, these results suggest that Rab25 may function as a tumor suppressor in intestinal epithelial cells through regulation of protein trafficking to the cell surface.

Authors

Ki Taek Nam, Hyuk-Joon Lee, J. Joshua Smith, Lynne A. Lapierre, Vidya P. Kamath, Xi Chen, Bruce J. Aronow, Timothy J. Yeatman, Sheela G. Bhartur, Benjamin C. Calhoun, Brian Condie, Nancy R. Manley, R. Daniel Beauchamp, Robert J. Coffey, James R. Goldenring

×

Figure 3

Construction of Rab25-knockout mouse.

Options: View larger image (or click on image) Download as PowerPoint
Construction of Rab25-knockout mouse. (A) Schematic of the targeting con...
(A) Schematic of the targeting construct for disruption of the Rab25 gene with insertion of the neomycin sequence into exon II of the Rab25 gene. The NEO cassette contained termination codons in all 3 frames. (B) Southern blot of EcoRV digests of DNA from heterozygote (+/–), homozygote wild-type (+/+), and 2 Rab25-knockout mice (–/–) probed with sequence from exon V. (C) 3 oligonucleotide PCR-based screening assays showed discrimination of Rab25-knockout mice from heterozygotes and wild-type littermate mice. (D) Western blot of extracts of protein (50 mg) from the gastric mucosa of littermate mice probed with rabbit anti-mouse Rab25 showing reduction of Rab25 expression in heterozygotes and complete absence of detectable Rab25 in Rab25-knockout mice. The distribution of molecular mass standards is indicated at left (kDa). The lane images shown are from noncontiguous lanes of the same gel and Western blot. (E) Immunohistochemical localization of Rab25 in the intestinal mucosa of wild-type mice (+/+) using either immunohistochemistry (left 2 panels) or immunofluorescence (third panel; green is Rab25 and blue is DAPI). The cells in the transition zone between the crypt and the villus stained most strongly for Rab25. In Rab25-deficient mice (–/–, far right), no Rab25 staining was observed. Scale bars: 50 μm.