SUNDAY, DECEMBER 4- Late Abstracts 1 - Molecular Biology of the ...

SUNDAY
suppressed Akt and Smad1/5 phosphorylation, calcium deposition and Col I α2 mRNA
expression in hADSCs. Most importantly, blockage of Akt activation significantly inhibited the
synergistic enhancement of Col I α2 mRNA expression, which was induced by BMP-6 and
VEGF combined treatment.
Conclusion and Discussion:
Our data showed a striking observation that VEGF and BMP-6 synergistically enhanced Col I α2
gene expression in hADSCs. Furthermore, biochemical blockage of Akt activation significantly
inhibited the enhancing effects of Col I α2 mRNA expression by BMP-6 and VEGF combined
treatment. It is reported that PI3-kinase catalytic isoform is coupled to VEGFR2 and regulates
the bulk of Akt activity. Moreover, dominant-negative Akt inhibited Smad5 phosphorylation. Our
data and these reported observations together establish a novel axis of control of Col I
expression as VEGF-VEGFR2-PI3K-Akt-Smad1/5-Col I in hADSCs. However, Akt-dependent
BMP-6 modulation of this axis remains to be investigated in near future.
2032
A novel gene encoding a candidate CK2 substrate involves in cancer cell proliferation.
H. Yang 1 , S. Liu 1 , H. Zheng 1 , H. Jiang 1 , Y. Qu 1 , Y. Li 1 ; 1 Harbin Institute of Technology, Harbin,
China
CK2, a ubiquitous protein serine/threonine kinase, is essential for the decision of cell survival
and cell death. In the present study, we identified a novel gene, Homo Sapiens Chromosome 1
ORF109 which protein product might be a candidate CK2 substrate. The C1ORF109 protein
presented in the phosphorylated form in vivo, and could be phosphorylized by the protein kinase
CK2 in vitro. Truncated constructs and site direct mutagenesis analysis revealed that Ser104,
Ser 134, and Ser182 were phosphorylated sites. Both TATA box and CAAT box within the
crucial region of c1orf109 promoter are required for maximal transcription of the c1orf109 gene.
The C1ORF109 protein was mainly located in the nucleus and cytoplasm. Upregulated
expression of c1orf109 was detected in multiple breast cancer cells. Exogenous expression of
C1ORF109 in breast cancer Hs578T cells promoted colony formation and cell proliferation
along with increased level of PCNA and cyclinD1. Meanwhile, knockdown of c1orf109 by siRNA
in breast cancer MDA-MB-231 cells resulted in inhibition of cell proliferation. Taken together, our
findings suggest that C1ORF109 may be the down stream target of protein kinase CK2 and
involved in the regulation of cancer cell proliferation.
Key words: promoter, transcription, CK2 kinase, c1orf109, proliferation
2033
Quantitative detection of phosphorylation of STAT3 and p38 by Western blotting.
K. Söderquist 1 , S. Grimsby 1 , M. Winkvist 1 ; 1 GE Healthcare, Uppsala, Sweden
Introduction
Western blotting is a well established technique used to study proteins from a wide variety of
sources. The technique is used throughout life sciences from basic research to medical
diagnostic applications. Western blotting is at best considered as semi-quantitative and hence
limited to studies involving large protein differences. However development of detection
reagents as well as imagers has open up for quantitative applications when high sensitivity and
broad dynamic range is needed.
Here we demonstrate how the Western blotting technique can be used to determine
phosphorylation levels of various proteins in whole cell lysates upon stimulation with various
well known inducers.