Werner syndrome is an autosomal recessive disorder associated with premature aging and cancer predisposition caused by mutations of the WRN gene. WRN is a member of the RecQ DNA helicase family with functions in maintaining genome stability. Sir2, an NAD-dependent histone deacetylase, has been proven to extend life span in yeast and Caenorhabditis elegans. Mammalian Sir2 (SIRT1) has also been found to regulate premature cellular senescence induced by the tumor suppressors PML and p53. SIRT1 plays an important role in cell survival promoted by calorie restriction. Here we show that SIRT1 interacts with WRN both in vitro and in vivo; this interaction is enhanced after DNA damage. WRN can be acetylated by acetyltransferase CBP/p300, and SIRT1 can deacetylate WRN both in vitro and in vivo. WRN acetylation decreases its helicase and exonuclease activities, and SIRT1 can reverse this effect. WRN acetylation alters its nuclear distribution. Down-regulation of SIRT1 reduces WRN translocation from nucleoplasm to nucleoli after DNA damage. These results suggest that SIRT1 regulates WRN-mediated cellular responses to DNA damage through deacetylation of WRN. Werner syndrome (WS)3 is a human autosomal recessive disorder that displays symptoms of premature aging, including graying and loss of hair, wrinkling and ulceration of skin, atherosclerosis, osteoporosis, and cataracts. WS patients also exhibit an increased incidence of diabetes mellitus type 2, hypertension, and are highly disposed to cancers (1). WS results from mutation of the WRN gene, a member of the RecQ DNA helicase family (2). Mutations in other family members, BLM and RECQ4, are responsible for the two other cancer-prone and premature aging syndromes, Bloom (3) and Rothmund-Thomson (4), respectively. Consistent with other RecQ helicases, WRN protein possesses 3Ј to 5Ј DNA helicase activity; however, it is the only human RecQ member to also have a 3Ј to 5Ј exonuclease activity. Although its physiological substrate is not yet clear, WRN appears to preferentially act on replication and recombination structures. Cells from WS patients show premature replicative senescence compared with cells derived from normal individuals (5). WS cells also show hypersensitivity to selected DNA-damaging agents including 4-nitroquinoline-1-oxide (4NQO; 6), topoisomerase inhibitors (7), and certain DNA cross-linking agents (8). Compared with normal cells, WS cells also exhibit increased genomic instability including higher levels of DNA deletions, translocations, and chromosomal breaks (9, 10), suggesting that WRN plays an important role in one or more genome maintenance pathways (11).WRN protein shows dynamic relocalization within the nucleus under different conditions of growth. The WRN protein localizes to the nucleoli in a variety of cell types (12), and this localization is modulated by DNA damage and cell cycle. Upon serum starvation or treatment with hydroxyurea (HU), aphidicolin, 4NQO, etoposide, or camptothecin, WRN migrates from nucleoli to discrete nuclear foci (13-17). ...

In order to model squamous cell carcinoma development in vivo , researchers have long preferred hairless mouse models such as SKH-1 mice that have traditionally been classified as ‘wild-type’ mice irrespective of the genetic factors underlying their hairless phenotype. The work presented here shows that mutations in the Hairless ( Hr ) gene not only result in the hairless phenotype of the SKH-1 and Hr −/− mouse lines but also cause aberrant activation of NFκB and its downstream effectors. We show that in the epidermis, Hr is an early UVB response gene that regulates NFκB activation and thereby controls cellular responses to irradiation. Therefore, when Hr expression is decreased in Hr mutant animals there is a corresponding increase in NFκB activity that is augmented by UVB irradiation. This constitutive activation of NFκB in the Hr mutant epidermis leads to the stimulation a large variety of downstream effectors including the cell cycle regulators cyclin D1 and cyclin E, the anti-apoptosis protein Bcl-2, and the pro-inflammatory protein Cox-2. Therefore, Hr loss results in a state of uncontrolled epidermal proliferation that promotes tumor development, and Hr mutant mice should no longer be considered merely hairless 'wild-type' mice. Instead, Hr is a crucial UVB response gene and its loss creates a permissive environment that potentiates increased tumorigenesis.

Hairless (HR) is a nuclear protein with co-repressor activity that is highly expressed in the skin and hair follicle. Mutations in Hairless lead to hair loss accompanied by the appearance of papules (atrichia with papular lesions) and similar phenotypes appear when the key polyamine enzymes ornithine decarboxylase (ODC) and spermidine/spermine N1-acetyltransferase (SSAT) are overexpressed. Both ODC and SSAT transgenic mice have elevated epidermal levels of putrescine, leading us to investigate the mechanistic link between putrescine and HR. We show here that HR and putrescine form a negative regulatory network, since epidermal ODC expression is elevated when HR is decreased and vice versa. We also show that regulation of ODC by HR is dependent on the MYC superfamily of proteins, in particular MYC, MXI1 and MXD3. Furthermore, we found that elevated levels of putrescine lead to decreased HR expression but that the SSAT-TG phenotype is distinct from that of HR mutants. Transcriptional microarray analysis of putrescine-treated primary human keratinocytes demonstrated differential regulation of genes involved in protein-protein interactions, nucleotide binding, and transcription factor activity, suggesting that the putrescine-HR negative regulatory loop may have a large impact on epidermal homeostasis and hair follicle cycling.

Hairless (HR) is a nuclear protein with corepressor activity whose exact function in the skin remains to be determined. Mutations in both human and mouse Hairless lead to hair loss accompanied by the appearance of papules, a disorder called atrichia with papular lesions. Furthermore, mice with mutations in HR are known to have a higher susceptibility to ultraviolet radiation-induced tumorigenesis, suggesting that HR plays a crucial role in the epidermal UVB response. Using normal human keratinocytes (NHKs) and keratinocytes containing a mutation in HR, we found that HR is an early UVB response gene that negatively regulates NFκB mRNA expression. HR mutant keratinocytes have a dysregulated UVB response that includes increased proliferation and the aberrant activation of NFκB effector genes. Additionally, we show that another UVB response gene, TNFα, negatively regulates HR mRNA expression. TNFα-induced negative regulation of HR occurs through a direct interaction of the p65 subunit with a single NFκB-binding domain located in the HR promoter region. Therefore, we show for the first time that HR and NFκB participate in a positive feedback loop that can be initiated either by UVB or TNFα.