Epigenetic regulation of RhoB loss of expression in lung cancer.
BACKGROUND
RhoB is down-regulated in most lung cancer cell lines and tumor tissues when compared with their normal counterparts. The mechanism of this loss of expression is not yet deciphered.


METHODS
Since no mutation has been reported in the RhoB sequence, we investigated the epigenetic regulation of RhoB expression by analyzing the effect of HDAC inhibitors and methyltransferase inhibitors, by direct sequencing after bisulfite treatment and by methylation specific PCR.


RESULTS
We first showed that histone deacetylase (HDAC) inhibitors induce a significant RhoB re-expression in lung cancer cell lines whereas only a slight effect was observed with methyl transferase inhibitors. As promoter methylation is the most common epigenetic process in lung cancer, we performed methylation specific PCR and sequence analysis after bisulfite treatment and demonstrated that RhoB was methylated neither in lung cancer cell lines nor in tumor tissues. We also showed that a variable number of tandem repeats sequences in the 5' region of the RhoB gene was involved in HDAC response.


CONCLUSIONS
We thus propose that RhoB regulation of expression occurs mainly by histone deacetylation rather than by promoter hypermethylation and that this process can be modulated by specific 5' sequences within the promoter.
INSERM U563 - Département Innovation Thérapeutique et Oncologie Moléculaire, F-31052, Toulouse, France. mazieres.j@chu-toulouse.fr
Julien
J
Mazières
Daniel
D
Tovar
Biao
B
He
Jacobo
J
Nieto-Acosta
Claire
C
Marty-Detraves
Carine
C
Clanet
Anne
A
Pradines
David
D
Jablons
Gilles
G
Favre
D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D001483:Base Sequence; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors; D019175:DNA Methylation; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D044127:Epigenesis, Genetic; D015870:Gene Expression; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D015151:Immunoblotting; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013447:Sulfites; D020080:Tandem Repeat Sequences; D020743:rhoB GTP-Binding Protein / Q000096:biosynthesis* / Q000235:genetics
D004791:Enzyme Inhibitors; D056572:Histone Deacetylase Inhibitors; D013447:Sulfites; D004248:DNA (Cytosine-5-)-Methyltransferases; D020743:rhoB GTP-Binding Protein; D001374:Azacitidine; C042345:hydrogen sulfite