Expression of ral GTPases, their effectors, and activators in human bladder cancer.
OBJECTIVE
The Ral family of small G proteins has been implicated in tumorigenesis, invasion, and metastasis in in vitro and animal model systems; however, a systematic evaluation of the state of activation, mutation, or expression of these GTPases has not been reported in any tumor type.


METHODS
We determined the activation state of the RalA and RalB paralogs in 10 bladder cancer cell lines with varying Ras mutation status. We sequenced RalA and RalB cDNAs from 20 bladder cancer cell lines and functionally evaluated the mutations found. We determined the expression of Ral, Ral activators, and Ral effectors on the level of mRNA or protein in human bladder cancer cell lines and tissues.


RESULTS
We uncovered one E97Q substitution mutation of RalA in 1 of 20 cell lines tested and higher Ral activation in cells harboring mutant HRAS. We found overexpression of mRNAs for RalA and Aurora-A, a mitotic kinase that activates RalA, in bladder cancer (both P < 0.001), and in association with tumors of higher stage and grade. RalBP1, a canonical Ral effector, mRNA and protein was overexpressed in bladder cancer (P < 0.001), whereas Filamin A was underexpressed (P = 0.004). We determined that RalA mRNA levels correlated significantly with protein levels (P < 0.001) and found protein overexpression of both GTPases in homogenized invasive cancers. Available data sets suggest that RalA mRNA is also overexpressed in seminoma, glioblastoma, and carcinomas of the liver, pancreas, and prostate.


CONCLUSIONS
These findings of activation and differential expression of RalA and RalB anchor prior work in model systems to human disease and suggest therapeutic strategies targeting both GTPases in this pathway may be beneficial.
Smith
Steven Christopher
SC
Department of Molecular Physiology, University of Virginia, Charlottesville, Virginia 22908, USA.
Oxford
Gary
G
Baras
Alexander S
AS
Owens
Charles
C
Havaleshko
Dmytro
D
Brautigan
David L
DL
Safo
Martin K
MK
Theodorescu
Dan
D
D000483:Alleles; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008954:Models, Biological; D008957:Models, Genetic; D008958:Models, Molecular; D009154:Mutation; D011487:Protein Conformation; D014018:Tissue Distribution; D001749:Urinary Bladder Neoplasms / Q000201:enzymology* / Q000473:pathology; D020725:ral GTP-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics
C483163:Ralb protein, human; C492198:RALA protein, human; D020725:ral GTP-Binding Proteins