ReportsThe expression of p16INK4a, the product of a tumor suppressor gene for melanoma, is upregulated in human melanocytes by UVB irradiation☆,☆☆
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Melanocyte culture and UVB irradiation
Primary melanocyte cultures were established from human neonatal foreskins (from whites) with selective melanocyte growth medium (MGM; Clonetics, San Diego, Calif), as reported.9 At near confluence of the asynchronous monolayers, the cells were washed with warm HEPES-buffered saline, leaving but a thin film over the cells. Single UVB (290-320 nm) exposure was administered with a UltraLum UVB-28 lamp, calibrated with a UltraLum CDR-2 All Wave UV Intensity Meter (UltraLum, Inc, Paramount, Calif).
RESULTS
After irradiation at several UVB doses, or no irradiation, melanocyte cultures were harvested at increasing incubation times thereafter for analysis of p16INK4a protein expression by immunostaining. Cellular labeling intensities of the immunoperoxidase-stained cells fixed in situ on slide chambers were quantified by digital image analysis, as previously described.9 It was found that expression of p16INK4a protein increased over time after irradiation and was maximal at approximately 5 to 12
DISCUSSION
With these studies we have evaluated a rule for UVB irradiation in regulating the expression of p16INK4a in cultured human neonatal melanocytes. Trends in p16INK4a protein expression after dosing with UVB were analyzed by quantitative immunolabeling and by Western blotting, and complementary Northern blotting was used to assess coordinate changes in the levels of p16INK4a mRNA. The data demonstrated significant upregulation of p16INK4a protein and a greater than 2-fold increase in melanocyte p16
Acknowledgements
Note added in proof: A recent report showed that UV irradiation of skin organ cultures upregulates p16INK4a expression in epidermal melanocytes and keratinocytes. These results are consistent with those reported herein (Pavey S, Conroy S, Russell T, Gabrielli B. Ultraviolet radiation induces p16CDKN2A expression in human skin. Cancer Res 1999;59:4185-9).
The technical assistance of Linda Foy, Terrance Peterson, and Dr Deniz Seçkin is gratefully acknowledged.
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2017, Oral OncologyCitation Excerpt :Telomere attrition in the malignant tumor results in induction of CDKN2A/p16 and stabilization of telomeres by telomerase activation thus suppresses the expression of CDKN2A/p16 [19–23]. CDKN2A/p16 is also reported to respond to other telomere independent stress signal such as constitutive oncogenic signalling and DNA damage signalling [24–26]. Telomerase activity is reported to be upregulated in solid tumors including OSCC [4,27].
Suppression of autophagy dysregulates the antioxidant response and causes premature senescence of melanocytes
2015, Journal of Investigative DermatologyCitation Excerpt :The changes in the morphology of autophagy-deficient MC were accompanied by significantly higher expression of p16Ink4a (CDKN2A) and p21 (CDKN1A) mRNAs (Figure 4e, P<0.05), and a significantly higher proportion of mutant cells exhibited nuclear p16Ink4a protein as determined by immunofluorescence staining (Figure 4f and g). Nuclear translocation of p16Ink4a increased upon UVB irradiation (20 mJ cm-2) (Piepkorn, 2000) in autophagy-competent cells, whereas it could not be elevated above the high basal level in autophagy-deficient cells (Figure 4f and g). Taken together these data suggest that the absence of autophagy leads to premature senescence of MC.
Molecular pathology of malignant melanoma: Changing the clinical practice paradigm toward a personalized approach
2014, Human PathologyCitation Excerpt :INK4 encodes the protein p16, which is also located in the 9p21 region. This gene is upregulated after ultraviolet B exposure in human melanocytes [105]. Progressive loss of p16 is seen in transformation of nevi toward melanoma.
Lentigines, nevi, and melanomas
2009, Weedon's Skin Pathology: Third EditionPhotochemoprevention of ultraviolet B signaling and photocarcinogenesis
2005, Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis
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Supported in part by US Public Health Service grant AR 21557 from the National Institutes of Health, Department of Health and Human Services.
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Reprint requests: Michael Piepkorn, MD, PhD, Division of Dermatology, Box 356524, Seattle, WA 98195-6524.