Furthermore, and mice develop a broader spectrum of tumors than null mice and ARF overexpression can induce cell cycle arrest in cells lacking p53. the molecular mechanisms of a novel part played by ARF will become offered and discussed, both in pathological and physiological contexts. locus, chemoresistance, FAK sumoylation, actin cytoskeleton 1. Intro The ARF (alternate reading framework) protein is definitely encoded by the Alternative Reading Frame of the locus, probably one of the most regularly mutated sites in human being cancers after the p53 locus [1,2,3]. The locus, located on human chromosome 9p21, encodes two completely unrelated proteins, p16INK4a and p14ARF, both of which are potent tumor suppressors. The mechanism by which the two proteins are produced is quite unusual. Each gene is usually endowed with its own promoter that guides the transcription of an – or -transcript. Each transcript has a specific 5 exon, E1 or E1 for INK4a and ARF respectively, spliced to a common exon 2 (Physique 1a) in which two overlapped ORFs (Open Reading Frame) are translated into two proteins sharing no amino acid sequence identity at all. Open in a separate window Physique 1 Genomic structure of the CDKN2a locus and produced transcripts. (a) Arrows above each exon 1 indicate promoters, continuous and dashed lines above and below the genomic structure indicate and splicing patterns respectively. Transcription of exon 1, and its splicing to exons 2 and 3 results in the -transcript, encoding p16INK4a, whereas transcription starting upstream of exon 1 produces the -transcript in which the exon1, and the common exons 2 and 3 encode ARF (p14ARF in human, p19Arf in mouse). In yellow and in reddish are indicated the open reading frames (ORFs) of p16 and ARF respectively, with exon 2 displaying two overlapped ORFs. White boxes represent untranslated regions at the 3 and 5 ends while asterisks (*) indicate quit codons (b) Pathways regulated by the two ADL5747 proteins: while p14ARF inhibits Mdm2 (Mouse Double Minute-2) functions with consequential p53 stabilization [4,5], p16INK4a inhibits the cyclinD-CDK4/6 complex thus maintaining the retinoblastoma protein pRb in its growth-suppressive mode . The alpha transcript encodes the p16INK4a protein, a member of the INK4 family of inhibitors of the cyclin-dependent kinases 4 and 6 (Inhibitor of CDK4). In response to specific signals, they block the assembly and/or inhibit the kinase activity of the cyclin D-CDK4/6 complex required for G1 to S cell cycle progression [6,7]. In this way, the retinoblastoma protein pRB is usually maintained in an active hypo-phosphorylated state and sequesters the transcription factors of the E2F family causing G1-phase cell cycle arrest [7,8] (Physique 1b). The ARF protein instead inhibits the functions of the MDM2 oncoprotein (Mouse Double Minute 2, HDM2 in human) thus inducing p53 stabilization and the activation of p53-dependent pathways (Physique 1b). In humans, the transcript results in a polypeptide of 132 amino acids (14 kDa) named p14ARF while, in mice, the transcript is usually translated into a 169 amino acid polypeptide named p19ARF (19kDa). Human and mouse proteins share only 50% of identity. Interestingly, the exon 1-encoded N-terminal region, that is necessary and sufficient to fulfil almost all of the known ARF tumor suppressor functions, is only modestly conserved between species, whereas the exon 2-encoded C-terminal region shows a stronger degree of identity between human and mouse (57% of identity) . By comparison, mouse and human INK4a are more conserved, sharing the 65% of identity overall . ARF proteins are highly basic ( 20% arginine content) and hydrophobic molecules. The basic nature of ARF renders this protein highly insoluble and this is likely the reason for which neither NMR (nuclear magnetic resonance) nor crystal structure Rabbit polyclonal to ZNF182 has been decided, despite its small size. ARF probably needs to form complexes with other ADL5747 molecules to presume specific spatial conformation and to neutralize its charge at physiological pH, thus explaining the incredible quantity of ARF binding partners . ARF is usually a potent tumor suppressor, regulating cell cycle arrest and/or apoptosis by both p53-dependent and impartial pathways . Interestingly, it represents a link between the pRb ADL5747 and the p53 pathway,.