Department Molecular Cell Biology Weizmann Institute of Science







Laboratory of David Givol

Microarray Comparative Analysis
of Normal (NHEK) and Tumoral (SCC) Human Keratinocytes
response to UVB Irradiation





Figures :


Dose and time dependent effects of a single UVB irradiation on normal keratinocytes (NHEK)


(A) Effect of UVB irradiation on cell viability as determined by Trypan blue uptake assay.
(B) Cell culture morphologies.
(C) Cell cycle analysis as determined by FACS histogram plots of DNA content.
(D) Western blot analysis of MDM2, DNp63, and p53 protein expression





Effects of a single or multiple doses of UVB irradiation on NHEK and SCC12B2


(A) Cell culture morphologies before (0h) and after UVB irradiation (48h).
(B) Percent (%) of sub G1 DNA content 48h after a single, or multiple UVB irradiations.


(C) Western blot analysis of PARP and caspase cleavage. Both full length and cleaved forms (p89) of PARP are shown.
For caspases, only the low MW active forms of caspase-8 (p18), caspase-3 (p15/17 doublet), and caspase-9 (p37) are shown.
(D) Western blot analysis of protein expression of proliferation-related genes.





Effect on cell death of exchange of UVB-conditioned medium between NHEK and SCC12B2 cells


Western blot analysis of PARP cleavage, percent (%) of sub G1 DNA content, and cell morphology 48h following a single UVB irradiation.
NHEK maintained in conditioned medium from NHEK (lane 1) or SCC12B2 (lane 2),
SCC12B2 cells maintained in conditioned medium from NHEK (lane 3) or SCC12B2 (lane 4)





Change of gene expression in NHEK and SCC12B2 cells following a single dose of UVB irradiation


(A) Venn diagram showing the number of up and down regulated genes in NHEK and SCC using a filter of twofold up- and threefold down-regulation in at least two time points.


(B) Scatter plot of changes of expression of monitored genes in NHEK and SCC cells.
Left panel: changes of gene expression in NHEK between sham irradiated controls.
Right panel: changes of gene expression in NHEK between control 0h (sham irradiated) and 24h following UVB irradiation.





Common Top Regulated Genes Both In
- UVB-Irradiated Normal Keratinocytes (NHEK)
- Squamous Cell Carcinoma (SCC)
- Colorectal Carcinoma (CRC)








Clustering of the UVB-regulated genes in NHEK using the SPC algorithm


(A) Dendrogram showing clusters of genes with four or more members. Six stable clusters of genes (G1-G6) could be identified.
Each cluster is represented by a box colored according to the proportion (left colorbar) of genes belonging to
the functional category 'cytokines and growth factors'.
The distribution of these cytokines and growth factors genes (cluster G2 and red crosses),
and that of chemokine genes of the CXC/CC subfamilies (blue crosses) is shown at the right.
(B) Expression matrix of the up-(red) and down-(blue) UVB regulated genes (right colorbar).
Centered and normalized log2 signals of the five time points post irradiation are shown.
(C) Average time course expression profiles of the genes in clusters G1-G6.





Coupled Two Way Clustering (CTWC) results showing partitions of genes and samples between NHEK and SCC


Each stable cluster identified by the CTWC method contains genes and samples reordered in an unsupervised coupled two way manner.
(A) Genes downregulated in NHEK, but showing high expression and no response to UVB in SCC.
(B) Genes that are UVB-responsive in normal keratinocyte cells only.


(C) Genes that are constitutively expressed at low levels in both SCC and NHEK before UVB, but are differentially regulated after UVB exposure.
P-values associated to each gene were determined in a permutation test by comparing the expression levels
in NHEK vs.SCC following UVB irradiation. Genes with a low p-value are those differentially regulated in NHEK vs. SCC after UVB exposure.


(D) Genes with low expression and no response to UVB in SCC, but high expression and moderate response to UVB in NHEK.





Tables :


Table 1 :  Selected Regulated Genes in NHEK and SCC12B2 in Response to UVB Irradiation





Supplementary Data :


Table 2 :  Full List of Regulated Genes in NHEK in Response to UVB Irradiation

Table 3 :  Full List of Regulated Genes in SCC12B2 in Response to UVB Irradiation





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Weizmann Institute of Science
Department Molecular Cell Biology
76100 Rehovot, Israel
Phone: +972-8-934-3634
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