RESUMO
The low-fidelity DNA polymerases thought to be specialized in DNA damage processing are frequently misregulated in cancers. We show here that DNA polymerase kappa (polkappa), prone to replicate across oxidative and aromatic adducts and known to function in nucleotide excision repair (NER), is downregulated in colorectal tumour biopsies. Contrary to the replicative poldelta and polalpha, for which only activating domains were described, we identified an upstream 465-bp-long repressor region in the promoter of POLK. We also found an activating 237-bp region that includes stimulating protein-1 (SP1) and cyclic AMP-responsive element (CRE)-binding sites. Mutations at one CRE-binding site led to a dramatic 80% decrease in promoter activity. Alterations of the SP1-binding site also affected, to a lesser extent, the transcription. Gel shift assays confirmed the role played by CRE/SP1 recognition sequences. Moreover, ectopic expression of SP1 or CRE-binding protein (CREB) protein favoured polkappa transcription. Finally, we found that polkappa downexpression in colorectal biopsies correlated with a decreased level of CREB and SP1 transcripts. This work shows that the promoter of POLK is cis-controlled and suggests that silencing of CREB and SP1 proteins could contribute to downregulation of this repair polymerase in colorectal tumours.
Assuntos
Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , DNA Polimerase Dirigida por DNA/metabolismo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Regiões Promotoras Genéticas/genética , Acetilação , Biópsia , Neoplasias Colorretais/patologia , DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , DNA Polimerase Dirigida por DNA/genética , Humanos , Pessoa de Meia-Idade , Mutação/genética , RNA Mensageiro/genética , Sequências Reguladoras de Ácido Nucleico , Transcrição Gênica/genéticaRESUMO
A major tolerance mechanism that functions to replicate damaged genomic DNA across lesions that have escaped elimination by repair mechanism is translesion DNA synthesis (TLS). DNA polymerase kappa (Pol kappa), a specialised low-fidelity DNA polymerase which is able to perform DNA synthesis across several damaged bases, is one of the enzymes involved in the process. The mutagenic nature of Pol kappa implies that its expression must be tightly regulated to prevent the formation of excessive genetic disorders along undamaged parts of the genome. Indeed, Pol kappa overexpression, which is notably observed in lung cancer, results not only in increased spontaneous mutagenesis, but also in pleiotropic alterations such as DNA breaks, genetic exchanges and aneuploidy. This review will discuss both aspects of DNA polymerase kappa, which can be considered as a genomic supervisor participating in genome maintenance and when misregulated as a genetic instability enhancer as well.
Assuntos
Dano ao DNA , DNA Polimerase Dirigida por DNA/fisiologia , Instabilidade Genômica , Animais , Cricetinae , Reparo do DNA , Replicação do DNA , DNA Polimerase Dirigida por DNA/genética , Regulação da Expressão Gênica , Humanos , Modelos Genéticos , MutagêneseRESUMO
A solution to the long-standing problem presented by the oxidative cyclization of a phenolic 3-arylpropionamide to a spirolactam has been developed in this laboratory via oxazoline chemistry. This research was motivated by our interest in some novel tricyclic azaspirane natural products formally derived from tyrosine, such as FR901483 and TAN1251C. In this paper, we disclose full details of the total synthesis of these substances.