ABSTRACT
BacKground:Fanconi anemia( FA)pathway as a DNA crossIink damage repair pathway pIays an important roIe in maintaining genome stabiIity. In recent years,FA pathway was wideIy studied in DNA damage and cancer pathogenesis. Aims:To investigate the interaction between RAD18 and FANCD2 protein in human coIorectaI cancer ceII Iine SW480. Methods:Antigen-antibody compIex was co-precipitated by immunoprecipitation. Expressions of rabbit anti-human FANCD2 and RAD18 protein in antigen-antibody compIexes were detected by Western bIotting. The pIasmids of GST-RAD18 and GST were transferred into BL21 ceIIs and induced to express the target proteins. TotaI proteins of the ceII was extracted and GST-beads were used to conjugate the GST-RAD18 protein,and then incubated with the SW480 ceII Iysates, and Western bIotting was performed with the addition of rabbit anti-human FANCD2 antibodies. Results:RAD18 protein was detected in the antigen-antibody compIex from immunoprecipitation by using anti-FANCD2 antibody,and FANCD2 protein was detected by using anti-RAD18 antibody. FANCD2 protein was aIso detected by using anti-GST-RAD18 antibody. GST-RAD18 protein used as bait protein couId capture the FANCD2 protein in SW480 ceIIs. Conclusions:There is an interaction between RAD18 and FANCD2 protein in SW480 ceIIs,and aIso an interaction between GST-RAD18 and FANCD2 protein.
ABSTRACT
Background:Fanconi anemia( FA ),an autosomal or x-linked recessive inherited disease,is caused by gene mutation related to FA pathway of DNA damage and with the clinical features of congenital malformation,bone marrow failure and susceptibility to cancer. Aims:To investigate the expression of Fanconi anemia complementation group D2 (FANCD2)in colorectal cancer and its correlation with prognosis. Methods:Fifty-six surgical resected specimens of colorectal cancer tissue and para-cancer noncancerous tissue from May 2012 to September 2013 at Guangzhou First People’s Hospital were obtained. Ninety-three patients with colorectal cancer from January 2008 to April 2009 at Guangzhou First People ’s Hospital were enrolled and the specimens of colorectal cancer tissue were obtained. The expression of FANCD2 mRNA in 56 specimens of colorectal cancer tissue and para-cancer noncancerous tissue was determined by qPCR. The expression of FANCD2 protein in 49 of 56 specimens of colorectal cancer tissue and para-cancer noncancerous tissue and 93 specimens of colorectal cancer tissue was determined by immunohistochemistry. Follow up was conducted in 93 patients and the correlation between the expression of FANCD2 and prognosis was analyzed. Results:Expression of FANCD2 mRNA was significantly higher in colorectal cancer tissue than that in para-cancer noncancerous tissue [0.102(0.047,0. 163)vs. 0. 051(0. 025,0. 095)](P =0. 007). Expression of FANCD2 mRNA was significantly correlated with lymph node metastasis and Dukes stage( P <0. 05 ). Positivity rates of FANCD2 protein expression in colorectal cancer tissue and para-cancer noncancerous tissue were 77. 6%(38/49)and 22. 4%(11/49),respectively. Expression of FANCD2 protein was significantly correlated with lymph node metastasis and Dukes stage(P<0. 05). The overall 5-year survival rate was significantly lower in patients with positive expression of FANCD2 protein than that in patients with negative expression of FANCD2 protein(35. 5% vs. 71. 0%)(P<0. 01). Conclusions:FANCD2 is associated with malignant potential of colorectal cancer and could be a potential marker for prognosis.